title: TEAD1 condensates are transcriptionally inactive storage sites on the pericentromeric heterochromatin

city: Baltimore

first: Danfeng

last: Cai

address: 615 N Wolfe St

institute: Johns Hopkins Bloomberg School of Public Health

summary: TEA domain transcription factor 1 (TEAD1), a Hippo pathway transcription factor important in cellular homeostasis and development, is increasingly implicated in cancer biology. Here, we reveal a novel role for TEAD1 in organizing nuclear condensates, independent of active transcription. Using high-resolution imaging, ChIP-seq, RNA-seq and proximity-based proteomics, we demonstrate that in patient-derived renal cell carcinoma cells, TEAD1 forms micron-sized foci by binding to the heterochromatic pericentromeric regions using its DNA-binding domain. These TEAD1 foci do not mediate transcription but instead serve as depots for excess TEAD1. This contrasts with TEAD1 organization in other genomic regions of both RCC and normal kidney cells, where TEAD1 associates with markers of active transcription. Our findings provide a mechanistic framework for TEAD1’s dual regulatory roles, offering new insights into its contribution to transcriptional dysregulation and tumor progression.

overall_design:
Chromatin immunoprecipitation (Ch-IP) was performed based on previous studies81. In general, 108 cells are used per each ChIP-seq experiment. HK2 and UOK cells were first treated with 1% formaldehyde in UOK media, followed by the addition of 0.125M glycine to halt cross-linking. The cells, washed in ice-cold PBS, were then lysed in cell lysis buffer, with subsequent nutation at 4°C. Cells were treated by dounce homogenization on ice, followed by centrifuge at 500 x g for 10min. The supernatant was discarded, and the pellets was resuspended using nuclear lysis buffer. Nuclear lysates were then sheared by Covaris E220 to achieve average chromatin length of 300-400 bp. The protein/chromatin complexes are captured using antibodies specific to the TEAD1, H3K9me3, YAP1, H3K27ac, and mouse IgG, and the mixtures were incubated overnight on nutator at 4°C.

Please note that the records have been updated with the following files on April 28, 2026:
GSM9011941_HK2-TEAD1-rep1_S8.k50.bw
GSM9011942_HK2-TEAD1-rep2_S24.k50.bw
GSM9011950_UOK121-TEAD1-rep1_S5.k50.bw
GSM9011951_UOK121-TEAD1-rep2_S27.k50.bw
GSM9011958_UOK122-TEAD1-rep1_S6.k50.bw
GSM9011959_UOK122-TEAD1-rep2_S16.k50.bw
GSM9011967_UOK342-TEAD1-rep1_S7.k50.bw
GSM9011968_UOK342-TEAD1-rep2_S19.k50.bw


title: IL-9 and Blimp-1 protect the transcriptional identity of group 2 innate lymphocytes in allergic asthma [ATAC-seq]

status: Public on May 07 2025

city: Pittsburgh

first: Amanda

middle: Catherine

last: Poholek

state: Pennsylvania

address: 4401 Penn Ave., Rangos Research Center 8129

department: Pediatrics

country: USA

institute: University of Pittsburgh

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Type 2 inflammation in the lung underlies allergic asthma and promotes tumor metastasis. Type 2 innate lymphoid cells (ILC2s) respond to tissue damage signals including IL-33 and IL-25 and are implicated in type 2 inflammatory diseases of the lung, but the factors that maintain ILC2s ability to produce type 2 cytokines are not known. We show Blimp-1 is a key transcriptional repressor of type 1 genes induced by the type 2 cytokine IL-9 in ILC2s in response to tissue damage signals. Loss of Blimp-1 in ILC2s altered inflammation in response to allergens, but also limited metastatic melanoma in the lung by limiting type 2 cytokines in favor of type I cytokines including IFNg and TNF. Thus, Blimp-1 protects the type 2 identity of ILC2s during lung inflammatory diseases.

overall_design: 2 female Blimp-1IL7RaCre and 2 female ControlIL7RaCre mice match with age were used. Whole lung was harvested and ILC2s were sorted from lung by the following markers: live, lineage-, CD90.2+, CD3-. After sorting, ~15,000 ILC2s cells from each sample were cultured in complete RPMI in 96-well plate with 20ng/mL IL-33 and 20ng/mL IL-25 for 72 hours. After 72 hours, ILC2s were collected passed down for sequencing.

first: Yibo

last: Zheng

first: Aaron

last: Yang

middle: C


title: Generation of a biliary tract cancer cell line atlas identifies molecular subtypes and therapeutic targets

status: Public on Apr 30 2025

city: Boston

first: Vindhya

last: Vijay

state: MA

address: 185 Cambridge Street

institute: MGH Cancer Center/ Harvard Medical School

type: Expression profiling by high throughput sequencing

summary: Biliary tract cancers (BTCs) are aggressive malignancies encompassing intrahepatic and extrahepatic cholangiocarcinoma, gallbladder carcinoma, and ampullary carcinoma. Here, we report integrative analysis of 63 BTC cell lines via multi-omics and genome-scale CRISPR screens. We identify widespread EGFR dependency in BTC, alongside dependencies selective to anatomic subtypes. Additionally, we delineate strategies to overcome therapeutic resistance, with combined EGFR inhibition potentiating targeting of KRAS-mutant and FGFR2-fusion-driven models, and SHP2 inhibition effective in the latter context. Clustering RNA/protein expression and dependencies data revealed functional relationships transcending single-gene alterations, with biliary, squamous, or dual biliary/hepatocyte lineage signatures stratifying BTC models. These subtypes exhibit distinct dependency profiles— including cell fate transcription factors GRHL2, TP63, and HNF1B, respectively— and demonstrate prognostic significance in patient samples. Potential subtype-specific targetable vulnerabilities include Integrin-a3 and the detoxification enzyme UXS1. This cell line atlas reveals therapeutic targets in molecularly-defined BTCs, unveils disease subtypes, and provides a resource for therapeutic development.

overall_design: RNASeq profiling of newly developed and established patient-derived biliary tract cancer cell lines.

first: Vijay

last: Vindhya

first: Karisan

last: Negin

first: Vazquez

last: Francisca

first: Getz

last: Gad

first: Bardeesy

last: Nabeel


title: IL-9 and Blimp-1 protect the transcriptional identity of group 2 innate lymphocytes in allergic asthma [RNA-seq]

overall_design: For bulk RNA sequencing evaluating Blimp-1 deficiency (including library WT_1, WT_2, WT_3, KO_1, KO_2 and KO_3), 3 female Blimp-1IL7RaCre and 3 female ControlIL7RaCre mice match with age were used. Whole lung was harvested and ILC2s were sorted from these individual mice by the following markers: live, lineage-, CD90.2+, CD3-. After sorting, ~15,000 ILC2s cells from each sample were cultured in complete RPMI in 96-well plate with 20ng/mL IL-33 and 20ng/mL IL-25 for 72 hours. After 72 hours, ILC2s were collected and RNA was isolated with RNeasy Plus Micro Kit (74034, Qiagen). RNA quality was assessed by RIN score and passed down to library preparation.  For bulk RNA sequencing comparing unstimulated ILC2s ex vivo from lung with stimulated ILC2s (including library 1_unstim, 2_unstim, 3_unstim, 4_unstim, 1_stim, 2_stim, 3_stim and 4_stim), 8 female B6 mice were separated into two groups. Whole lung was harvested and ILC2s were sorted from these individual mice lung by the following markers: live, lineage-, CD90.2+, CD3-. After sorting, ~15,000 ILC2s cells from each sample were cultured in complete RPMI in 96-well plate with or without IL-33 (20ng/mL) according to their group for 24 hours. After 24 hours, ILC2s were collected and RNA was isolated with RNeasy Plus Micro Kit (74034, Qiagen). RNA quality was assessed by RIN score and passed down to library preparation.


title: Diet-induced obesity dysregulates rapid epigenetic response to hypoxia in macrophages (RNA-Seq)

status: Public on May 05 2025

city: Syracuse

first: Norifumi

last: Urao

state: New York

address: 750 E. Adams St.

department: Pharmacology

institute: SUNY Upstate Medical University

summary: Macrophages play important roles in tissue homeostasis and tissue healing process. The response to hypoxic environment is a trigger to induce key signaling pathways in macrophages. Functions of macrophages are dysregulated in metabolic diseases such as diabetes and obesity resulting in poor tissue healing. Epigenetic status including histone modification in macrophages is regulated by hypoxia. However, whether and how obesity influences histone modification under hypoxia is unknown. Using chromatin immunoprecipitation-sequencing (ChIP-seq), RNA-seq on mouse bone marrow-derived macrophages, we sought to determine whether hypoxia alters histone 3-lysine 4 trimethylation (H3K4me3) and gene expression profile in macrophages and how obesity influences H3K4me3 enrichment on the genome in hypoxia-dependent and -independent manner.

overall_design: Gene expression profile analysis of RNA-seq data for bone marrow-derived macrophages (BMDMs) cultured from C57B6/J mice fed a high-fat diet or a normal-fat diet.

first: Kentaro

last: Takahashi


title: CaMKK2 regulates mechanically-induced macrophage fate

city: Madison

first: Jianhong

last: Ou

state: Wisconsin

address: 330 N Orchard Street

institute: Morgridge Institute For Research

summary: Macrophages play crucial roles in immunity and tissue homeostasis, exhibiting diverse phenotypes influenced by their microenvironment. While matrix stiffness is recognized as a key regulator of macrophage fate, the underlying mechanism remains elusive.  Here, we investigate primary bone marrow-derived macrophages (BMDMs) in a physiologically relevant 3D environment using a dynamic stiffening collagen-based hydrogel system. We show that increasing matrix stiffness promotes BMDM polarization towards a pro-regenerative phenotype, characterized by elevated CD206 levels, and M2-like gene signature. Mechanistically, we identify the calcium/calmodulin-dependent kinase kinase 2 (CaMKK2) as an important component of molecular machinery controlling BMDM response to stiff microenvironment. Surprisingly, the blocking of CaMKK2 does not interfere with the IL-4-induced pro-regenerative programs. On the contrary, the inhibition of this kinase abolishes phenotypic and transcriptional changes induced by stiffness, highlighting the specific function of the CaMKK2-STAT6 axis in macrophage mechanosensory signaling. Functionally, matrix stiffening promotes M2-like/pro-tumoral polarization of BMDM in a CaMKK2-dependent manner, confirming the critical role of this kinase in the tumor microenvironment and identifying a direct pro-tumoral effect mediated by Camkk2-signaling in macrophages. In a murine wound healing model, stiffened matrices drive immune cell recruitment and macrophage polarization towards a pro-regenerative phenotype, facilitating vascularization, while deletion of Camkk2 impairs this phenomenon. Our findings elucidate the mechanistic basis of matrix stiffness-mediated macrophage fate determination, identifying CaMKK2 as a druggable target to selectively modulate the mechanoresponsiveness of macrophages in malignant and injured tissues.

overall_design: To investigate the effect of matrix stiffness on the regulation of macrophages activation programs, we leveraged 3D dynamics hydrogels cultures. Immunoprofiling, transcriptomics, and phospho-proteins analysis were used to determine the effects of matrix stiffness on BMDM. WT and Camkk2-/- BMDM were used to determine the role on CaMKK2 in mechanosensory signaling.

first: Ya

last: Guan

first: Min

last: Zhang

first: Luigi

last: Racioppi

first: Sharon

last: Gerecht


title: UHRF1 regulates AR ubiquitination to promote the loss of AR Signaling and Enzalutamide Resistance in Progression of Prostate Cancer

status: Public on Apr 29 2025

city: Guangzhou

first: Yifan

state: Guangdong

address: Department of Urology,Nanfang Hospital,Southern Medical University,Guangzhou,Guangdong,China

country: China

summary: In this study, we demonstrated the critical role of the epigenetic regulator UHRF1 in the enzalutamide resistance development of prostate cancer. We have shown that UHRF1 is highly expressed in enzalutamide-resistant prostate cancer cells and its expression correlates with the loss of androgen receptor (AR)-dependent glandular features. Knocking down UHRF1 led to increased AR expression and enhanced the activity of canonical AR signaling pathway in prostate cancer cells.

overall_design: Expression profiling by high throughput sequencing.


title: HIF transcriptome in clear cell renal carcinoma cells

status: Public on Apr 24 2025

city: Dallas

first: Weibo

last: Luo

state: Texas

address: 5323 Harry Hines Blvd.

department: Pathology

institute: UT Southwestern Medical Center

summary: Global gene changes by HIF-1/2alpha loss or VHL expression in cancer cells

overall_design: Parental, HIF-1/2alpha double knockout, VHL reconsituted RCC10 cells were subjected to mRNA-seq.

first: Yong

last: Wang


title: HIF-1 ChIP-seq in hypoxic breast cancer cells

summary: The genome-wide HIF-1alpha binding to chromatin under hypoxia

overall_design: MDA-MB-231 cells were exposed to 1% O2 for 24 hrs and subjected to HIF-1alpha ChIP-seq.

first: Lei

last: Bao


title: Transcriptome induced by DNMT degrader in ccRCC cells

summary: Global gene changes by DNMT degrader SGI1027 in VHL-deficient ccRCC cells

overall_design: RCC10 cells were treated with vehicle or SGI1027 for 2 days and subjected to mRNA-seq.


title: LSD1 inhibition attenuates targeted therapy-induced lineage plasticity in BRAFV600E colorectal cancer

city: Bloomington

first: Heather

last: O'Hagan

state: IN

address: 1001 E. 3rd St. Bloomington, IN 47405

institute: Indiana University

summary: Background: BRAF activating mutations occur in approximately 10% of metastatic colorectal cancer (CRCs) and are associated with worse prognosis in part due to an inferior response to standard chemotherapy. Standard of care for patients with refractory metastatic BRAFV600E CRC is treatment with BRAF and EGFR inhibitors and recent FDA approval was given to use these inhibitors in combination with chemotherapy for patients with treatment naïve metastatic BRAFV600E CRC. Lineage plasticity to neuroendocrine cancer is an emerging mechanism of targeted therapy resistance in several cancer types. Enteroendocrine cells (EECs), the neuroendocrine cell of the intestine, are uniquely present in BRAFV600E CRC as compared to BRAF wildtype CRC. Methods: BRAF plus EGFR inhibitor treatment induced changes in cell composition were determined by gene expression, imaging and single cell approaches in multiple models of BRAF mutant CRC. Furthermore, multiple clinically relevant inhibitors of the lysine demethylase LSD1 were tested to determine which inhibitor blocked the changes in cell composition. Results: Combined BRAF and EGFR inhibition enriched for EECs in all BRAF mutant CRC models tested. Additionally, EECs and other secretory cell types were enriched in a subset of BRAFV600E CRC patient samples following targeted therapy. Importantly, inhibition of LSD1 with a clinically relevant inhibitor attenuated targeted therapy-induced EEC enrichment through blocking the interaction of LSD1, CoREST2 and STAT3. Conclusions: Our findings that BRAF plus EGFR inhibition induces lineage plasticity in BRAFV600E CRC represents a new paradigm for how resistance to BRAF plus EGFR inhibition occurs. Additionally, our finding that LSD1 inhibition blocks lineage plasticity has the potential to improve responses to BRAF plus EGFR inhibitor therapy in patients.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Patient derived xenograft colon organoids were emplanted into the colons of NSG mice and tumors were allowed to grow. Resulting tumors were then treated with vehicle or a combination of encorafenib and cetuximab for three weeks and 10X Flex single cell RNA sequencing was performed. For the mouse syngenic model, TPKO organoids were emplanted into the colons of wild type C57Bl/6 mice and tumors were allowed to form. Tumors were then treated for 3 weeks with vehicle or a combination of encorafenib and gefitinib for three weeks and 10X Flex single cell RNA sequencing was performed

first: Christopher

middle: A

last: Ladaika

first: Averi

last: Chakraborty

first: Ashiq

last: Masood

first: Galen

last: Hostetter

first: Joo Mi

last: Yi

middle: M

last: O’Hagan


title: Transcription factor NFYA directs male meiotic entry by facilitating accessible chromatin at the promoters of genes expressed during meiosis [scRNA-seq & scATAC-seq]

status: Public on May 10 2025

city: Stockholm

first: Masomeh

last: Askari

address: Svante Arrhenius väg 20C

country: Sweden

institute: Stockholm University

summary: Meiotic prophase I characterized by homologous recombination and synapsis is an intricate step for spermatogenesis. This process entails extensive changes to chromatin and transcription. Recent studies have revealed that prior to prophase I, accessible chromatin bound by paused Pol II at meiotic gene promoters is essential for their timely activation during later stages of prophase I. However, the factor responsible for promoting accessible chromatin at meiotic gene promoters before entry into prophase I is unknown. Here, we discovered that NFYA expressed in pre-meiotic germ cells promotes accessible chromatin at meiotic gene promoters. Concordantly, conditional germline deletion of Nfya in male mice blocks meiotic entry. Functionally, our spatial and single-cell ATAC-seq data revealed that loss of NFYA in pre-meiotic cells disrupts accessible chromatin at meiotic gene promoters. Our study identifies a pioneer role for NFYA in facilitating permissive chromatin at meiotic gene promoters before meiosis, thereby controlling the timely activation of meiotic genetic program during later meiosis.

overall_design: We performed single-cell RNA sequencing (scRNA-seq) and single-cell ATAC sequencing (scATAC-seq) on adult Testis tissue to simultaneously profile the whole transcriptome and assess genome-wide chromatin accessibility at single-cell resolution.

first: Martin

last: Säflund

first: Atiyeh

last: Eghbali

first: Mukhtar

last: Mohammed Abdi

first: Ann-Kristin Iréne

last: Östlund Farrants

first: Tianxiong

last: Yu

first: Deniz

last: Özata


title: Trpv4 specifically inhibits p53-driven Dkk1 and Sost secretion in a novel Kcnk2+ osteolineage subpopulation to rejuvenate the aged osteo-brain axis

status: Public on Apr 23 2025

city: Shanghai

first: Kai

last: Yang

address: 197 Ruijin 2nd Road, Shanghai, 200025, China

department: Shanghai Institute of Traumatology and Orthopedics

institute: Shanghai Jiao Tong University School of Medicine

summary: This comprehensive multi-omics study aims to elucidate the multi-dimensional transcriptional and epigenetic mechanisms underlying osteogenic lineage fate remodeling and bone homeostasis during aging and exercise. The existing samples encompass bulk RNA-sequencing (RNA-seq) to evaluate global transcriptomic alterations in brain and bone tissues. The new samples include single-cell RNA sequencing (scRNA-seq) of an 8-month-old (8M) mouse model to map osteogenic heterogeneity at high resolution, alongside CUT&Tag profiling to delineate the direct genome-wide binding landscapes of Trp53.

overall_design:
Bulk RNA-sequencing (existing, n=18 samples): Long bones were first digested with Collagenase A to remove the periosteum. The bones were then sagittally split, and the bone marrow was completely removed via extensive flushing. The remaining cleaned bone fragments were subjected to total mRNA isolation and subsequent bulk RNA-seq analysis to evaluate systemic transcriptomic changes.
Single-cell RNA-sequencing (new, n=1 sample): Following the initial preparation steps, Collagenase A digestion to remove the periosteum, sagittal splitting, and thorough flushing of the bone marrow, the remaining long bone fragments from an 8-month-old (8M) mouse were minced into fine pieces. The tissue was then digested with EDTA and Collagenase A for total 20 minutes with continuous agitation at 37°C. The resulting cell suspension was filtered through a 20-μm cell strainer to generate a high-quality, non-FACS-sorted single-cell suspension for library preparation.
CUT&Tag profiling (new, n=3 samples): Differentiating IDG-SW3 cells were treated with Nutlin-3a for 4 hours prior to harvest. The treated cells were then subjected to the CUT&Tag protocol to profile the direct genome-wide chromatin binding landscape of Trp53.

first: Rong

last: Zhou

first: Wei

last: Yan

first: Yuxin

last: Zhang

first: Jiahui

first: Yichen

last: Zhao

first: Ruonan

last: Ning

first: Min

last: Jiang

first: Lianfu

last: Deng


title: Pbrm1 loss induces a permissive chromatin state for cholangiocytic differentiation and cholangiocarcinoma formation [ATAC-seq]

status: Public on Apr 21 2025

city: Taipei City

first: Li-Wen

last: Chiou

address: Chung Shan S. Rd.

country: Taiwan

institute: National Taiwan University

summary:
Background and Aims:
The SWI/SNF ATP-dependent chromatin remodeling complex regulates transcrip-tional machinery access and is critical in normal physiology and cancer development. Pbrm1, a key subunit of this complex, is frequently mutated in intrahepatic cholan-giocarcinoma (ICC), though how its loss contributes to tumorigenesis remains poor-ly understood. This study aims to explore the role of Pbrm1 in liver physiology and its involvement in ICC development.
Methods:
Liver-specific Pbrm1 knockout (Pbrm1 KO) mice were generated to assess the ef-fects of Pbrm1 loss under various conditions. These mice were exposed to a 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet to induce cholestatic injury and were also subjected to a high-fat diet to evaluate susceptibility to liver steatosis. Chromatin accessibility and gene expression under both normal and injury condi-tions were examined. Additionally, the impact of Pbrm1 loss was analyzed in com-bination with an activating KrasG12D mutation to study cancer development.
Results:
Pbrm1 KO mice exhibited increased susceptibility to cholestatic injury, with an en-hanced ductular reaction. Loss of Pbrm1 reduced chromatin accessibility at hepato-cyte-specific and metabolically important genes, though RNA expression remained unaffected during homeostasis. Following cholestatic injury, hepatocyte-specific gene expression was significantly reduced compared to wild-type controls. Pbrm1 KO mice also showed heightened vulnerability to high-fat diet-induced liver steato-sis. When combined with KrasG12D mutation, Pbrm1 KO/KrasG12D mice had shorter survival and were more likely to develop cholangiocarcinomas, whereas Pbrm1 wild type/KrasG12D mice predominantly developed hepatocellular neoplasms.
Conclusion:
Pbrm1 maintains chromatin accessibility for hepatocyte differentiation-related genes. Its loss promotes differentiation toward cholangiocytes during injury or tu-morigenesis, driving ICC development.

overall_design: To investigate the role of Pbrm1 in liver function and tumorigenesis, we generated Alb-Cre-mediated Pbrm1 knockout mice. We performed ATAC-seq to examined chromatin accessibility changes between Pbrm1 KO and WT liver.

first: Liwen

first: Yulin

last: Jhuang

first: Chialang

last: Hsu

first: Yungming

last: Jeng


title: Therapeutic base editing alleviates restrictive cardiomyopathy

city: Xihu District, Hangzhou

first: Xing

last: Chang

state: Zhejiang

address: No.18 Shilongshan Road Cloud Town

department: School of Life Sciences

institute: Westlake University

summary: Restrictive cardiomyopathy (RCM) is a severe cardiac disorder characterized by impaired ventricular filling and diastolic dysfunction, with mutations in sarcomeric proteins representing major causative factors. Mutations of TNNI3 gene (e.g. p.R192H) constitute major genetic causes of RCM, particularly affecting pediatric patients and being associated with poor prognosis. Here, we demonstrate that adenine base editor (ABE) is able effectively correct RCM-causing mutation and alleviate RCM in a murine model. We first developed a novel murine model harboring the Tnni3R193H mutation that recapitulates the hallmark features of human RCM. Importantly, targeted delivery of ABE via adeno-associated virus (AAV) achieved efficient and precise correction of the Tnni3R193H mutation in adult RCM mice, leading to significant improvement of cardiac functions. Our findings establish base editing as a therapeutic strategy for RCM and highlight its broader potential for treating genetic cardiomyopathies in clinical settings.

overall_design: RNA sequencing (RNA-seq) was employed to identify RNA off-target effects induced by ABE8e treatment in vivo. Untreated mice were included as a control group to serve as mock treatment for comparison, ensuring a baseline for evaluating the specificity and potential off-target impacts of the ABE8e intervention

first: Chong

first: Xiulin

first: Geng

last: Li


title: RNA sequencing to study the molecular relationships between temperature, aging, and OCTR-1

city: Spokane

first: Yiyong

last: Liu

state: WA

address: 412 E Spokane Falls Blvd

institute: Washington State University

summary: To gain molecular insights into OCTR-1-dependent longevity responses to temperatures, we employed RNA-seq to compare gene expression in young and old wild-type and octr-1(ok371) animals propagated at different temperatures (15°C, 20°C and 25°C).

overall_design: Five replicates of eight groups of RNA samples (young and old wild-type and octr-1(ok371) animals propagated at 15°C, 20°C or 25°C) were collected and subjected to RNA-seq analysis.

first: Phillip

last: Wibisono

first: Jingru

last: Sun


title: Human CLOCK enhances cognitive flexibility by altering cortical excitatory neuron function

status: Public on Apr 18 2025

city: Los Angeles

first: Genevieve

last: Konopka

state: CA

address: 650 Charles Young Drive South, 73-214 CHS

department: Neurobiology

institute: UCLA David Geffen School of Medicine

summary: A BAC containing human CLOCK gene with flanking regulatory region was transferred to mice. The humazied  (HU) mice outperformed wildtype (WT) in a reversal learning task, and hence suggested enhanced cognitive flexibility in HU mice. IHC of frontal cortex suggested that the enhanced cognitive flexibility in HU mice might result from the increased neuronal density, dendritic arborization, and spine density. In order to determine the molecular mechanism, we conducted single-nuclei RNA-seq on frontal cortex from postnatal 7 day (P7) and P56 mice. In the excitatory neuron, genes that upregulated in HU mice were enriched for  dendritic growth (e.g. Tenm2 and Flrt2), spine formation (e.g. Cntn5 and Sorcs2), and energy metabolism (e.g. Suclg1 and Cox8a).

overall_design: P7 and P56 of humanized (CLOCKhum;Clock-/-, HU), knockout (Clock-/-, KO), and wildtype (Clock+/+, WT) mice were harvested rostral part of frontal cortex which is corresponding to the cortex part of Coronal level 22-36 in Allen Brain Atlas: Reference Atlas, Version 2 (2011), and extracted single nuclei for sequencing.

first: Joseph

middle: S

last: Takahashi

first: Yuxiang

first: Ashwinikumar

last: Kulkarni

first: Miles

last: Fontenot


title: T-cell receptor repertoire alterations drive pathological gut immunological and microbial signatures in humans and mice with hypomorphic RAG deficiency

status: Public on Apr 09 2025

city: BETHESDA

first: Cihan

last: Oguz

state: MD

address: NIHBC 29B BG RM 1NN20, 9000 ROCKVILLE PIKE

institute: NIH

summary: Mechanistic knowledge about the homeostatic interplay between T cell-mediated immunity and gut microbiota is still limited. We aimed to investigate how restriction of the T-cell receptor (TCR) repertoire (VDJ region) may influence gut microbiota composition, causing tissue inflammation and disruption of peripheral immune tolerance. Based on the clinical observation of colonic inflammation in patients with partial RAG deficiency (pRD), we applied a multi-omics strategy to study a mouse model of hypomorphic Rag1 deficiency (Rag1 mutant mice) with spontaneous development of colitis.

overall_design: To characterize the transcriptomic and immune repertoire differences between the wild-type and Rag1 mutant mice at 3 weeks and 12 weeks of age, with and without ANMV (ampicillin, neomycin, metronidazole and vancomycin) treatment (only at 12 weeks for the Rag1 mutant mice), we performed scRNA-seq and TCR repertoire (VDJ region) sequencing of colonic lamina propria-specific CD4+ T cells. The scRNA-seq and TCR libraries were coupled with hashtag oligo (HTO) libraries for labeling the samples in order to demultiplex the pooled libraries into wild-type and Rag1 mutant mice at 3/12 weeks of age and with/without ANMV treatment cell populations for downstream analysis. The number of libraries sequenced is six for scRNA-seq, six for TCR (VDJ) repertoire, and six for HTOs.

first: Riccardo

last: Castagnoli

first: Francesca

last: Pala

first: Poorani

last: Subramanian

first: Benjamin

last: Schwarz

first: Ai Ing

last: Lim

first: Andrew

last: Burns

first: Elena

last: Fontana

first: Marita

last: Bosticardo

first: Cristina

last: Corsino

first: Angelina

last: Angelova

first: Ottavia

last: Delmonte

first: Heather

last: Kenney

first: Deanna

last: Riley

first: Grace

last: Smith

first: Lisa

last: Ott de Bruin

first: Vasileios

last: Oikonomou

first: Lucas

last: Dos Santos Dias

first: Danielle

last: Fink

first: Eric

last: Bohrnsen

first: Cole

middle: D

last: Kimzey

first: Gian Luigi

last: Marseglia

first: Guisela

last: Alva-Lozada

first: Jenna

last: Bergerson

first: Ana

last: Brett

first: Karlla

last: W Brigatti

first: Dimana

last: Dimitrova

first: Cullen

last: Dutmer

first: Alexandra

middle: F

last: Freeman

first: Ale

last: Hanadys

first: Steven

last: Holland

first: Francesco

last: Licciardi

first: Srdjan

last: Pasic

first: Laura

last: Poskitt

first: David

middle: E

last: Potts

last: Dasso

first: Svetlana

middle: O

last: Sharapova

first: Kevin

last: Strauss

first: Brant

last: Ward

first: Melis

last: Yilmaz

first: Douglas

middle: B

last: Kuhns

first: Michail


title: Transcription factor reveals interconnected regulation of carbon source utilization and carbon catabolite repression in an oleaginous yeast

status: Public on Apr 10 2025

city: Ithaca

first: Lori

last: Huberman

state: NY

address: 236 Tower Rd.

department: Plant Pathology and Plant-Microbe Biology, School of Integrative Plant Science

institute: Cornell University

summary: Cells must sense and respond to available nutrients to utilize available resources and establish fungal colonies. Saprophytic microbes harvest carbon from plant biomass, which contain insoluble, complex carbohydrates that must be degraded extracellularly prior to import into the cell. Filamentous fungi can degrade these complex carbohydrates. However, while many single cellular microbes can utilize the building blocks of these insoluble polysaccharides, they frequently are unable to degrade insoluble carbohydrates. Cellobiose is a disaccharide breakdown product of cellulose, the most abundant component of plant biomass. We investigated the genetic regulation of cellobiose utilization in the oleaginous, basidiomycete yeast Rhodotorula (Rhodosporidium) toruloides using a combination of phylogenetic, genetic, and genomic approaches. We tested the closest R. toruloides homolog of the cellulose degradation regulator CLR-2/ClrB for a role in cellobiose utilization. Deletion of this transcription factor, which we named Cbr1, eliminated the ability of cells to grow on media containing cellobiose as the sole carbon source. Although, the role of CLR-2/ClrB is limited to regulating the expression of genes involved in the utilization of cellulose and hemicellulose, Cbr1 plays a role not only in cellobiose utilization, but also utilization of tricarboxylic acid (TCA) cycle intermediates. Transcriptional profiling revealed that the genes activated by Cbr1 included genes encoding beta-glucosidases and transporters. These beta-glucosidases are secreted and are necessary and sufficient for cellobiose utilization, while one of the transporters is required for utilization of the TCA cycle intermediate, citrate. Additionally, Cbr1 inhibits carbon catabolite repression, specifically during utilization of disaccharides, which may have evolved to limit the repression of genes encoding secreted proteins that cleave disaccharides into glucose extracellularly. The co-regulation of cellobiose and TCA cycle utilization with carbon catabolite repression may shed light on alternative methods of the regulation of carbon catabolite repression in fungi.

overall_design: mRNA-seq profling of Rhodotorula toruloides wild type and cbr1 deletion strains exposed to cellobiose or succinate as the sole carbon source or media lacking a carbon source for 8h. Each experiment was performed in biological triplicate.

first: Brandon

last: Reyes-Chavez

first: Joshua

last: Kerkaert


title: Endometrial assembloid model reveals endometrial adenogenesis regulation by estradiol-driven WNT7B expression [bulk RNA-seq]

status: Public on Apr 08 2025

city: Hong Kong

first: Yimeng

last: Li

address: 21 Sassoon Road, Pokfulam Hong Kong

institute: The university of Hong Kong

summary: Little is known about the adenogenesis of human endometrial glands. This study explored endometrial adenogenesis using a novel three-dimensional endometrial assembloid model that integrates human endometrial organoids (EOs) and human endometrial stromal cells (HESCs). The model effectively recapitulated endometrial tubular gland formation, underscoring the critical role of stromal-epithelial interactions. Transcriptomic analyses identified WNT7B as a key intrinsic regulator for EO-derived tubular gland formation, which is extrinsically regulated by TGFβ1-VDR interaction between HESCs and EOs. Estradiol stimulated endometrial gland development via WNT7B downregulation in EOs. This finding was validated in an estradiol-stimulated mouse model and clinical samples from women undergoing in vitro fertilization (IVF) cycles. Uterine-specific WNT7B knockout in mice further confirmed its inhibitory role in endometrial gland development. This study offers insights into endometrial adenogenesis and potential therapeutic targets for related endometrial disorders.

overall_design: As differential tubular gland development ability has been identified in different endometrial organoid samples, we tried to identify the differential gene expression patterns in EOs with and without tubular gland development capacity. EOs were collected separately that with and without tube development potential for RNA sequencing analysis.

first: Xintong

first: Jianlin

first: Cheuk

middle: L

last: Lee

first: Philip

last: Chiu


title: E-cigarette Vapor Extract Alters Human Eosinophil Gene Expression in an Effect Mediated by Propylene glycol, Glycerin, and Nicotine

city: San Diego

first: Gregory

last: Seumois

address: 9420 Athena Cir, La Jolla

institute: La Jolla Institute for immunology

summary: E-cigarette use (vaping) has become widespread, and its effects on airway inflammation and disease are not fully delineated. E-cigarette vapor extract (EVE) profoundly affects neutrophil function. We hypothesized that EVE also alters eosinophil function and thus could impact allergic airways disease. We employed RNA-sequencing to measure the ex vivo effect of EVE components on human eosinophil transcription. Eosinophils from 7 non-vaping subjects without asthma were isolated by negative selection from venous blood. Cells were incubated for 48 hours with EVE consisting of glycerin, propylene glycol and nicotine (EVE+), EVE without nicotine (“EVE-”), Extract Buffer (EB), and untreated media. Bulk RNA-sequencing was performed. Principal component analysis revealed that the EVE+ and EVE- conditions clustered closely together. Differential gene expression analysis revealed a “negative regulation of interferon gamma production” and “apoptotic mitochondrial changes” pathways for genes upregulated in EVE- samples and a down regulation of  “inflammatory response” and “chemokine signaling” in comparison to the EB control condition. 93 genes were differentially expressed genes between EVE+ and EVE-, indicating a relatively small effect of nicotine on eosinophil gene expression. These results demonstrate that EVE significantly alters eosinophil gene expression, predominantly mediated by propylene glycol and glycerin rather than by nicotine. This study motivates further research into the pathogenic effects of vaping on airway eosinophils and allergic airways disease.

overall_design: Circulating Eosinophils from 7 healthy subjects non-smokers are isolated and in vitro submitted to different E-cigarette vaping compounds including PEG, Glycerin and Nicotine. After 48 hours of treatment, cells are harvested and totRNA is extracted and sequencing libraries are prepared for whole trancriptome expression analysis.

first: Nicholas

middle: T

last: Hogan

first: F

last: Castaneda-Castro

first: Howard

last: Brickner

first: Ashmitaa

last: Ramamoorthy Premlal

first: Monalisa

last: Mondal

first: Sara

last: Herrera-De La Mata

first: Pandurangan

last: Vijayanand

last: Crotty Alexander

first: Praveen

last: Akuthota


title: Foxo1 regulates intestinal tissue-resident memory CD8 T cell biology in an anatomic compartment- and context-specific manner

status: Public on Apr 11 2025

first: John

last: Chang

address: UCSD

institute: UC San Diego

summary: Transcriptional regulation of diverse aspects of tissue-resident CD8 memory T cells (TRM) biology is nuanced and context-specific. Transcription factors (TF) that orchestrate TRM formation and maintenance are distinct between anatomic compartments, and predicting the roles of TF on TRM biology is challenging. Here, we applied the Taiji algorithm to map TF networks and identify Forkhead Box protein 01 (Foxo1) as differentially invested in intestinal TRM TF networks. Loss of Foxo1 in established TRM resulted in a survival advantage for small intestinal intraepithelial layer (siIEL) TRM compared with other intestinal compartments. SiIEL TRM were uniquely able to maintain expression of proliferative and anti-apoptotic cellular programs in the absence of Foxo1. This advantage was lost upon blockade of integrin αE (Itgae/CD103). These findings suggest that the TF networks maintaining siIEL TRM exhibit redundancy and highlight an underappreciated role of integrin signaling in TRM biology outside of tissue-retention.

overall_design: P14 WT CD45.1 and P14 Foxo1f/f Cre-ERT2+/- CD45.1.2 CD8 T cells were adoptively co-transferred into congenically distinct recipient mice. The recipient mice were infected with 2x105 plaque forming units of lymphocytic choriomeningitis virus 30 minutes after transfer. 21 days post-infection, mice were treated with 0.5mg/g tamoxifen intraperitoneally daily for 5 days, and then sacrificed 2 weeks later. CD8 T cells were isolated from spleen, small intestines, and colon, and incubated in an antibody cocktail containing antibody-derived tags against cell surface markers and fluorescence-conjugated antibodies. The congenically distinct CD8 T cells were separated by fluorescence-activated cell sorting. About 20,000 cells for spleen, siIEL, and siLPL samples were loaded onto Chromium Next GEM Chip M (10x Genomics) and 2,000 cells for cIEL and cLPL were loaded onto Chromium Next Gem Chip G (10x Genomics), and partitioned into Gel Beads. CITE-seq was then performed using previously described protocol (Stoeckius et al., 2017).

first: Paul

last: Hsu

first: Eunice

middle: J

last: Choi

first: William

middle: H

last: Wong

first: Yun

last: Lin

last: Vandenburgh

first: Yi

last: Liu

first: Priscilla

last: Yao

first: Cynthia

last: Indralingam

first: Ananda

middle: W

last: Goldrath

first: Wei

last: Wang


title: Endometrial assembloid model reveals endometrial adenogenesis regulation by estradiol-driven WNT7B expression

first: Yimeng

last: Li

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Pbrm1 loss induces a permissive chromatin state for cholangiocytic differentiation and cholangiocarcinoma formation

first: Li-Wen

last: Chiou

overall_design: To investigate the role of Pbrm1 in liver function and tumorigenesis, we generated Alb-Cre-mediated Pbrm1 knockout mice (referred to as PA in RNA-seq data, whereas wild-type mice referred to as P). Mice were fed a DDC-supplemented diet to initiate liver injury. We then performed RNA-seq in condition with (w/) and without (w/o) DDC to analyze the gene expression differences between Pbrm1 KO and WT mice during liver regeneration.

first: Liwen

first: Yulin

last: Jhuang

first: Chialang

first: Yungming

last: Jeng


title: PROTein Evolution Using Selection (PROTEUS) [Illumina]

status: Public on Apr 03 2025

city: Camperdown

first: G.

middle: Greg

last: Neely

state: NSW

address: Charles Perkins Centre, D17.

country: Australia

institute: University of Sydney

summary: Directed evolution (DE) is a process of mutation and iterative artificial selection to breed biomolecules with new or improved activity. DE platforms are primarily prokaryotic or yeast-based, and stable highly mutagenic mammalian systems have been challenging to establish and apply. To this end, we developed PROTein Evolution Using Selection (PROTEUS), a new platform that uses chimeric virus-like vesicles (VLVs) to enable extended mammalian DE campaigns without loss of system integrity. This platform, consisting of a minimal modified Semliki Forest virus genome controlling expression of the Indiana vesiculovirus G coat protein, is stable and can generate sufficient diversity for DE in mammalian systems. Using PROTEUS, we altered the doxycycline responsiveness of tetracycline-controlled transactivators, generating a more sensitive TetON-4G tool for gene regulation. PROTEUS is also compatible with intracellular nanobody evolution, and we use it to design a novel DNA damage-responsive anti-p53 nanobody. Overall, PROTEUS is a robust, efficient, and stable platform to direct evolution of biomolecules within mammalian cells.

sample_organism:
Homo sapiens
Semliki Forest virus
synthetic construct

overall_design: Long-read nanopore sequencing and short-read Illumina sequencing of VLV transgenes to determine transgene sequence proportions and mutations. Long-read sequencing samples arise from pooled (N=6) VLV samples at each sampling timepoint. Short-read sequencing samples arise from single VLV samples at each sampling timepoint.

first: Alexander

last: Cole

first: Christopher

last: Denes

first: Daniel

last: Hesselson

first: Graham

middle: G


title: PROTein Evolution Using Selection (PROTEUS)

platform_organism:
Homo sapiens
Semliki Forest virus
synthetic construct


title: A Living Organoid Biobank of Crohn’s Disease Patients Reveals Distinct Clinical Correlates of Molecular Subtypes of Disease

status: Public on Apr 02 2025

city: La Jolla, CA 92093

first: Pradipta

last: Ghosh

address: 9500 Gilman Drive (MC 0651)

department: Departments of Medicine and Cellular and Molecular Medicine

institute: University of California San Diego

summary: In this work, Penrose et al. demonstrate the potential of PDOs as predictive tools in Crohn’s disease, that that capture the past, present, and future of disease behavior, thereby advancing PDO-informed precision medicine beyond oncology into complex inflammatory diseases.

overall_design: PDOs were isolated from colonic tissue specimens collected from CD subjects using established crypt isolation protocols as previously summarized [PMID:  39332415]. These intestinal crypts, which contain crypt-base columnar (CBC) cells, were enzymatically dissociated and processed for 3D organoid culture. Organoids and monolayers were lysed using 200 μl of RNA lysis buffer followed by RNA extraction per Zymo Research Quick-RNA MicroPrep Kit instructions (see key resources table). Tissue samples were lysed in TRI-Reagent and RNA was extracted using Zymo Research Direct-zol RNA Miniprep.

first: Saptarshi

last: Sinha

first: Harrison

last: Penrose

first: Brigid

last: Boland

first: Courtney

last: Tindle


title: Electrostatic properties of disordered regions control transcription factor search and pioneer activity [ATAC-Seq]

city: Lausanne

first: Sim

last: Sakong

state: Vaud

address: AI 1133, Station 19

department: SV-IBI

country: Switzerland

institute: EPFL

summary: Transcription factors (TFs) recognize specific target sites within cis-regulatory regions to control gene expression. Prokaryotic TFs combine 1D and 3D diffusion mechanisms to efficiently locate their binding sites. In contrast, the determinants of target search efficiency of eukaryotic TFs within the chromatinized genome are poorly characterized. Here we combine in vivo and in vitro single molecule imaging to dissect the role of charged disordered regions of the mammalian Sox2 and Sox17 TFs in their ability to search for their binding sites. We demonstrate that the DBD flanking region of Sox2 (DFRSox2) dramatically enhances Sox2 search efficiency as compared to the negatively-charged DFRSox17 through distinct mechanisms on naked versus nucleosomal DNA. Enhanced search on naked DNA is primarily mediated by an increase in target recognition rate during 1D sliding. In contrast, target site recognition within nucleosomal DNA is mainly enhanced by increased nonspecific interactions with nucleosomes, facilitating binding to closed chromatin and reinforcing pioneering activity. These findings provide critical insights into the biophysical mechanisms governing TF target search beyond the DBD, and demonstrate how charged disordered regions modulate TF target search on the chromatinized genome.

overall_design: ATAC-seq experiments were performed on CGR8-derived mouse embryonic stem cells induced with doxycycline to express Halo-Sox2, Halo-Sox2a, as well as on CGR8-derived cells lacking doxycycline-inducible transgenes (Ctrl).

first: Beat

last: Fierz

first: David

last: Suter


title: GRHL and PGR control WNT4 expression in the mammary gland via 3D 2 looping of conserved and species-specific enhancers [T47DS RNA-seq]

city: Amsterdam

first: Renee

last: van Amerongen

address: Science Park 904

department: Swammerdam Institute for Life Sciences

country: Netherlands

institute: University of Amsterdam

summary:
This dataset belongs to a study in which we functionally dissect the cis-acting enhancer network and spatiotemporal transcriptional mechanisms that regulate WNT4 expression in the mouse mammary gland and the human breast. As part of these efforts, we identified GRHL as a luminal lineage transcription factor. In follow up work, we studied the interplay between GRHL2 and PGR in the presence and absence of progesterone signaling. This bulk RNAseq dataset allows investigation of the PGR signaling response in human T47DS breast cancer cells in either the presence or absence of GRHL2 gene expression.

The 0, 4 and 24 hours timepoints in T47DS cells were first used and reported in https://doi.org/10.1101/2025.04.11.648333.
The complete dataset, including the GRHL2 knockdown samples, was used and reported in https://doi.org/10.1101/2025.09.01.673508.

overall_design: Bulk RNA sequencing of control (wildtype T47DS) or GRHL2 knockdown (polyclonal shGRHL2) T47DS cells treated with the progesterone analogue R5020 for either 4 or 24 hours.

first: Marleen

last: Aarts

first: Anthonius

last: van Boxtel

first: Renée


title: PROTein Evolution Using Selection (PROTEUS) [Nanopore]

first: G.

overall_design: Long-read nanopore sequencing and short-read Illumina sequencing of VLP transgenes to determine transgene sequence proportions and mutations. Long-read sequencing samples arise from pooled (N=6) VLP samples at each sampling timepoint. Short-read sequencing samples arise from single VLP samples at each sampling timepoint.


title: scRNA-seq based characrization of Cdh16-expression alveolar epithelial progenitor during mouse lung development

status: Public on Apr 01 2025

city: WenZhou

first: Jin

last: Wu

state: ZHejiang

address: Jingguan street 1#

institute: WenZhou Medical University

summary: We have employed a single cell sequencing approach using 10x Genomics scRNAseq to study Cdh16-expression alveolar epithelial progenitor during mouse lung development. Alveolar morphogenesis and regeneration rely on progenitors to replenish alveolar epithelia. Cdh16 positive cells constitute an epithelial progenitor pool capable of differentiating into both AT2 and AT1 cells.In this study, we present the first scRNA-seq analysis on of Cdh16 lineage traced cells during mouse lung alveologenesis, revealing their plasticity with a dynamic differentiation trajetory along with the embryonic stages as well as the postnatal days. Overall, our study elucidates the crucial role of the Cdh16 lineage as alveolar epithelial progenitors in lung development and regeneration.

overall_design: Cdh16CreERT2/wt;R26-tdTomatoflox/wt mice were used for in vivo lineage tracing experiments to label Cdh16Pos cells in embryonic lung tissues.Tamoxifen (Sigma-Aldrich) was administered to timed-pregnant females either via IP injection (0.1 mg/kg body weight) or through diet (0.4 g/kg food) since embryonic day 12.5 (E12.5), with embryonic lungs collected at E16.5, E18.5 and postnatal day 10 (P10). TomatoPos cells were isolated by Fluorescence-activated cellsorting (FACS) and analyzed using scRNA-seq.

first: Jin-San

last: Zhang


title: Mapping and quantifying nascent transcript start sites using TT-TSS-seq

status: Public on Mar 31 2025

city: London

first: Folkert

last: van Werven

address: 1 Midland Road

country: United Kingdom

institute: Francis Crick Institute

summary: Transcription initiation is a highly dynamic and tightly regulated process involving the coordinated action of transcription factors, chromatin remodelers, and RNA polymerase which determine where and when transcription begins. Accurately mapping and quantifying transcription start sites (TSSs) from nascently transcribed RNAs remains a key area of interest, as it provides critical insights into transcription dynamics. Here, we combined transient transcriptome sequencing with transcription start site sequencing (TT-TSS-seq) to accurately map and quantify transcription initiation sites from nascent transcripts. Since transient metabolic labelling yields low-input RNA, we optimized the TSS-seq protocol to enhance sensitivity and accuracy. Specifically, we refined enzymatic reactions for decapping and RNA ligation and incorporated 5' oligonucleotides including unique molecular identifiers (UMIs) and barcodes to enable accurate quantification and sample multiplexing. The TT-TSS-seq approach detected transcription initiation of unstable transcripts, such as enhancer RNAs. Moreover, we identified that a large fraction of genes use multiple transcription initiation sites, yet often produce only a single stable transcript. Overall, TT-TSS-seq provides precise mapping and quantification of transcription initiation sites, offering new insights into transcriptional dynamics and expanding the toolkit for studying gene regulation.

sample_organism:
Saccharomyces cerevisiae
Mus musculus

platform_organism:
Saccharomyces cerevisiae
Mus musculus

overall_design: S288C S. cerevisea, and HM1 mESC cells.

first: Eleanor

last: Elgood Hunt

first: Claudia

last: Vivori


title: Nurr1 orchestrates claustrum development and functionality

city: Berlin

first: Andrew

last: Newman

address: Charitéplatz 1

department: Institute for Cell and Neurobiology

country: Germany

institute: Charité Universitätsmedizin Berlin

summary: Claustrum is the part of the forebrain known to be most widely interconnected tissue with almost all global regions. It is believed to be involved in coordinating multiple cognitive behaviors including consciousness formation. However, little is known about the molecular mechanisms underlying its development and involvement in behavioral control. Here we show that Nurr1 (Nr4a2) is the key transcription factor orchestrating claustral morphogenesis, functional connectivity and thus behaviors. Nurr1 deficient claustral cells aberrantly migrate into insular cortex, shaping claustrum into an abnormal wing-like structure, and ectopically turn on insular cortex genetic program. Accordingly, functional connectivity of the claustrum is not properly formed and relevant behaviors are dysregulated in Nurr1 deficient mice. We show that Nurr1 regulates claustral neuron positioning by suppressing G-protein signaling.

overall_design: P0 anterior claustro-insular parts of mouse cerebral cortices (amygdala, hippocampal and posterior cortical tissues were manually excluded) were dissociated. Approximately 10,000 nuclei were prepared for single-nuclei encapsulation using a 10X single cell controller for each genotype.10x Genomics 3' v3 gene expression (GEX) libraries were prepared and sequenced on a single Novaseq sequencing system. Sex was not discriminated in this project or single cell transcriptomic analysis.

first: Kuo

last: Yan

first: Pauline

last: Lange

first: Susanne

last: Mueller

first: Marco

last: Foddis

first: Stefan Paul

last: Koch

first: Philipp

last: Boehm-Sturm

first: Maron

last: Mantwill

first: Carsten

last: Finke

first: Penghui

last: Deng

first: Melissa

last: Long

first: Victor

last: Tarabykin


title: PTEN mutations impair CSF dynamics and cortical networks by dysregulating periventricular neural progenitors

city: Boston

first: Kris

last: Kahle

address: 55 Fruit Street

institute: MGH

summary: Enlargement of the cerebrospinal fluid (CSF)-filled brain ventricles (ventriculomegaly) is a defining feature of congenital hydrocephalus (CH) and an underrecognized concomitant of autism. Here, we show that de novo mutations in the autism risk gene, PTEN, are among the most frequent monogenic causes of CH and primary ventriculomegaly. Mouse Pten-mutant ventriculomegaly results from aqueductal stenosis due to hyperproliferation of periventricular Nkx2.1+ neuroprogenitor cells (NPCs) and increased CSF production from hyperplastic choroid plexus. Pten-mutant ventriculomegalic cortices exhibit network dysfunction from increased activity of Nkx2.1+ NPC-derived inhibitory interneurons. Raptor deletion or post-natal everolimus treatment corrects ventriculomegaly, rescues cortical deficits, and increases survival by antagonizing mTORC1-dependent Nkx2.1+ NPC pathology. Thus, PTEN mutations concurrently alter CSF dynamics and cortical networks by dysregulating Nkx2.1+ NPCs. These results implicate a non-surgical treatment for CH, demonstrate a genetic association of ventriculomegaly and ASD, and help explain neurodevelopmental phenotypes refractory to CSF shunting in select CH patients.

overall_design: Examine the knockout effect of PTEN on choroid plexus, subventricular zone and cortex

first: Kristopher

first: Emre

last: Kiziltug

first: Tyrone

last: DeSpanza Jr.


title: DAXX governs the silencing of LINE1 during 1  spermatogenesis in mice

city: Wuhan

first: shen

last: jin

state: Hubei

address: No. 299, Bayi Road, Wuchang District, Wuhan City

institute: Wuhan university

type: Methylation profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

summary: The disinhibition of transposon elements (TEs) is a significant threat to male reproduction, particularly during the delicate process of spermatogenesis. Here we identify that Death associated protein 6 (Daxx), a potent transcription repressor and an H3.3 chaperone essential for the TEs silencing during spermatogenesis. DAXX-deficient mouse display delayed meiotic progression, reduced production of spermatids, deformed spermatozoa and age-dependent decline in male fertility. We demonstrate that young long-interspersed nuclear elements (LINE1) and endogenous retroviral elements (ERVs) subfamilies, were upregulated in meiotic spermatocytes of Daxx null testes. Further study found that the Daxx mutant meiotic cells exhibit DNA hypomethylation in TEs. In summary, we have identified DAXX as a previously unknown key regulator of spermatogenesis that may function as an epigenetic regulator to silence young LINE1 by DNA methylation.

overall_design: This submission includes RNA-Seq(3 replicates), smallRNA-Seq(2 replicates), ATAC-seq(3 replicates), and WGBS sequencing(2 replicates) data from testicular spermatogonia and round sperms derived from wild-type and DAXX gene knockout mice. The samples aim to investigate the effects of DAXX gene deletion on reproductive health. The dataset comprises biological replicates for each condition, including both control (wild-type) and experimental (knockout) groups, to ensure statistical robustness and reproducibility of results

first: Zejia

last: Li

first: Chaoyang

last: Xiong

first: Jin

last: Shen

first: Chen

last: Tan

first: Dupeng

last: Ma

first: Jun

last: Chen

first: Rong

last: Liu

first: Qi

last: Sun

first: Weihao

last: Gong

first: Wenming

last: Yuan

first: Mengya

last: Huang

first: Li

first: Yueqiu

first: Guohong

first: Mengcheng

last: Luo


title: DNA supercoiling impacts alternative transcription start site selection in yeast

status: Public on Mar 24 2025

first: Folkert

last: van Werven

summary: Most genes are transcribed from multiple transcription start sites (TSSs), defined as alternative TSSs, which are highly regulated and can lead to various gene regulatory outcomes including changes in translation efficiency and protein isoforms. Transcription factors and chromatin regulators control alternative TSS selection. DNA supercoiling affects multiple aspects of transcription including transcription initiation. However, its regulatory effect on genes with multiple TSSs is not known. Here we investigated how DNA supercoiling impacts alternative TSS usage in Saccharomyces cerevisiae. We depleted topoisomerases during early meiosis, where alternative TSS usage is prevalent, and applied an improved TSS sequencing protocol. We show that supercoiling affects alternative TSS usage of almost 600 genes. Increased alternative and aberrant TSS usage were observed near and within open reading frames, likely resulting from transcription-induced supercoiling originating from upstream alternative TSSs. DNA supercoiling had the greatest impact on genes with a dominant alternative TSS, significant spacing between alternative TSSs, and greater overall length. Our results establish that DNA supercoiling release during transcription is critical for correct TSS selection.

overall_design: Top1/2 depletion at 0h and 6h in SPO. Controls Top1/2-AID, TIR, Top1/2 AID+TIR at 0h and 6h. Total 18 samples.

first: Eleanor

last: Elgood Hunt

first: Claudia

last: Vivori


title: Electrostatic properties of disordered regions control transcription factor search and pioneer activity [ChIP-Seq]

first: Sim

last: Sakong

overall_design: ChIP-seq experiments, utilizing a Halo-tag antibody, were conducted in CGR8-derived mouse embryonic stem cells expressing Halo-Sox2, Halo-Sox2a, Halo-Sox17, or Halo-Sox17b upon doxycycline addition. Two biological replicates of the ChIP procedure were carried out in the cells expressing Halo-Sox2 or Halo-Sox2a.

first: Beat

last: Fierz

first: David

last: Suter


title: Glycocalyx micro- and nanodomains in cell-cell and cell-matrix adhesions revealed by sensitive click chemistry

status: Public on Mar 18 2025

city: Nijmegen

first: Daan

last: Smits

address: Geert Grooteplein 26

department: MBS

institute: RadboudUMC

summary: The glycocalyx consists of glycoproteins, glycolipids and extracellular polysaccharides at the cell surface which mediate viscoelastic and electrostatic barrier function. In molecular interactions the glycocalyx is thought to segregate locally to facilitate receptor-ligand binding, yet high-resolution maps of glycocalyx domains in cell-cell and cell-matrix interactions are lacking. We here applied TMTH-sulfoximine (THS)-based biorthogonal chemistry in live-cell culture and demonstrate much-enhanced glycocalyx detection, compared to established  dibenzocyclooctyne-based labeling. Using superresolution microscopy, we identified micron-scale moderately diminished glycocalyx in cell-cell contacts and subtotal depletion in protrusions at the leading and trailing edges and membrane blebs when cells invaded 3D fibrillar matrix. At contacts to collagen fibrils, focal integrin clusters segregated in outward-protrusive nanodomains from glycocalyx (distance: 350 nm), forming adhesion sites of low glycocalyx content. Thus, THS-based next-generation bioorthogonal labelling of live cells identifies micro- and nanodomains with altered glycocalyx density, implicating local glycocalyx downregulation in functional cell-cell and cell-matrix interactions.

overall_design: RNA-sequencing to determine transcriptomic interference by metabolic labelling using Ac4ManNAz (5-day treatment), so comparing treated vs. untreated.

first: Yuri

last: Damen

first: Tianfu

last: Li

first: Floris

last: van Delft

first: Bauke

last: Albada

first: Peter

last: Friedl


title: Endometrial assembloid model reveals endometrial adenogenesis regulation by estradiol-driven WNT7B expression [scRNA-seq]

status: Public on Mar 29 2025

first: Yimeng

summary: To establish an in vitro endometrial co-culture model, endometrial organoid was co-cultured with endometrial stromal cells in a 3D hydrogel matrix. During this co-culture process, tubular gland development was identified from endometrial organoids, which showed the molecular, morphological and functional properties of human endometrial glands. To further characterize the genetic drive for this phenomenon, we conducted single-cell RNA sequencing analysis at day 1 and day 9 after co-culture.

overall_design: Three-dimensional co-culture models were collected separately at day 1 and day 9 after co-culture, digested into single cells and send for single-cell RNA sequencing analysis to identify the regulatory genes.

first: Xintong

first: Jianlin

first: Cheuk

last: Lee

first: Philip

last: Chiu


title: Transcriptomic analysis for temporomandibular joint arthritis

status: Public on Mar 06 2025

city: Sharjah

first: Rifat

last: Hamoudi

state: Sharjah

address: M32 Sharjah Institute for Medical Research, College of Medicine, University of Sharjah, , Sharjah

department: Clinical Sciences

country: United Arab Emirates

institute: University of Sharjah

summary: Background: Temporomandibular joint osteoarthritis (TMJOA) is a degenerative disease marked by the progressive degeneration of cartilage and underlying bone, resulting in pain and functional impairment. Despite current conservative treatments such as physical therapy and NSAIDs, the etiology and pathophysiology of TMJOA are unclear, making effective management difficult. Finding reliable biomarkers for early identification and new therapeutic targets is crucial to improve treatment outcomes.

overall_design:
Methods: The goal of this study was to identify possible salivary biomarkers for TMJ-OA through an integrative strategy that included both in silico  and clinical validation. Transcriptomic data from the Gene Expression Omnibus (GEO) dataset GSE205389 were analysed to identify differentially expressed genes (DEGs) in TMJ-OA patients versus healthy controls. RNA sequencing was carried out on saliva samples from both group, TMJ-OA patients and healthy controls. DEGs were validated with quantitative reverse transcription PCR (qRT-PCR). Gene set enrichment analysis (GSEA) and pathway annotation tools such as Metascape were used to uncover biological pathways linked to TMJ-OA.
Results: In silico analysis revealed 81 differentially active pathways, with 3554 DEGs substantially associated with TMJOA. Among the top 20 DEGs, CRIP1, PPA1, TARS1 and GCLC were significantly increased in TMJ-OA patients compared to controls. These genes were engaged in inflammation, metabolic processes, and oxidative stress, indicating a role in illness progression. Actin filament polymerization, osteoclast differentiation, and immune regulatory pathways were shown to be significantly enriched.
Conclusion: This work reveals CRIP1, PPA1,TARS1 and GCLC as possible salivary biomarkers for TMJ-OA, offering novel insights into disease pathophysiology. The findings imply that these genes could be used as diagnostic markers and therapeutic targets for TMJ-OA, particularly in the Arabic population. Additional large-scale research is required to confirm these biomarkers and investigate targeted therapies for illness management.
Temporomandibular joint osteoarthritis (TMJOA) and healthy control

first: Nawal

last: Alketbi

first: Alaa

last: Altaie

first: Reem

last: Alhamidi

first: Mohamed

last: Haider

first: Wael

last: Talaat


title: Methylation pseudotime analysis for label-free profiling of the temporal chromatin landscape with long-read sequencing

status: Public on Mar 05 2025

city: Irvine

first: Annie

last: Trinh

address: 419 S. Circle View Drive

department: Microbiology & Molecular Genetics

institute: University of California, Irvine

summary: Faithful epigenetic inheritance across cell divisions is essential to maintaining cell identity and involves numerous epigenetic modifications, whose roles in coordinating chromatin architecture are less understood. Technological approaches to temporally order epigenetic modifications throughout the cell cycle often face limitations in sequence resolution and rely on potentially damaging mitotic labeling or conversion steps. Herein, we present Methylation Pseudotime Analysis Through read-level Heterogeneity (MPATH), a label- and conversion-free method to infer post-replication DNA strand maturity from methylation patterns across single molecules. We showed that MPATH is generally applicable across human embryonic stem cell (hESC) lines in which CpG methylation heterogeneity is coupled to replication, but not applicable to breast cancer cells, in which CpG methylation heterogeneity may also be associated with other factors. We used MPATH to track hydroxymethylated regions throughout mitotic inheritance in hESCs and observed that in the absence of ten-eleven translocation (TET) enzymes, these regions appeared to progress more quickly across replication-associated timescales, suggesting that enzymatic competition may drive temporal heterogeneity in CpG methylation patterns. When applied to long reads generated by NOMe-seq, MPATH revealed that nucleosome compaction and CpG methylation uniformity are coupled across replication-associated timescales. Finally, extension of MPATH to phased reads reveals allele-specific trends in pseudotime distribution associated with X chromosome inactivation. Our findings suggest that when coupled with multimodal sequencing strategies, MPATH could provide valuable insights into chromatin restoration dynamics.

overall_design:
HUES64 hESCs, H9 hESCs, HUES8 TKO hESCs and MDA-MB-231 breast cancer cells were subjected to several different experimental conditions prior to DNA extraction for long-read sequencing. In the first experiment, HUES64 hESCs were treated with 1µg/ml nocodazole for 16h. BrdU pulse-chase experiments were conducted on all cell lines to investigate DNA methylation patterns during chromatin maturation. For the pulse optimization, cells were subjected to 50 µM BrdU for durations of 10 minutes, 20 minutes, 30 minutes, and 1 hour, followed by genomic DNA collection at 0h (nascent chromatin) and 16h (mature chromatin). Mock controls were included for both 0h and 16h timepoints. Optimized labeling conditions included a 500 µM BrdU pulse (10 minute duration) with 0h and 16h chase points. Unlabeled HUES64 samples were also generated (sequenced to a total of 62x coverage across 2 technical replicates). All long-read data was acquired using ONT sequencing platforms. In a separate sequencing experiment, hESCs were subjected to Repli-ATAC sequencing in order to temporally profile chromatin accessibility across replication-associated timescales.

>>>Raw data in POD5 or FAST5 will be available upon request by the submitter.<<<

first: Kehan

last: Ren

first: Nandor

last: Laszik

first: Navied

last: Akhtar

first: Kwadwo

last: Bonsu

first: Tanye

last: Wen

first: Asia

last: Mendelevich

first: Yoshiko

last: Takahashi

first: Julien

last: P Morival

first: Katelyn

last: Diune

first: Mitchell

last: Frazeur

first: Justin

last: Vega

first: Edouard

last: Aleman

first: Minji

last: Byun

first: Alexander

last: Gimelbrant

first: Elizabeth

last: Read

first: Timothy

last: Downing


title: Construction of a miRNA panel as possible biomarker for separation of glioblastoma and brain metastasis patients with adenocarcinoma origin using next-generation sequencing

city: Debrecen

first: Zsuzsanna

last: Birkó

address: Egyetem tér 1.

department: Humangenetics

country: Hungary

institute: University of Debrecen

summary: Next Generation Sequencing technique was performed to compare the miRNA expression pattern of tumor  brain tissue sample of 6 Glioblastoma patients, and 6 patients with brain metastases (BM) originated from lung adenocarcinoma (LUAD).  In order to analyse the difference on miRNA expresion level between GBM and LUAD-BM cases, we applied cluster analysis on the NGS dataset of 6 samples for each of the two goups  with iDEP 1.1 software. Log2FC values were calculated to determine the exact level of up- and downregulation in case of the deregulated miRNAs, using the iDEP 1.1 web tool applying the DESeq2 algorithm. Evaluate the results of analysis 99 known miRNAs were detected with altered expression using a threshold of false discovery rate (FDR) <0.05 and fold-change> 2. Among them, 62 miRNAs were upregulated (log2FC > 2) and 37 miRNAs were downregulated (log2FC < -2) with biological revelance in tissue LUAD-BM samples compared with the GBM samples. To validate our results obtained by NGS, four upregulated miRNAs: hsa-miR-200c-5p, hsa-miR-141-5p, hsa-miR-200a-5p, hsa-miR-375-3p  and two downregulated miRNAs: hsa-miR-410-3p, hsa-miR-9-5p were chosen for RT-qPCR analysis. As the result of that  hsa-miR-200c-5p, hsa-miR-141-5p, hsa-miR-200a-5p, hsa-miR-375-3p was significantly upregulated, while hsa-miR-410-3p, hsa-miR-9-5p was significantly downregulated in LUAD-BM - GBM comparison.

overall_design: Tissue samples were gathered from brain tumor region of six individuals diagnosed with GBM, and from brain tumor region of six patients with LUAD-BM. All samples in both group were confirmed histopathologically. These intraoperative quick-frozen tissue samples were stored at -80⁰C until further processing. Total RNA was isolated from  the tissue samples of GBM and LUAD-BM as well. Global transcriptomic profile was studied by RNA-sequencing applying the Illumina NextSeq500 platform including microRNA sequencing. In the case of all samples 3 replicates were included. In order to identify up-, or down-regulated microRNAs the LUAD-BM samples were compared to GBM samples..

first: Bernadett

last: Torner

first: Dóra

last: Géczi

first: Álmos

last: Klekner

first: István

last: Balogh

first: András

last: Penyige


title: METTL5 deficiency induces lung cancer regression via mitochondrial translation reduction

status: Public on Mar 01 2025

city: nanjing

first: Zhou

last: Jun

state: jiangsu

address: longmiandadao

department: College of life science and technology

institute: China Pharmaceutical University

summary: Cellular protein synthesis occurs through two types of translation: cytoplasmic and mitochondrial. These two types of translation operate independently but in coordination to regulate cell survival and death. Methyltransferase-like 5 (METTL5) has been found to play a role in directing the m6A modification of rRNA, which in turn controls cytoplasmic translation and impacts the decisions related to cell fate. In this study, we employed Ribo-seq to explore the effects of METTL5 deficiency on both cytoplasmic and mitochondrial translation.

overall_design: To investigate the potential role of METTL5 in the regulation of non-small cell lung cancer, a stable METTL5 knockout cell line was initially constructed using classical A549 and H1299 NSCLC cells using a CRISPR Cas9 sgRNA-based gene interference method.

first: Qiujie

last: Li

first: Jun

last: Zhang


title: Anti-VEGF treatment potentiates ICB responses through a BAFF and IL-12-dependent reprogramming of the TME

status: Public on Mar 16 2025

city: BETHESDA

first: Cihan

last: Oguz

address: NIHBC 29B BG RM 1NN20, 9000 ROCKVILLE PIKE

institute: NIH

summary: Anti-VEGF treatment has shown clinical activity in combination with checkpoint inhibitor therapy, but the exact mechanism is not known. We show that adding VEGF blockade to the ICB combination of anti-CTLA4 + anti-PD-L1 in cholangiocarcinoma unleashed a therapeutic efficacy driven through a novel multimodal mechanism dependent on BAFF production by T and myeloid cells, leading to a BAFF-dependent proinflammatory B cell response characterized by Germinal center B cell and plasma cell expansion, as well as IL-12 production. Using single cell RNA sequencing (scRNA-Seq) of paired pre- and post-treatment tumor biopsy samples from patients treated with anti-CTLA4, anti-PD-L1 and anti-VEGF, we characterized the treatment-associated immunological changes.

overall_design: We performed single cell RNA sequencing (scRNA-Seq) of liver cancer tumor biopsy samples (pre- and post-treatment) from three patients that contributed a total of seven samples (paired pre- and post- treatment samples from patients 1 and 2, and one pre-treatment and two post-treatment samples from patient 3).

first: Mohamed-Reda

last: Benmebarek

first: Matthias

last: Seifert

first: Benjamin

last: Ruf

first: Yuta

last: Myojin

first: Kylynda

last: Bauer

first: Patrick

last: Huang

first: Chi

last: Ma

first: Marina

last: Villamor-Payà

first: Francisco

last: Rodriguez-Matos

first: Marlaine

last: Soliman

first: Rajiv

last: Trehan

first: Cecilia

last: Monge

first: Changqing

last: Xie

first: David

last: Kleiner

first: Bradford

last: Wood

first: Elliot

last: Levy

first: Anuradha

last: Budhu

first: Noemi

last: Kedei

first: Christian

last: Mayer

first: Xin

last: Wang

last: Lack

first: William

last: Telford

first: Firouzeh

last: Korangy

first: Tim

last: Greten


title: Dbf4-Dependent Kinase Finetunes INO80 Function at Chromosome Replication Origins

city: Planegg

first: Priyanka

last: Bansal

state: Bavaria

address: Großhaderner Str. 9

department: Molecular Biology

institute: Biomedical Center

summary: The highly conserved Dbf4-Dependent Kinase (DDK) plays a pivotal role in the nucleus during S phase, where it directly phosphorylates the replicative helicase, the minichromosome maintenance (MCM) complex. This leads to the initiation of chromosome replication. However, aside from the MCM complex, few other targets have been identified to date, leaving DDK an understudied kinase. Here, we describe a two-pronged mass spectrometry-based approach and define the nuclear DDK-dependent phosphoproteome, which consists of approximately 400 phosphorylation events. Within this network, we found that DDK directly phosphorylates the Arp8 subunit of the multi-subunit chromatin remodeler complex INO80. Arp8 phosphorylation stabilises INO80's intramolecular complex integrity, which finetunes its nucleosome spacing activity at replication origins. This adjustment of origin chromatin architecture stimulates replication and is important for the response to replication stress. Our results represent a significant advance in our understanding of the molecular mechanisms underlying the regulation of replication origins.

overall_design: It was tested whether DDK phosphorylation of INO80 might influence nucleosome positiong at origins of relication. Nucleosome positioning analysis was performed using MNase-seq data obtained remodeling of SGD (salt gradient dialysis) chromatin by INO80 WT and INO80-AA mutant purified protein. Futher, to test if changes in nucleosome positioning is because of DDK dependent phosphorylation, we treated our purified INO80 and INO80-AA protein with Lamda phosphatse, rephosphorylated with DDK and repurified the proteins to perform nucleosome positioing assay. We also tested nucleosome positioning in vivo in WT and ARP8 mutant strain by isolating nuclei. Atleast two biological for each experiment were performed.

first: Shibojyoti

last: Lahiri

first: Chandni

last: Kumar

first: Lorenzo

last: Galanti

first: Erika

last: Chacin

first: Jessica

last: Furtmeier

first: Garp

last: Linder

first: María

last: Ortíz-Bazán

first: Marisa

last: Müller

first: Petra

last: Vizjak

first: Tobias

last: Straub

first: Felix

last: Mueller-Planitz

first: Johannes

last: Stigler

first: Andrés

last: Aguilera

first: Belen

last: Gómez-González

first: Boris

last: Pfander

first: Axel

last: Imhof

first: Christoph

last: Kurat


title: Mycobacterium tuberculosis MutT4 is an RNA pyrophosphohydrolase that forms biomolecular condensates and sensitizes mRNAs to degradation

status: Public on Mar 26 2025

city: Worcester

first: Junpei

last: Xiao

state: MA

address: 100 Institute Rd

institute: Worcester Polytechnic Institute

summary: Bacterial adaptation to stress involves changes in transcription and mRNA degradation rates. In Escherichia coli, the Nudix hydrolase RppH initiates mRNA degradation by removing pyrophosphate from mRNA 5’-ends, converting 5’-triphosphates to 5’-monophosphates. We aimed to identify the RppH homolog in the globally important pathogen Mycobacterium tuberculosis (Mtb). We deleted each non-essential Nudix gene from Mtb to determine their impacts on mRNA 5’-phosphorylation. Deletion of mutT4 (Rv3908) increased the relative abundance of 5’-triphosphates on myriad mRNAs across the transcriptome. Purified MutT4 converted mRNA 5’-triphosphates into monophosphates, and stimulated degradation by RNase E and RNase J. MutT4 has intrinsically disordered regions (IDRs), a common domain for biomolecular condensate formation. Microscopy showed that MutT4 forms condensates that dissociate upon addition of rifampicin, and that the N-terminal IDR is sufficient for condensate formation. These MutT4 condensates localize with RNase E and RNase J. Deletion of mutT4 in Mtb leads to a higher outer membrane permeability and resistance to oxidative stress. We conclude that MutT4 is the RppH of Mtb, assembling in condensates that may act as degradation hubs. Our data indicate that MutT4 is unlikely to participate in DNA repair or nucleotide pool cleansing, and as such would more accurately be called RppH.

overall_design: The mutT4 gene of M. tuberculosis strain mc26030 (H37Rv ΔRD1 ΔpanCD) was partially replaced by a zeocin resistance gene by recombineering. Care was taken to disrupt the function of mutT4 without disrupting adjacent genes. The deletion strain was complemented by integration of a plasmid containing mutT4 with its native promoter and 5' UTR into the Giles phage integration site. Triplicate cultures of the parental, deletion, and complemented strains strains were grown to log phase in 7H9 media supplemented with OADC (0.05 g/L oleic acid, 5 g/L bovine serum albumin fraction V, 2 g/L glucose, 0.85 g/L NaCl, and 4 mg/L catalase), 0.2% glycerol, 0.05% Tween 80, and 24 μg/mL pantothenate. RNA was harvested and rRNA was depleted as described in (Culviner, P.H., Guegler, C.K. and Laub, M.T. (2020) A simple, cost-effective, and robust method for rRNA depletion in RNA-sequencing studies. MBio, 11, 10.1128/mbio. 00010-00020.). Samples were then treated with E. coli RppH or mock-treated, and 5'-end directed RNAseq libraries were constructed as described (Martini, M.C., Sun, H. and Shell, S.S. (2021) RNA Sequencing for Transcript 5'-End Mapping in Mycobacteria. Methods Mol Biol, 2314, 513-531).

first: Juan Hilario

last: Cafiero

first: Irene

last: Lepori

first: Abigail

middle: R

last: Rapiejko

first: Manchi

last: Reddy

first: Louis

last: Roberts

first: M Sloan

last: Siegrist

first: James

last: Sacchettini

first: Scarlet

last: Shell


title: Functional chromatin signatures premark future lineage-specific enhancers [scRNA-seq]

city: New York

first: Julian

last: Pulecio

department: Developmental Biology

institute: Memorial Sloan Kettering Cancer Center

summary: It remains unknown whether early embryonic cells harbor a blueprint for future enhancers that regulate the expression of lineage-specific genes in adult tissues. Here, we demonstrate that embryonic stem cells (ESCs) have transcriptionally competent chromatin regions (CCRs) prepared to induce the expression of lineage genes prior to differentiation. CCRs represent activatable pre-enhancers within the topological chromatin domains of lineage genes, marked by chromatin signatures distinguishable from primed/poised enhancers, enabling their genome-wide identification. The pioneer transcription factor (TF) FOXA2 preferentially binds CCRs during early lineage specification, promoting their conversion into active enhancers. CCRs can be harnessed to boost the expression of master TFs and promote the direct reprogramming of ESCs into differentiated cells, showcasing their potential for practical applications. Our findings identify a mechanism by which ESCs rapidly establish enhancer activity during early lineage differentiation and expand our understanding of the epigenetic features supporting transcriptional regulation and cellular plasticity.

overall_design: Single-cell 10x genomics experiments were performed using hESCs after interrogating candidate CCRs through CRISPRa screens. We employed a gRNA backbone inspired on the CROP-Seq system that allows the dual identification of gRNAs and single-cell transcriptomics. gRNA and transcriptome libraries were generated and sequenced. Final analyses were done using CellRanger and the SCEPTRE algorithm.

first: Renhe

last: Luo

last: Hu

first: Jielin

last: Yan

first: Danwei

last: Huangfu


title: Melanoma plasticity is regulated by mTOR-mediated crosstalk between MITF and TFE3

status: Public on Mar 20 2025

city: Iowa City

first: Jeremy

last: Chang

state: IA

address: 285 Newton Rd

department: Surgery

institute: University of Iowa

summary: Bulk RNAseq was performed to identify differences in gene expression in SKMEL28 cells as related to MITF and TFE3 expression.

overall_design:
TFE3KO cell lines were created in the D2-MITFKO SKMEL28 and A375 background via CRISPR-cas9 strategies
500k SKMEL28: WT, D2 MITFKO, D2 MITFKO/TFE3KO (clone 3 and clone 13) and A375: WT and TFE3KO cells were plated in triplicate for biologic replicates. RNA was extracted utilizing RNeasy Plus Kit (Qiagen).

first: Colin

last: Kenny


title: Arcuate nucleus of the hypothalamus cells are sensitive to nutritional state in a sex dependent manner.

status: Public on Feb 24 2025

city: Houston

first: Jonathan

middle: Carter

last: Bean

state: TX

address: 1100 Bates Ave

department: Pediatrics

institute: Baylor College of Medicine

summary: Obesity has reached epidemic levels and obesity increases the risk for negative health outcomes including type 2 diabetes. Sex differences are evident in metabolic homeostasis with differences seen both in amount and location of adipose stores. Males store more adipose viscerally whereas females store adipose subcutaneously. Interestingly, this can be affected by the brain. The arcuate nucleus of the hypothalamus is a key brain region that exerts control over feeding behavior and metabolism. The cells of the arcuate are heterogeneous in their transcriptomic expression and in their functions. Here we perform single nuclei RNA-sequencing of the arcuate in females and males under varying nutritional states. We found that Agrp neurons were the most sensitive to nutritional state and this was more pronounced in females than in males. KNDy neurons were also sensitive to nutritional state and major sex differences were observed. Dopamine neurons of the arcuate had major sex differences. Ghrh neurons saw moderate nutritional regulation and sexual dimorphism. Both microglia and oligodendrocytes had moderate differences observed between nutritional states. This study represents the first major effort to characterize sex differences across nutritional states in the arcuate.

overall_design: Female and male mice were either ad. Libitum fed or fasted for 28 hours, had their arcuate nucleus of the hypothalamus collected and processed for snRNA-seq.

middle: C

first: Yong

last: Xu


title: Influenza virus exposure shapes the B cell response to influenza vaccination fifty years later

status: Public on Feb 28 2025

city: Bethesda

first: Sarah

last: Andrews

state: Maryland

address: 9000 Rockville Pike

institute: National Institutes of Health

summary: Pre-existing immunity impacts vaccine responses to endemic respiratory viruses such as influenza, but directly connecting influenza infections early in life with immune responses decades later is difficult.  However, H2N2 stopped circulating in the human population in 1968, creating the opportunity to directly evaluate the impact of early H2N2 exposure on vaccine responses 50 years later.  Here, we vaccinate individuals born before (H2 Exposed) or after (H2 Naïve) 1968 with an H2 HA DNA plasmid and/or a ferritin nanoparticle vaccine.  H2 Exposed individuals generated a rapid B cell recall response that differed in potency, cross-reactivity, immunoglobulin repertoire, epitope targeting and phenotype from the de novo response in H2 Naïve individuals.  Furthermore, vaccinating with a DNA versus a protein nanoparticle vaccine altered the response in H2 Naïve but not H2 Exposed individuals.  This study clearly establishes and describes the life-long impact of influenza HA-specific memory B cells formed early in life on vaccine responses decades later.

overall_design: Memory b cells and plasmablasts were isolated at various timepoints from PBMCs of individuals undergoing vaccination against H2 influenza. Antibody derived tag, whole transcriptome, and VDJ libraries were generated for all samples.


title: Gene expression profile at single cell level of hepatocytes and non parenchymal cells (NPCs) isolated from a murine model of MASLD

city: MIlano

first: Paola

last: Dongiovanni

address: Via Pace 9

country: Italy

institute: Fondazione IRCCS Cà Granda ospedale Policlinico

summary: Metabolic dysfunction-associated steatotic liver disease (MASLD) embraces different conditions, including metabolic dysfunction-associated steatohepatitis (MASH), fibrosis, cirrhosis and hepatocellular carcinoma (HCC). The landscape of cellular abnormalities occurring in the different stages of MASLD as well as the processes which drive MASLD evolution are not completely clarified. We used single cell RNAsequencing (scRNAseq) to unravel cellular heterogeneity affecting livers during the switching from simple steatosis towads MASH and and MASH-fibrosis.

overall_design: Heps and NPCs were isolated by in situ digestion and collagenase perfusion. Cells were separed by centrifugation, and live cells were FACS sorted. Heps fraction was CD45 depleted.

first: Marica

last: Meroni

first: Ettore

last: Mosca


title: RpdD and RpoS ChIP-exo in Klebsiella pneumoniae

city: Ulsan

first: Donghyuk

last: Kim

address: 50 UNIST-gil, Eonyang-eup, Ulju-gun

department: Department of Chemical Engineering

country: South Korea

institute: UNIST

summary: With increasingly concerning strains of antimicrobial resistant strains of the commensal, gram-negative bacteria Klebsiella pneumoniae emerging, there is a pressing need to better understand the pathogen and mechanisms behind its pathogenicity. This study investigated the regulatory mechanisms in strain MGH 78578 of two major sigma factors, the house-keeping sigma factor RpoD, and the general stress response sigma factor RpoS, in mid-exponential and early stationary phase using chromatin immunoprecipitation with exonuclease treatment (ChIP-exo) followed by deep sequencing. Combining ChIP-exo and transcriptome analysis allowed for the determination of sigma factor binding sites, binding motifs, and genes included in the phase-specific sigmulons. The number  of genes included in the RpoS sigmulon was greater than in the RpoD sigmulon, with 1,833 and 1,690 genes included, respectively; however, a majority of sigmulon genes were found in all phase-specific sigmulons. Focussing on pathogenicity  genes, 20 antimicrobial resistance genes (ARGs) and 155 virulence genes, only two ARGs were found exclusively in one phase-specific sigmulon, an oxacillin-hydrolysing class D beta-lactamase and chloramphenicol efflux MFS transporter CmlA5, which were found in the RpoD sigmulon in early stationary phase. Notably, six unnamed proteins that are or pertain to fimbrial proteins were found uniquely in the RpoS sigmulon in early stationary phase. From this, it can be hypothesised that early stationary phase might be an important phase for pathogenicity gene regulation. While there is little conservation in RpoS sigmulons from strain to strain, RpoS appears to have a consistent overarching role across strains, including a role as a regulator of pathogenicity genes.

overall_design: [ChIP-exo] Klebsiella pneumoniae MGH 78578 cells were cultured in M9 minimal media with 0/2% glucose to etiher mid-exponential or stationary phase in biological duplicate or triplicate

first: Zoe

middle: K

last: Scott-Nevros


title: CEBPB-high dormant tumor cells drive immune evasion [ChIP-Seq]

status: Public on Feb 21 2025

city: Shanghai

first: Yang

last: Dong

state: 上海

address: Shanghai

institute: Tongji University

summary: Triple negative breast cancer (TNBC) poorly responds to immune checkpoint blockade (ICB) therapy. Dormant tumor cells are recognized as immunotherapy-resistant reservoirs, potential leading to tumor relapse, highlighting the importance to elucidate the mechanisms underlying immunotherapy resistance. Herein, we demonstrated that dormant tumor cells were resistant to ICB therapy and occupied an immunosuppressive niche with enhanced tumor-associated macrophages (TAMs) and decreased CD8+T cells infiltration in mouse TNBC allografts via a label-retention system. CEBPB was highly expressed in dormant tumor cells and maintained tumor dormancy by transcriptionally activating cell cycle negative regulators like CCNG2 and p27.

overall_design: ChIP-seq of CEBPB in 4T07 cells.

first: Jin

last: Bai

last: Gao


title: Stress drives plasticity in leaf aging transcriptional dynamics in Arabidopsis thaliana

status: Public on Feb 26 2025

city: La Jolla

first: Joseph

last: Ecker

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: We investigated the transcriptional basis of how drought stress reshapes Arabidopsis leaf development. We profiled 1,226 leaves at various developmental stages and levels of drought stress, and generated a single-nucleus transcriptome atlas comprising ~1 million individual nuclei.

overall_design: Using sci-RNA-seq3 and 10X transcriptional approaches, we analysed the gene expression changes of different individual cell types of Arabidopsis to drought stress

last: Swift

first: Xuelin

last: Wu

first: Tanvi

last: Jain

first: Jiaying

first: Natanella

last: Illouz-Eliaz

last: Nery

first: Joanne

last: Chory


title: Methylation pseudotime analysis for label-free profiling of the temporal chromatin landscape with long-read and single-molecule resolution

first: Annie

last: Trinh

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: CEBPB-high dormant tumor cells drive immune evasion [RNA-Seq]

overall_design: To investigate which genes were regulated by CEBPB in triple negative breast cancer, we transfected 4T07 cell lines with either pLenti-CMV-MCS or pLenti-CMV-Cebpb by lentivirus. Stable tracfected cells were selected by puromycin.We extracted RNA from each group with three biological replicates. We performed gene expression profiling assay using data obtained from RNA-seq of 6 samples (two groups with three biological replicates per group). Comparative gene expression profiling analysis of RNA-seq data for OE-Control and OE-Cebpb


title: Supinoxin Blocks Small Cell Lung Cancer Progression by Inhibiting Mitochondrial Respiration through DDX5

status: Public on Feb 20 2025

city: West Lafayette

first: Elizabeth

last: Tran

state: IN

address: 175 S. University Street

department: Biochemistry

institute: Purdue University

summary: DDX5 is a DEAD-box RNA helicase that is overexpressed and implicated in the progression of several cancers, including small cell lung cancer (SCLC).  Our laboratory has demonstrated that DDX5 is essential for the invasive growth of SCLC and mitochondrial respiration.  SCLC is an extremely lethal, recalcitrant tumor, and currently lacking effective treatments. Supinoxin (RX 5902), a compound having anti-cancer activity, is a known target of phosphor-DDX5.  We now report that Supinoxin inhibits the proliferation of chemo-sensitive and chemo-resistant SCLC lines, H69 and H69AR respectively.  Additionally, Supinoxin mitigates both the growth of H69AR xenograft tumors and SCLC PDX tumors in vivo.    Finally, we find that Supinoxin inhibits expression of mitochondrial genes and effectively blocks respiration.  These studies suggest that Supinoxin functions in anti-tumor progression by reducing cellular energy levels through DDX5.

overall_design: To investigate the molecular pathway through which Supinoxin works on SCLC, we conducted differential gene expression in H69AR chemoresistant SCLC cells with or without 70 nM of Supinoxin.

first: Subhadeep

last: Das

first: Maria

middle: P

last: Zea

first: Matthew

last: Russon

first: Zheng

last: Xing

first: Sandra

last: Torregrosa-Allen

first: Heidi

last: Cervantes

first: Haley

middle: A

last: Harper

first: Bennett

last: Elzey


title: Reduced TBX5 dosage causes human atrial disease by undermining developmental gene regulation of atrial cardiomyocyte identity [scRNA-seq]

status: Public on Feb 18 2025

city: San Francisco

first: Benoit

last: Bruneau

address: 1650 Owens St

institute: Gladstone Institutes

summary: In a TBX5-linked human cellular model of atrial disease, we examined the gene networks controlled by TBX5 during human atrial differentiation using single-cell RNA and ATAC sequencing.

overall_design: Wildtype, TBX5 heterozygous and homozygous mutant iPSCs were differentiated to atrial cardiomyocytes. At day 20 or day 45 of differentiation, cells were harvested for single-cell RNA sequencing and analyzed using Seurat. Two replicates per genotype per timepoint were harvested.

first: Kavitha

middle: S

last: Rao

first: Irfan

last: Kathiriya


title: Copy number normalization distinguishes differential signals driven by copy number differences in ATAC-seq and ChIP-seq

city: Tübingen

first: Yingguang Frank

last: Chan

address: Max Planck Ring 9

department: Chan group

institute: Friedrich Miescher Laboratory of the Max Planck Society

summary: A common objective across ATAC-seq and ChIP-seq analyses is to identify differential signals across contrasted conditions. However, in differential analyses, the impact of copy number variation is often overlooked. Here, we demonstrated copy number differences among samples could drive, if not dominate, differential signals. To address this, we propose a pipeline featuring copy number normalization. By comparing the averaged signal per gene copy, it effectively segregates differential signals driven by copy number differences from other factors. Further applying it to Down syndrome, we unveiled distinct dosage-dependent and -independent changes on chromosome 21. Thus, we recommend normalization as a general approach.

overall_design: To characterize the endogenously formed G-quadruplex structures in wildtype and BLM-/-  cells, we performed G4 ChIPseq with a specific antibody against the secondary DNA structures. During the process of characterizing the differential G-quadruplex forming sites, we noticed that copy number differences at local regions between the samples skew the differential analysis towards the sample with a higher copy number. We explored the impacts of copy number variation on the differential analysis and solutions to segregate the differential signals driven by copy number variation.

first: Dingwen

last: Su


title: A Novel EBF2 Nonsense Variant Drives Adipose Tissue Fibrosis and Dysfunction in Partial Lipodystrophy Syndrome

status: Public on Feb 17 2025

city: ANN ARBOR

first: Tae-Hwa

last: Chun

state: MI

address: 2800 PLYMOUTH RD, NCRC B10-A186, NCRC B10-A186

department: Internal Medicine

institute: University of Michigan

summary: Heterozygous Ebf2 p.E165X knock-in (KI) mice exhibited restricted adipogenesis and excess extracellular matrix (ECM) deposition during post-weaning AT expansion and high-fat-diet (HFD)-induced obesity.

overall_design: Bulk RNA-seqeuding of inguinal white adipose tissues (IWAT) isolated from  4-week-old wildtype (Ebf2+/+) and knock-in (Ebf2 p.E165X/+) male mice on regular chow diet.

first: Donatella

last: Gilio

first: Weisheng

last: Wu

last: Foss-Freitas


title: Sex-specific extracellular matrix remodeling in skeletal muscle after extended immobilization

status: Public on Feb 15 2025

city: Richmond

first: Jessica

middle: Rose

last: Williamson

address: 70 S Madison St

department: Biomedical Engineering

institute: Virginia Commonwealth University

summary: Extended periods of disuse can cause changes to the musculoskeletal system, particularly impacting the composition and strength of skeletal muscle. While many studies have investigated changes in muscle phenotype after unloading, majority of these studies were performed on animals of a single genetic background or sex. The effect of muscle immobilization on males compared to females, as well as genetic diversity, is not well understood. The objective of this study was to investigate differential effects of skeletal muscle immobilization in genetically diverse outbred (DO) males and females through large-scale transcriptomic analysis, histopathological, and functional analyses. Transcriptomic analysis revealed both male and female have downregulated metabolism pathways during unloading. Investigation of matrix modifying pathways revealed differential expression of collagen related pathways, not matrix degradation pathways, in males compared to females. Histopathological analysis of collagen and functional analysis of muscle cross sectional area and volume correlated directly with transcriptomic analysis.

overall_design: 16 week old Diversity Outbred (DO) mice were casted via single limb immobilization for a period of 3 weeks. RNA was isolated from gastrocnemius muscles and sequenced.

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