title: The GPR84 molecule is a mediator of a subpopulation of retinal microglia that promote TNF/IL-1α expression via the rho-ROCK pathway after optic nerve injury (scRNA-Seq)

status: Public on Sep 30 2023

city: Sendai

first: Kota

last: Sato

state: Miyagi

address: 1-1, Seiryo-machi, Aoba-ku

department: Ophthalmology

country: Japan

institute: Tohoku University

type: Expression profiling by high throughput sequencing

summary: We show that microglial gene expression patterns gradually change during RGC degeneration induced by optic nerve injury.

overall_design: Examination of 3 different timepoint in Iba1-positive retinal cells.

first: Michiko

last: Ohno-Oishi

first: Masaaki

last: Yoshida

first: Taimu

first: Takaharu

last: Aizawa

first: Yuto

last: Sasaki

first: Shigeto

last: Maekawa

first: Makoto

last: Ishikawa

first: Kazuko

last: Omodaka

first: Chihiro

last: Kawano

first: Ryuji

last: Ohue-Kitano

first: Ikuo

last: Kimura

first: Toru

last: Nakazawa


title: Oncogene EVI1 Drives Acute Myeloid Leukemia Via a Targetable Interaction with CTBP2 [RNA-seq]

city: Rotterdam

first: Ruud

last: Delwel

address: Dr. Molewaterplein 40

country: Netherlands

institute: Erasmus MC

summary: Acute myeloid leukemia (AML) driven by the activation of EVI1 due to chromosome 3q26/MECOM rearrangements is incurable with current chemotherapy regimens. Insight into the mechanism by which EVI1 drives myeloid transformation is needed to target EVI1 in those leukemias. Here we demonstrate recurrent interaction of CTBP1/2 with a unique PLDLS motif in EVI1, which is indispensable for leukemic transformation of 3q26/MECOM rearranged AML. A PLDLS competitor construct outcompetes EVI1 to CTBP1/2 binding and inhibits AML proliferation in xenotransplant models. This proof-of-concept study opens the possibility to target one of the most incurable forms of AML using specific inhibitors directed to EVI1/CTBP1/2 interaction. Our findings have important implications for other tumour types with aberrant expression of EVI1 or other oncogenic transcription factors that also depend on CTBP1/2 interaction.

overall_design: PolyA-selected, non-stranded RNA-seq of MUTZ3, an AML cell line with inv(3), under 3 experimental conditions: 1) treated with a construct that contains PLDLS, which interferes between EVI1 and CTBP interaction; treated with a construct that contains PLASS, a similar peptide that does not interfere with this interaction; and treated with an empty vector.

first: Dorien

last: Pastoors

first: Roger

last: Mulet

first: Eric

last: Bindels


title: IGF2BP1-HAX-1 positive feedback loop-mediated HAX-1 overexpression blocks autophagic flux and promotes chemoresistance in nasopharyngeal carcinoma

city: Nantong

first: Siyu

last: Zhang

address: Xisi Road 20

country: China

institute: Affiliated Hospital of Nantong University

summary: Autophagic flux is associated with chemoresistance, the leading cause of chemotherapeutic failure. Here, we showed that HAX-1 promotes chemoresistance by effectively blocking the fusion of autophagosomes with lysosomes. Mass spectrometric and functional studies demonstrated that HAX-1 recruited NEDD4 to promote Rab7a degradation and inhibited the binding of Rab7a with SNAREs by competitively binding to it. Furthermore, HAX-1 could bind to IGF2BP1 mRNA, thereby contributing to its stability and translation. Moreover, IGF2BP1 enhanced HAX-1 m6A methylation, thereby enhancing its stability. Via in vivo and in vitro experiments, we confirmed the positive role of the IGF2BP1-HAX-1 feedback loop in chemoresistance. Our data provide evidence that HAX-1, IGF2BP1, and SQSTM1 levels are useful predictors of clinical outcome and chemoresistance risk. In addition, our data provide new insights into the clinical applications of therapies related to autophagic flux and its associated molecular network in targeting cisplatin chemoresistance in nasopharyngeal carcinoma.

overall_design: Guide RNA (sgRNA) preparation: sgRNA was designed using the CRISPR online tool (https://zlab.bio/guide-design-resources). The effective targeting site for HAX-1 is 5′-GCTGAGGACTATGGAACCTT-3′.Plasmid construction: The CRISPR-SaCas9-KKH system was selected for this study because it can be packaged and delivered through a single AAV vector and has known PAM sequences (50-NNNRRT-30) that are compatible with larger sgRNA numbers. sgRNA was inserted into the BsaI site of the pCMV_SaCas9-KKH_hU6 plasmid using PCR and Gibson assembly techniques. Finally, the plasmid was transfected into the CNE2 cells for high-throughput RNA sequencing analysis.

first: Miao

last: Gu

first: Haimeng

last: Yin

first: Si

last: Pan

first: Haijing

last: Xie

first: Wenhui

last: Chen

first: Wenjie

last: Zheng

first: Yiwen

last: You

first: Bo


title: Differences in syncytia formation by SARS-CoV-2 variants modify host chromatin accessibility and cellular senescence via TP53

city: Boston

first: Jonathan

middle: David

last: Lee

state: MA

address: 3 Blackfan Circle, Rm. 424

country: USA

institute: BIDMC

type: Genome binding/occupancy profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Chlorocebus sabaeus

platform_organism:
Homo sapiens
Chlorocebus sabaeus

overall_design: Refer to individual Series


title: Intervention effect and mechanism of Tianhe Zhuifeng liquid extract on rheumatoid arthritis

city: Beijing

first: Liu

middle: xue

last: ting

address: Dongzhimen

institute: China Academy of Chinese Medical Science

summary:
In order to explore the intervention effect of Tianhezhuifeng liquid extract on rheumatoid arthritis, we constructed an animal model of rheumatoid arthritis
We then performed gene expression profiling analysis using data obtained from synovial tissues of typical samples

overall_design: Comparative gene expression profiling analysis of RNA-seq data for rheumatoid arthritis and Tianhe zhuifeng liquid extract improve rheumatoid arthritis

first: Xueting

last: Liu

first: Xia

last: Mao

first: Yanqiong

last: Lin


title: RNA Isolation Methods - Diverse Biofluids

city: La Jolla

first: Louise

middle: C

last: Laurent

state: CA

address: SCRM, 2880 Torrey Pines Scenic Drive

department: Obstetrics, Gynecology, and Reproductive Sciences

institute: UCSD

summary: To evaluate performance of different extra cellular RNA isolation methods in biofluids

sample_organism:
Homo sapiens
Rattus rattus

platform_organism:
Homo sapiens
Rattus rattus

overall_design: Serum and plasma were collected from healthy adult donors, 10 female and 10 male. Three Core exRNA isolation methods (ExoRNeasy (Qiagen), miRNeasy (Qiagen), and miRCURY Biofluids (Exiqon)) were tested in all labs, and 7 methods (Exosomal RNA Purification Kit (Norgen Biotek) w/o Amicon 3K, Exosomal RNA Purification Kit (Norgen Biotek) w/ Amicon 3K, Ultra + miRNeasy, Millipore + miRNeasy, ME Kit (New England Peptide) + miRNeasy, ExoQuick+Seramir (SBI), miRCURY Exosome Isolation + miRCURY Biofluids (Exiqon)) were tested in a subset of labs. CCCM was collected from three cell types in three different labs: human embryonic stem cells (hESCs); primary rat cardiomyocytes (NRVM); and cholangiocarcinoma cells (KMBC). exRNA isolation experiments in triplicate for the supernatants from each cell type. Bile was collected from three patients with cholangiocarcinoma using four methods: ExoRNeasy; Ultra; Millipore; and ExoQuick. exRNA isolation experiments in triplicate for the bile from each donor using five methods: ExoRNeasy; miRNeasy; Exiqon; Ultra; and Millipore. Urine was collected from healthy adult donors, 10 female and 10 male (the same donors as for plasma and serum).exRNA isolation experiments in triplicate for the urine from each donor using five methods: ExoQuick; ExoRNeasy; Homebrew; Millipore; and Ultra . Small RNA sequencing libraries were constructed using the NEB Next small RNA library kit.

first: Srimeenakshi

last: Srinivasan

first: Ashish

last: Yeri

first: Pike

middle: S

last: Cheah

first: Allen

last: Chung

first: Kirsty

last: Danielson

first: Peter

middle: D

last: Hoff

first: Justyna

last: Filant

first: Clara

first: Lucie

first: Rogan

last: Magee

first: Courtney

last: Moeller

first: Venkatesh

middle: L

last: Murthy

first: Parham

last: Nejad

first: Anu

last: Paul

first: Isidore

last: Rigoutsos

first: Rodosthenis

last: Rodosthenous

first: Ravi

middle: V

last: Shah

first: Bridget

last: Simonson

first: Cuong

last: To

first: David

last: Wong

first: Irene

middle: K

last: Yan

first: Xuan

first: Xandra

middle: O

last: Breake?eld

first: George

last: Daaboul

first: Roopali

last: Gandhi

first: Jodi

last: Lapidus

last: Londin

first: Tushar

last: Patel

first: Robert

last: Raffai

first: Anil

last: Sood

middle: P

last: Alexander

first: Saumya

last: Das


title: Differences in syncytia formation by SARS-CoV-2 variants modify host chromatin accessibility and cellular senescence via TP53 [miRNA-Seq]

first: Jonathan

last: Lee

summary: COVID-19 remains a significant public health threat due to the ability of SARS-CoV-2 variants to evade the immune system and cause breakthrough infections. Although pathogenic coronaviruses such as SARS-CoV-2 and MERS-CoV lead to severe respiratory infections, how these viruses affect the chromatin proteomic composition upon infection remains largely uncharacterized. Here we used our recently developed integrative DNA And Protein Tagging (iDAPT) methodology to identify changes in host chromatin accessibility states and chromatin proteomic composition upon infection with pathogenic coronaviruses. SARS-CoV-2 infection induces TP53 stabilization on chromatin, which contributes to its host cytopathic effect. We mapped this TP53 stabilization to the SARS-CoV-2 spike and its propensity to form syncytia, a consequence of cell-cell fusion. Differences in SARS-CoV-2 spike variant-induced syncytia formation modify chromatin accessibility, cellular senescence, and inflammatory cytokine release via TP53. Our findings suggest that differences in syncytia formation alter senescence-associated inflammation, which varies among SARS-CoV-2 variants.

overall_design: Small RNA-seq of VeroE6 or A549ACE2 cell lines expressing SARS-CoV-2 cDNAs.

first: Craig

middle: B

last: Wilen

first: Frank

middle: J

last: Slack


title: Plantain flour - a beneficial material for the organ and transcriptional profile of kidney of diabetic rats

city: Guangzhou

first: jinfeng

last: fu

state: Guangdong

address: Wushan Street

department: School of Food Science and Engineering

institute: South China University of Technology

summary: This study investigated the ameliorative effects of plantain flour on the organs of diabetic rats, and explored the mechanism from the perspective of kidney transcription. Diabetes was induced in rats by streptozotocin (STZ) and a high-sugar and high-fat diet. The alleviation effects and molecular mechanism of diabetes by plantain flour at different doses (1g/kg·Bw, 2g/kg·Bw, and 4g /kg·Bw) were evaluated based on the analysis of biochemical indicators, histological observations, homeostasis model, and kidney transcriptome. The findings demonstrated that the intervention of plantain flour can reduce the level of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and urea (UREA). The pathological study revealed that plantain flour can alleviate the pathological symptoms of fat accumulation in liver and kidney tissue. According to the homeostasis model, plantain flour was able to reduce insulin resistance in diabetic rats. Transcriptome analysis showed that plantain flour improved dyslipidemia by up-regulating Acsl1, down-regulating Fabp4 and Plin1, and reduced kidney injury by up-regulating Slco1a6, Hsd17b2, Cyp2c24, and Cyp2c11 genes. These results suggested that plantain flour could recover the organ damage caused by diabetes and has the potential to be developed into healthy food.

overall_design: Kidney mRNA profiles of diabetic rats and diabetic rats treated with plantain flour at different doses (1g/kg•Bw, 2g/kg•Bw, and 4g /kg•Bw) were generated by deep sequencing, in triplicate, using

first: Jinfeng

last: Fu

first: Shiyun

last: Tu

first: Juan

last: Wang

first: Ou

last: Sheng


title: Sequential Exposure to Interleukin-21 and Interleukin-15 During Human Natural Killer Cell Expansion Optimizes Yield and Function

city: Philadelphia

first: Caimei

last: Zhang

state: PA

address: 3400 Civic Center Boulevard Atrium, Ground Floor

department: Center for Cellular Immunotherapies

institute: University of Pennsylvania

summary: NK cells are frequently expanded for the clinic using irradiated engineered K562 feeder cells expressing a core transgene set of membrane-bound mb IL-15 and/or mbIL-21 together with 41BBL. Prior comparisons of mbIL-15 to mbIL-21 for NK expansion lack comparisons of key attributes of the resulting NK cells including their high dimensional phenotype, polyfunctionality, the breadth and potency of cytotoxicity, cellular metabolism, and activity in xenograft tumor models. Moreover, despite multiple rounds of K562 stimulation, studies of sequential use of mbIL-15- and mbIL-21-based feeder cells are absent. We addressed these gaps and found that using mbIL-15 versus mbIL-21-based feeder cells drove distinct phenotypic and functional profiles. Feeder cells expressing mbIL-15 alone drove superior functionality by nearly all measures, while those expressing mbIL-21 alone drove superior yield. In combination, most attributes resembled those imparted by mbIL-21, whereas in sequence NK yield approximated that imparted by the first cytokine, and the phenotype, transcriptome, and function resembled that driven by the second cytokine highlighting the plasticity of NK cell differentiation. The sequence mbIL-21 followed by mbIL-15 was especially advantageous in achieving significant yields of highly functional NK cells that demonstrated equivalent in vivo activity to those expanded by mbIL-15 alone in 2 of 3 xenograft models. Our findings define the impact of mbIL-15 versus mbIL-21 during NK expansion and reveal a previously under appreciated tradeoff between NK yield and function for which sequential use of mbIL-21- followed by mbIL-15-based feeder cells may be the optimal approach in many settings.

overall_design: To characterize NK cells expanded with different K562 feeders expressing mbIL15 or/and mbIL21 and optimize the NK expansion process for producing sufficient functional NK cells, We co-cultured pure NK cells with irradiated K562-41BBL-CD40-mbIL15 or K562-41BBL-CD40-mbIL21 or K562-41BBL-CD40-mbIL15-mbIL21 or sequential K562-41BBL-CD40-mbIL15 and K562-41BBL-CD40-mbIL21. On day 20 of the expansion, NK cell yield, phenotype, functionality, metabolism, and transcriptional expression were assessed.

first: Siddhant

last: Kadu

first: Yansen

last: Xiao

first: Omar

last: Johnson

first: Linhui

last: Chen

first: James

last: Riley

first: Roddy

last: O’Connor

first: Carl

middle: H

last: June

first: Neil

last: Sheppard


title: Hiltonol and pRNA stimuli induce an immune activating profile in the transcriptomics of cDC1s

city: Nijmegen

first: Johannes

last: Textor

address: Geert Groteplein 26-28

institute: Radboud University Medical Center


title: The apocarotenoid β-ionone regulates the transcriptome of Arabidopsis thaliana and increases its resistance against Botrytis cinerea

city: Thuwal

first: Juan Camilo

last: Moreno Beltran

address: King Abdullah University of Science and Technology

country: Saudi Arabia

institute: KAUST

summary: Carotenoids are isoprenoid pigments vital for photosynthesis. Moreover, they are the precursor of apocarotenoids including the phytohormones abscisic acid (ABA) and strigolactones (SLs), and retrograde signaling molecules and growth regulators (e.g., β-cyclocitral and zaxinone). The apocarotenoid β-ionone (β-I) was previously reported to exert antimicrobial effects, however, at very high millimolar concentrations. Here, we showed that the application of this volatile to Arabidopsis plants at micromolar concentrations caused a global reprogramming of gene expression, affecting thousands of transcripts involved in stress tolerance, growth, hormone metabolism, pathogen defense and photosynthesis. These changes, along with modulating the levels of the phytohormones ABA, jasmonic acid and salicylic acid, led to enhanced Arabidopsis resistance to Botrytis cinerea (B.c.); one of the most aggressive and widespread pathogenic fungi affecting numerous plant hosts and causing severe losses of post-harvested fruits. Pre-treatment of tobacco and tomato plants with β-I followed by inoculation with B.c. confirms the conserved effect of β-I in crop plants. Moreover, there was reduced susceptibility to B.c. in LYCOPENE β-CYCLASE-expressing tomato fruits possessing elevated levels of endogenous β-I, indicating beneficial biological activities of this compound in planta. Our work unraveled β-I as a further carotenoid-derived regulatory metabolite and opens up new possibilities to control B.c. infection by establishing this natural volatile as an environmentally friendly bio-fungicide.

overall_design: To investigate the function of b-ionone we treated Arabidopsis plants by spraying this volatile compound on the plants and then they were kept in a closed environment. Leaf samples were taken at different time points. The same approach was used for pathogen infection of the plants with the fungus botrytis cinerea and leaf samples were collected at 24 hours post infection. We also combined both treatments and spray the plants with b-ionone for 24 hours followed by botrytis cinerea infection and take samples at 24 and 48 hours post infection.

first: Abrar

last: Felemban

last: Moreno

first: Jianing

last: Mi

first: Shawkat

last: Ali

first: Arjun

last: Sham

first: Synan

middle: F

last: AbuQamar

first: Salim

last: Al-Babili


title: Differences in syncytia formation by SARS-CoV-2 variants modify host chromatin accessibility and cellular senescence via TP53 [RNA-Seq]

first: Jonathan

last: Lee

overall_design: RNA-seq of VeroE6 or A549ACE2 cell lines expressing SARS-CoV-2 cDNAs.

first: Craig

last: Wilen

first: Frank

last: Slack


title: Longitudinal gene expression profiling of self-collected blood samples in COVID-19+ and healthy participants

city: Seattle

first: Fang Yun

last: Lim

state: WA

address: 1100 Eastlake Ave E

department: Vaccine and Infectious Disease Division

institute: Fred Hutchinson Cancer Center

summary:
Longitudinal cohort: 773 host response genes were profiled in previously vaccinated (n=16) and unvaccinated (n=14) COVID-19+ participants along with 5 healthy uninfected controls across a 2-week observational window
Single timepoint cohort: 773 host response genes were profiled in 6 healthy uninfected participants

overall_design:
Longitudinal cohort: 30 COVID-19+ and 5 uninfected participants were asked perform self-collection and stabilization of capillary blood using a novel technology (homeRNA) every other day for two weeks (7 longtiudinal timepoints per participant). Temporal kinetics of 773 immune genes were profiled using the nCounter direct digital counting of native mRNA.
Single timepoint cohort: 6 healthy uninfected participants were asked perform self-collection and stabilization of capillary blood using a novel technology (homeRNA). Temporal kinetics of 773 immune genes were profiled using the nCounter direct digital counting of native mRNA.

first: Fang


title: Hiltonol and pRNA stimuli induce an immune activating profile in the transcriptomics of cDC1s [Affy]

summary: Stimulation of dendritic cells (DC) is considered critical in cancer immunotherapy. BATF-3–dependent subsets, that express in humans BDCA-3 (CD141), promote CD8 T-cell cross-priming against tumor antigens. Here, we evaluate two clinical grade stimuli for peripheral blood cDC1 myeloid dendritic cells (mDCs), a rare DC subset that is currently being explored for use in immunotherapy. In contrast to routine evaluation methods, which focus on predefined maturation markers on the surface or factors released from the activated cells, we applied an unbiased transcriptome-based method using both RNA-sequencing (RNA-seq) and microarrays. Specifically, we analyzed the mRNA of cDC1 mDCs from five human donors upon activation with two clinical-grade adjuvants, Hiltonol (poly IC, a TLR3 ligand, called Stimulus 1 in the description below) and protamine RNA (pRNA, aTLR7/8 ligand, Stimulus 2), and compared these samples to unstimulated counterparts. Both methods, RNA-seq and microarray, gave very similar results and showed that Hiltonol and pRNA lead to almost identical changes in the transcriptome of cDC1 mDCs. A gene ontology (GO) term analysis suggested that these changes were mainly related to activation and maturation pathways, including induction of type I IFN and IL-12 transcription, while pathways related to adverse effects or cell damage were not strongly affected. The combination of both reagents in the DC cultures gave a very similar result as compared to either stimulus alone, suggesting no synergistic effect. Together, our results indicate that both stimuli are potent clinical grade adjuvants with comparable effects to mature cDC1 mDCs within an immunogenic profile in short-term cultures suitable to be used in immunotherapy.

overall_design: BDCA3 were isolated from blood from 5 donors, 2 treatments were tested separately or in combination (combination not administered to donor 2), an unstimulated control was taken along. Due to limited cell numbers, only microarray analysis was performed for Donor 1.

first: Georgina

last: Florez-Grau

first: Till

last: Mathan

first: Mihaela

last: Mihaylova

first: Tom

last: van Oorschot

first: Sonja

middle: I

last: Buschow

first: Gerty

last: Schreibelt

first: Inge

last: Reinieren-Beeren

first: David

last: Sancho

first: Ignacio

last: Melero

middle: G

last: Figdor

first: Jolanda

middle: M

last: de Vries I


title: The GPR84 molecule is a mediator of a subpopulation of retinal microglia that promote TNF/IL-1α expression via the rho-ROCK pathway after optic nerve injury (bulk RNA-Seq)

first: Kota

last: Sato

overall_design: Examination of 4 different timepoint in Iba1-positive retinal cells under optic nerve injury in mice

first: Michiko

last: Ohno-Oishi

first: Masaaki

last: Yoshida

first: Taimu

first: Takaharu

last: Aizawa

first: Yuto

last: Sasaki

first: Shigeto

last: Maekawa

first: Makoto

last: Ishikawa

first: Kazuko

last: Omodaka

first: Chihiro

last: Kawano

first: Ryuji

last: Ohue-Kitano

first: Ikuo

last: Kimura

first: Toru

last: Nakazawa


title: Differences in syncytia formation by SARS-CoV-2 variants modify host chromatin accessibility and cellular senescence via TP53 [ATAC-Seq]

first: Jonathan

middle: David

last: Lee

overall_design: ATAC-seq of VeroE6 or A549ACE2 cell lines, either infected with coronaviruses or expressing SARS-CoV-2 cDNAs.

first: Craig

last: Wilen

first: Frank

last: Slack


title: Oleic Acid Availability Preprograms Early Thymocytes for Treg Cell Commitment in the Periphery [Rna-seq]

city: Shanghai

first: Mingyuan

last: Hu

address: 320, Yueyang Road

institute: Chinese Academy of Science

summary: The natures of activation signals are essential in determining T cell subset differentiation, however, the properties of T cell subset preference acquired during intrathymic development remain elusive. Here we show that naïve CD4+ T cells generated in the thymic microenvironment lacking SCD1, the enzyme catalyzing oleic acid production, exhibit enhanced Treg cell differentiation and confer on mice the ability to resist EAE. SCD1 deletion in K14+ thymic epithelia recapitulated the enhanced Treg cell differentiation phenotype of SCD1 knockout mice. The dearth of oleic acid permitted Dot1L to increase H3K79me2 level at the Atp2a2 locus of thymocytes at the DN2-DN3 transition stage. Such epigenetic modification persisted to naïve CD4+ T cells and facilitated Atp2a2 expression. Upon TCR activation, Atp2a2 enhanced the activity of calcium-NFAT1-Foxp3 axis to promote naïve CD4+ T cells to differentiate into Treg cells. Therefore, oleic acid availability is critical for preprogramming thymocytes with Treg differentiation propensities in periphery.

overall_design: RNA-seq data of cTECs (Ly51+ UEA1−) and mTECs (Ly51− UEA1+) derived from SCD1 knockout mice and littermate control mice

first: Liangyu

last: Lin


title: LncRNA DCRT protects against dilated cardiomyopathy via binding to PTBP1 (RIP-seq)

city: WUHAN

first: heng

middle: zhi

last: du

address: 1095 Jiefang Dadao, Wuhan City, Hubei Province, China

institute: Wuhan Tongji Hospital

summary: RNA immunoprecipitation sequencing (RIP seq) using PTBP1 antibody to identify the potential targets of in DCRT-KO and WT cell

overall_design: We report the application of sequencing technology for high-throughput profiling of mRNA expressions in DCRT-KO and WT cells. DCRT-KO and WT cells were lysed and then incubated with anti-PTBP1 antibody . After incubating at 4 °C overnight, protein A/G magnetic beads (Thermo Scientific, Shanghai, China) were added into the solution and incubated for another 2 h at 4 °C. Then, the beads were washed five times with PBST and the remaining products were extracted with TRIzol agent. In this project, rRNA was removed from the total RNA by ribosomal RNA removal kit, and the way of ion breaking were used to break the RNA into 200-300bp fragments. By using RNA as template, the first strand of cDNA was synthesized with 6 base random primers and reverse transcriptase. The first strand cDNA was used as the template to synthesize the second strand cDNA. When the second strand cDNA is synthesized, the base T is replaced by U to achieve the goal of a chain specific library. All of our RNA-seq projects used strand specific kits to construct the libraries. Finally, we show that mRNA different expression in AC16 cells. This study provides abundance of mRNA that may predict heart diseases.

first: Hengzhi

last: Du

first: Yanru

last: Zhao


title: The GPR84 molecule is a mediator of a subpopulation of retinal microglia that promote TNF/IL-1α expression via the rho-ROCK pathway after optic nerve injury

first: Kota

last: Sato

overall_design: Examination of 3 different timepoint in Iba1-positive retinal cells.

first: Michiko

last: Ohno-Oishi

first: Masaaki

last: Yoshida

first: Taimu

first: Takaharu

last: Aizawa


title: Nrp1 downregulation correlates with enhanced ILC2 responses during IL-33 challenge

city: Tianjin

first: Jie

last: Zhou

address: No22.Qixiangtai Rd.,Heping Dist,Tianjin P.R. China

institute: Tianjin Medical University

summary: Group 2 innate lymphoid cells (ILC2s) play critical roles in driving the pathogenesis of allergic airway inflammation. The mechanisms underlying the regulation of ILC2s remain to be fully understood. Here, we identified neuropilin-1 (Nrp1) as a surface marker of ILC2s in response to IL-33 stimulation. Nrp1 was abundantly expressed in ILC2s from lung under steady state, which was significantly reduced upon IL-33 stimulation. ILC2s with high expression of Nrp1 (Nrp1high) displayed lower response to IL-33, as compared with Nrp1low ILC2s. Transcriptional profiling and flow cytometric analysis showed that downregulation of AKT-mTOR signaling participated in the diminished functionality of Nrp1high ILC2s. These observations revealed a potential role of Nrp1 in ILC2s responses under allergic inflammatory condition.

overall_design: To evaluate the transcriptional differences between Nrp1high and Nrp1low ILC2s , we sorted  Nrp1high and Nrp1low ILC2s from lungs at IL-33 treatment condition. We then performed gene expression by KEGG analysis, Heat map analysis and volcano plot analysis

first: Ying

last: Wang

first: Gaoyu

last: Liu


title: Single-cell RNA sequencing to reveal non-parenchymal cell heterogeneity and immune network of acetaminophen-induced liver injury in mice

first: Li

last: Xiaoyun

state: other

address: china

institute: Gastroenteroloy

summary: The role of non-parenchymal cells (NPCs) in the early phase of acetaminophen (APAP)-induced liver injury (AILI) remains unclear. Therefore, single-cell sequencing (scRNA-seq) was performed to explore the heterogeneity and immune network of NPCs in the livers of mice with AILI.  Mice were challenged with saline, 300 mg/kg APAP, or 750 mg/kg APAP (n=3 for each group). After 3 h, the liver samples were collected, digested, and subjected to scRNA-seq. Immunohistochemistry and immunofluorescence were performed to confirm the expression of Makorin ring finger protein 1 (Mkrn1).  We identified 14 distinct cell subtypes among the 120,599 cells. A variety of NPCs were involved even in the early stages of AILI, indicating highly heterogeneous transcriptome dynamics. Cholangiocyte cluster 3 with high Deleted in malignant brain tumors 1 (Dmbt1) expression was found to perform drug metabolism and detoxification functions. Liver sinusoidal endothelial cells displayed fenestrae loss and angiogenesis. Macrophage cluster 1 displayed an M1 polarization phenotype, whereas cluster 3 tended to exhibit M2 polarization. Kupffer cells (KCs) particularly highly expressed Cxcl2 exhibiting pro-inflammatory effects. qRT-PCR and western blotting verified that LIFR-OSM axis might promote the activation of MAPK signaling pathway in RAW264.7 macrophages. Mkrn1 was highly expressed in the liver macrophages of AILI mice and patients with AILI. Interaction patterns between macrophages/KCs and other NPCs were complex and diverse.  NPCs were highly heterogeneous and involved in the immune network in the early phase of AILI. In addition, we proposed that Mkrn1 may serve as a potential biomarker of AILI.

first: Xiaoyun

last: Li

first: Yang

last: Zhi

first: Jing

first: Xiaohong

last: Lei

first: Yi

last: Ju

first: Yuting

last: Zhang

first: Yufan

last: Zheng

first: Xiaming

last: Kong

first: Feng

last: Xue

first: Wei

last: Zhong

first: Xiaoyu

last: Chen

first: Jieting

last: Tang

first: Xiaobo

first: Yimin

last: Mao


title: The effects of atorvastatin (AVS) and afidopyropen on developing zebrafish

city: Gainesville

first: Christopher

last: Martyniuk

state: FL

address: 8891 SW 79th Avenue

institute: University of Florida

summary: For RNA-seq experiments, 20 embryos were placed into 5 mL final volume of sterile ERM. Each treatment group were exposed within a 10-minute window. Exposure groups included ERM, [100 ng/L AVS or 1 µg/L AVS] or [100 ng/L or  1 µg/L afidopyropen] (N=5 beakers per treatment). On 7 days post fertilization (dpf), fish were transferred to a labeled and pre-weighed 1.7 mL tube. The tube was then immediately flash frozen in liquid nitrogen prior to RNA-seq analysis.

overall_design: Exposure groups included 5 treatments:  ERM, [100 ng/L AVS or 1 µg/L AVS] or [100 ng/L or  1 µg/L afidopyropen] Four replicates out of the 5 were ultimately selected for RNA-seq.

first: Chris

last: Souders II

first: Ciara

last: Saccente

first: Cole

last: English


title: Time of administration and cryptochrome-dependent changes in tolerability and pharmacokinetics of oxaliplatin

city: Istanbul

first: Halil

last: Kavakli

address: Rumelifeneri Yolu, Sariyer

department: Chemical and Biological Engineering

country: Turkey

institute: Koc University

summary: The circadian clock controls many physiological functions in mammals, including responses to cancer therapy, by influencing the pharmacokinetics, efficacy, and toxicity of anticancer drugs. Our objective was to investigate how the timing of administration, as well as the influence of cryptochrome (Cry), a key gene in the circadian clock, impact oxaliplatin's pharmacokinetics and toxicity. Additionally, we aimed to shed light on the underlying mechanism through RNA sequencing.

overall_design:
Cryptochrome (Cry) 1/2 heterozygous mice in C57BL/6J background were generously provided by Aziz Sancar (University of North Carolina, Chapel Hill, North Carolina, USA) and used to obtain Cryptochrome double-knockout (Cry1-/-Cry2-/-, DKO) mice. Animals were kept in Koc University Research Center for Translational Medicine (KUTTAM) and genotyping was done to determine the knockouts. Animals (12-16 weeks old) were housed under 12 h of light- 12 h of dark cycle (LD 12:12) and synchronized for 3 weeks prior to the beginning of experiments. Mice were kept in constant darkness (DD) throughout the experiments, starting 3 days before drug administration
A single dose of oxaliplatin (12 mg/kg, i.p.) was administered to wild-type and CRYDKO mice over two circadian times (CT8/ CT16, Circadian Time). Drug-induced toxicity; assessed by death rate and weight loss. Oxaliplatin pharmacokinetics were studied by ICP-MS. 12 mg/kg i.p. for three days. Following administration of oxaliplatin, RNA obtained from liver of mice was sequenced.
Total RNA from liver samples (WT and CRYDKO mice treated with oxaliplatin at CT8 and CT16) were isolated by using RNeasy Mini Kit (QIAGEN, Germany). Quality of amount of RNA were assessed by using Qubit (Thermo Fisher Scientific, USA). High quality RNA samples (OD260/280 1.8-2.0) were used to prepare library for RNA Sequencing analysis. Briefly, adenylated mRNAs were isolated by using Poly(A) mRNA Magnetic Isolation Module (New England Biolabs, US). Then mRNAs were fragmented, converted to cDNA and second strand synthesis was performed. The 150bp paired-end sequencing of libraries were carried out via Illumina platform (Macrogen, Netherlands).

first: Seref

last: Gul

first: Yasemin

last: Akyel

first: Alper

last: Okyar


title: Transcriptome profiling of Phalaenopsis reveals that photoperiod regulates floral induction

city: Hangzhou

first: Wang

last: Shuang

address: 666 Wusujie Linan

institute: Zhejiang A & F University

summary: Floral induction; RNA-seq; Differentially transcribed Genes; WGSCN; qRT-PCR

overall_design: The topmost leaves of the Phalaenopsis orchid were pooled at 0 d, 20 d, and 40 d, with three biological replicates at each treatment, each biological replicate consisting of 3 plants.120 phalaenopsis plants were divided equally into two groups of long  photoperiod treatment and short photoperiod treatment. Young leaves were extracted for RNA sequencing.

first: Shuang

last: Wang

first: Ping

last: Luo

first: Jingjing

last: Xin

first: Yongyi

last: Cui


title: Gene expression and functional analysis of GJB2 mutated HaCaT cell lines

city: Jinan

first: Li

last: Zhang

address: Department of Dermatology, Shandong Provincial Hospital, Cheeloo College of Medicine, Shandong University, Jinan, Shandong 250021, China.

overall_design: Refer to individual Series


first: Hengyu

last: Du

address: Huangpu Road West 601

institute: Department of Plastic Surgery of the First Affiliated Hospital of Jinan University

summary: Human dermal microvascular endothelial cells (HDMECs) were isolated via the magnetic-activated cell sorting (MACS) method from skin biopsies of patients with DFUs and healthy controls. We then performed gene expression profiling analysis using data obtained from RNA-seq of these cells.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HDMECs isolated from skin biopsies of patients with DFUs and healthy controls

first: Hongwei

last: Liu


title: Hiltonol and pRNA stimuli induce an immune activating profile in the transcriptomics of cDC1s [RNA-seq]

first: Johannes

last: Textor

summary: Stimulation of dendritic cells (DC) is considered critical in cancer immunotherapy. BATF-3–dependent subsets, that express in humans BDCA-3 (CD141), promote CD8 T-cell cross-priming against tumor antigens. Here, we evaluate two clinical grade stimuli for peripheral blood cDC1 myeloid dendritic cells (mDCs), a rare DC subset that is currently being explored for use in immunotherapy. In contrast to routine evaluation methods, which focus on predefined maturation markers on the surface or factors released from the activated cells, we applied an unbiased transcriptome-based method using both RNA-sequencing (RNA-seq) and microarrays. Specifically, we analyzed the mRNA of cDC1 mDCs from five human donors upon activation with two clinical-grade adjuvants, Hiltonol (poly IC, a TLR3 ligand, called "Stimulus 1" in the description below) and protamine RNA (pRNA, aTLR7/8 ligand, "Stimulus 2"), and compared these samples to unstimulated counterparts. Both methods, RNA-seq and microarray, gave very similar results and showed that Hiltonol and pRNA lead to almost identical changes in the transcriptome of cDC1 mDCs. A gene ontology (GO) term analysis suggested that these changes were mainly related to activation and maturation pathways, including induction of type I IFN and IL-12 transcription, while pathways related to adverse effects or cell damage were not strongly affected. The combination of both reagents in the DC cultures gave a very similar result as compared to either stimulus alone, suggesting no synergistic effect. Together, our results indicate that both stimuli are potent clinical grade adjuvants with comparable effects to mature cDC1 mDCs within an immunogenic profile in short-term cultures suitable to be used in immunotherapy.

first: Georgina

last: Florez-Grau

first: Till

last: Mathan

first: Mihaela

last: Mihaylova

first: Tom

last: van Oorschot

first: Sonja

last: Buschow

first: Gerty

last: Schreibelt

first: Inge

last: Reinieren-Beeren

first: David

last: Sancho

first: Ignacio

last: Melero

first: Carl

last: Figdor

first: Jolanda

last: de Vries I


title: Effect of FTY720 on gene expression in KMT2A-R AML cell line THP1

city: London

first: Maria Teresa

last: Esposito

address: Holybourne Avenue

department: School of life and health

country: United Kingdom

institute: Centre for integrated research in  life and health

summary: KMT2A-rearranged (KMT2A-R)- leukemia is a distinct and rare form of leukemia which mostly affects children. Despite extensive research, KMT2A-R-patients have a dismal prognosis and no targeted therapy is yet available in the clinics. SET is a potent oncoprotein and an endogenous inhibitor of the phosphatase PP2A. In this study we show that SET is specifically enriched in KMT2A-R-leukemic stem cells (LSCs) and it is essential for the self-renewal of KMT2A-R-cells. Treatment with SET-PP2A inhibitor FTY720 disrupted SET-PP2A interaction leading to cell cycle arrest, cellular death and increased sensitivity to chemotherapy treatment in KMT2A-R-models.  Genetic and pharmacological inhibition of SET via FTY720 led to down-regulation in the expression of several genes belonging to the KMT2A-R-leukemia signature, including MYC, HOXA10 and HOXA9/MEIS1 target genes.  Taken together these results demonstrated that SET represents an interesting and novel  player of KMT2A-R-leukemia and its antagonism through FTY720 could serve as a novel strategy to treat this disease.

overall_design: We treated the KMT2A-R cell line THP1 with either vehicle (DMSO) or FTY720 (5uM) for 24hr and collected RNA for RNAseq. Every sample was run in triplicate.

first: Maria

middle: T


title: Gene expression and functional analysis of GJB2 mutated HaCaT cell lines [D66H]

first: Li

institute: Department of Dermatology, Shandong Provincial Hospital, Cheeloo College of Medicine, Shandong University, Jinan

summary:
Vohwinkel syndrome, VS (OMIM#124500), a rare autosomal dominant genetic disease, with less than 50 reported cases in the literature. Although clinical symptoms of VS are complex, which are caused by GJB2 mutation is more typical.
To explore related differential genes and signaling pathways of Vohwinkel syndrome (VS) caused by mutations of GJB2. Human Gene Expression Array of two types of GJB2-VS mutated (G130V and D66H) HaCaT cell lines and identified distinct classes of up- and down- regulated genes during this process.

overall_design: Using GJB2 gene mutant p.Asp66His(D66H) primers as templates,  the corresponding cDNA sequence of GJB2: NM_004004 (D66H) was amplified respectively, and then was ligated with GV308 vector. The recombinant plasmids D66H-GV308, and empty vector GV308 were identified and sequenced. Then GJB2-D66H-overexpression lentiviruses. Human Gene Expression Array of two types of GJB2-VS mutated (D66H) and NC HaCaT cell lines for RNA extraction and hybridization on Affymetrix microarrays. And combined Ingenuity Pathway Analysis, combined apoptosis experiment, mRNA expression of LOR, FLG, and differentially expressed genes.

first: Keying

last: GUO


title: The PENGUIN approach to reconstruct  protein interactions at enhancer-promoter regions and its application to prostate cancer

city: Badalona

first: François

middle: José

last: Serra

address: Ctra de Can Ruti, Camí de les Escoles s/n

country: Spain

institute: Institut de Recerca Contra la Leucèmia Josep Carreras-IJC

summary:
We introduce the Promoter-ENhancer-GUided Interaction Networks (PENGUIN) approach to identify protein-protein interactions (PPI) within enhancer-promoter (E-P) interactions. By integrating high-coverage H3K27ac-HiChIP data and tissue-specific PPI networks, PENGUIN identifies functional clusters in E-P networks.
Here, we applied PENGUIN to E-P networks of  prostate cancer (PrCa) cell line LNCaP. We validated PENGUIN's structural classification by observing clear differential enrichment of the architectural protein CTCF. One of our 8 main clusters, comprising 273 promoters, showed significant enrichment for PrCa-associated single nucleotide polymorphisms (SNPs) and oncogenes. Our approach provides a mechanistic explanation for 208 PrCa SNPs located within DNA-binding protein (DBP) binding sites or intermediate protein-encoding genes involved in E-P contacts. CRISPR analysis in the LNCaP cell line confirmed the relevance of these SNPs in PrCa.
PENGUIN confirms the importance of key regulators in PrCa and identifies new intervention candidates, offering new directions for identifying molecular targets in disease treatment.
Data was generated in the Matthew L. Freedman lab.

overall_design: H3K27ac HiChIP in LNCaP. Chromatin digested with MboI restriction enzyme. H3K27ac ChIP-seq ??

first: Ji-Heui

last: Seo

first: Alexandros

last: Armaos

first: Iker

last: Núñez-Carpintero

first: Sylvan

middle: C

last: Baca

first: Stefano

last: Gustincich

first: Alfonso

last: Valencia

first: Davide

last: Cirillo

first: Claudia

last: Giambartolomei

first: Gian

last: Gaetano Tartaglia

first: Matthew

last: Freedman


title: miR profile in pagetic osteoclasts: from large-scale sequencing to gene expression study

city: Sherbrooke

first: Sophie

last: Roux

state: Quebec

address: 3001, 12th avenue N

department: Division of Rheumatology

country: Canada

institute: Université de Sherbrooke

summary:
Background: Paget's Disease of Bone (PDB) is characterized by excessive and disorganized bone remodeling, in which bone-resorbing osteoclasts play a key role. Strong genetic components and environmental factors contribute to the etiopathogenesis of PDB. We hypothesized that dysregulated microRNA (miR) expression contributed to the osteoclast phenotype in PDB.
Results: From deep sequencing, six mature miRs, miR-29b1-3p, miR-15b-5p, miR-181a-5p, let7i-3p, miR-500b-5p, and miR-1246, were found to be significantly decreased in overactive osteoclasts from the PDB patients compared to those in the healthy controls. The differential expression of the miRs was confirmed by the analysis of a larger independent cohort using qPCR.
Conclusion: We identified six miRNAs whose expression were deregulated in PDB condition. Many of these miRs are known to modulate many critical aspects of osteoclast generation and function by coordinating their regulation of gene expression. Our results enhance the understanding of osteoclast biology in PDB disease

overall_design: Blood samples were obtained from 9 patients with PDB and  9 sex- and age- matched controls. Peripheral blood mononuclear cells (PBMCs) were differentiated into osteoclasts in 3-week in-vitro cultures and osteoclast samples were sent to be sequenced using multiplex sequencing on a HiSeq (Illumina) instrument (Genome Quebec).

first: Hoang Dong

last: Nguyen

first: Martine

last: Bisson

first: Michelle

middle: S

last: Scott

first: Gilles

last: Boire

first: Luigi

last: Bouchard


title: Transcriptome of 5-week-old Col-0 and isi1-2 distal leaves 13 in untreated controls or upon wounding leave 8

city: Lausanne

first: Karolina

last: Bojkowska

address: Unil, Genopode

country: Switzerland

institute: University of Lausanne

summary: In our study, IMPAIRED SUCROSE INDUCTION 1 (ISI1) was identified as a player in long distance wound signaling in Arabidopsis. To further assess the impact of ISI1 on the wound response, we tested the transcriptomes of the wounded isi1-2 mutants in comparison with wounded wild type plants, in order to find genes that were deregulated in isi1 loss-of-function mutants.

overall_design: Two groups of plants: control (Col-0) and isi-mutant plants (isi1-2) with three biological replicates per group; Distal leaves 13 were collected one hour after wounding leave 8 (in wounded plants) or without any treatment (unwounded plants). Two individual plants were pooled for one replicate; single read design.

first: Qian

last: Wu

first: Stéphanie

last: Stolz

first: Archana

last: Kumaria

first: Gaétan

last: Glauser

first: Edward

middle: E

last: Farmer


title: Acetylation of EED by CBP/P300 Regulates the Targeting of PRC2 and is Required for mESC Differentiation

city: Macau

first: Gang

last: Li

address: E12 Avenida da Universidade, Taipa, Macau

department: Faculty of Health Sciences

country: Macau

institute: University of Macau

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


title: Acetylation of EED by CBP/P300 Regulates the Targeting of PRC2 and is Required for mESC Differentiation [ChIP-seq]

summary: Polycomb repressive complex 2 (PRC2) is a chromatin-modifying enzyme that catalyzes the methylation of lysine 27 on histone H3 (H3K27me1/2/3). This complex maintains the gene expression profiles in different cell types and plays an essential role in normal organismal development. The mechanisms by which PRC2 targets specific chromatin regions remain unclear. To address this question, we focused on the post-translational modifications (PTMs) of PRC2 subunits. Here we show EED, a core component of PRC2, is acetylated by acetyltransferase CBP/P300 at lysine 19 (K19). The acetylation of EED at K19 (EED-K19ac) reduces the binding affinity between PRC2 and native nucleosomes, causing PRC2 to leave its chromatin targets in vivo. Genome-wide location analysis (ChIP-seq) reveals that K19-acetylated EED preferentially accumulates at highly transcribed genes with low EZH2 and H3K27me3 levels. Using CRISPR/Cas9 technology, we generated mouse embryonic stem cells (mESCs) carrying non-acetylated EED mimic (EED-K19R) and showed acetylation of EED at K19 is necessary for mESC differentiation. In summary, our study reveals a novel mechanism that allows intracellular signalling pathways to regulate cellular differentiation and cell fate decisions by controlling the genomic targeting of PRC2 through acetylating EED.

overall_design: ChIP sequencing of wildtype mESCs, EED knockout mutant mESCs, FLAG-EED wildtype and FLAG-EED K19R mutant knock-in mESCs with or without TSA treatment.

first: Yiqi

last: Yang

first: Yunze

last: Liu


title: Oleic Acid Availability Preprograms Early Thymocytes for Treg Cell Commitment in the Periphery

status: Public on Sep 29 2023

first: Mingyuan

last: Hu

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modification H3K79me2 in in vitro developed DN3 thymocytes  with or without SCD1 inhibitor treatment.

first: Liangyu

last: Lin


title: IPSC derived cardiac fibroblasts of DMD patients show compromised actin microfilaments, metabolic shift and pro-fibrotic phenotype.

city: Toulouse

first: Nathalie

last: Pizzinat

address: 1 avenue du Pr Jean Poulhès

country: France

institute: INSERM

summary: Duchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy caused by mutations in the dystrophin gene. We characterized which isoforms of dystrophin were expressed by human induced pluripotent stem cell (hiPSC)-derived cardiac fibroblasts obtained from control and DMD patients. Distinct dystrophin isoforms were observed; however, the highest molecular weight isoform was absent in DMD patients carrying exon deletions or mutations in the dystrophin gene. The loss of the full-length dystrophin isoform in hiPSC-derived cardiac fibroblasts from DMD patients resulted in deficient formation of actin microfilaments and a metabolic switch from mitochondrial oxidation to glycolysis. The DMD hiPSC-derived cardiac fibroblasts exhibited a dysregulated mitochondria network and reduced mitochondrial respiration, with enhanced compensatory glycolysis to sustain cellular ATP production. This metabolic remodeling was associated with an exacerbated myofibroblast phenotype and increased fibroblast activation in response to pro fibrotic challenges. As cardiac fibrosis is a critical pathological feature of the DMD heart, the myofibroblast phenotype induced by the absence of dystophin may contribute to deterioration in cardiac function. Our study highlights therelationship between cytoskeletal dynamics, metabolism of the cell and myofibroblast differentiation and provides a new mechanism by which inactivation of dystrophin in non-cardiomyocyte cells may increase the severity of cardiopathy.

overall_design: In order to investigate the effect of dystrophin loss on cell metabolism, we generated hiPSC-derived cardiac fibroblasts from  control (CON) and DMD patients and performed gene expression profiling  analysis

first: Salwa

last: Soussi

first: Lesia

last: Savchenko

first: Davide

last: Rovina

first: Jason

last: Iacovoni

first: Andrea

last: Gottinger

first: Maxime

last: Viallettes

first: Josè-Manuel

last: Pioner

last: Farini

first: Sara

last: Mallia

first: Martina

last: Rabino

first: Giulio

last: Pompilio

first: Angelo

last: Parini

first: Olivier

last: Lairez

first: Aoife

last: Gowran


title: RNA-seq of polarized macrophages from wild-type and specifically delete FBXO38 in myeloid cells

city: shanghai

first: jiang

last: qi

address: No. 320 Yueyang Road, Xuhui District

institute: university of chinese academy of sciences

summary: We compare transcriptomes between macrophages polarized with IL-4 + IL-13 in the presence or absence of Fbxo38.

overall_design: PEMs were isolated from three pairs of WT and cKO littermate mice and stimulated with IL-4/IL-13 (20 ng/mL) for 24 hours.


title: Genetic variants that impact alternative polyadenylation in cancer represent candidate causal risk loci [RNA-seq]

city: Wuhan

first: Bin

last: Li

address: Donghu Road 115

institute: Department of Epidemiology and Biostatistics, School of Public Health; Wuhan University

summary: Alternative polyadenylation (APA) is emerging as a major mechanism of post-transcriptional regulation. APA can impact the development and progression of cancer, suggesting that the genetic determinants of APA might play an important role in regulating cancer risk. Here, we depicted a pan-cancer atlas of human APA quantitative trait loci (apaQTLs), containing approximately 0.7 million apaQTLs across 32 cancer types. Systematic multi-omics analyses indicated that cancer apaQTLs could contribute to APA regulation by altering poly(A) motifs, RNA binding proteins (RBP), and chromatin regulatory elements and were preferentially enriched in GWASidentified cancer susceptibility loci. Moreover, apaQTL-related genes (aGenes) were broadly related to cancer signaling pathways, high mutational burden, immune infiltration, and drug response, implicating their potential as therapeutic targets. Furthermore, apaQTLs were mapped in Chinese colorectal cancer (CRC) tumor tissues and then screened for functional apaQTLs associated with CRC risk in 17,789 cases and 19,951 controls using GWAS-ChIP data, with independent validation in a large-scale population consisting of 6,024 cases and 10,022 controls. A multi-ancestry-associated apaQTL variant rs1020670 with a C>G change in DNM1L was identified, and the G allele contributed to an increased risk of CRC. Mechanistically, the risk variant promoted aberrant APA and facilitated higher usage of DNM1L proximal poly(A) sites mediated by the RBP CSTF2T, which led to higher expression of DNM1L with a short 3'UTR. This stabilized DNM1L to upregulate its expression, provoking CRC cell proliferation. Collectively, these findings generate a resource for understanding APA regulation and the genetic basis of human cancers, providing insights into cancer etiology.

overall_design: Comparative gene expression profiling analysis of RNA-seq data was performed on 154 colorectal cancer samples from CRC patients

first: Can

last: Chen

first: Zequn

last: Lu

first: Yimin

last: Cai


title: CircRNA profiling  in laminar shear stress-treated human umbilical vein endothelial cells.

city: Guangzhou

first: ziqi

last: chen

address: Zhongshan 2nd Road

institute: Sun Yat-sen University

type:
Expression profiling by array
Non-coding RNA profiling by array

summary: Laminar shear stress (LSS) suppresses endothelial inflammation and protects the arteries from atherosclerosis. Circular RNAs (circRNAs) are powerful regulators of vascular homeostasis and atherosclerosis; however, their roles in mediating the effects of LSS remain unexplored. To identify the changes in circRNA expression patterns after shear stress stimulation, we conducted circRNA microarray analysis using RNA extracted from HUVECs cultured for 24 h under static or LSS conditions.

overall_design: Confluent HUVECs cultured on microscope slides were exposed to well-defined laminar flow with a shear stress of 15 dyne/cm2. HUVECs cultured under static conditions were used as the controls. N=3

first: Ruoyu

last: Wang

first: Yue

last: Zeng

first: Ziqi

first: Dongwei

last: Ma

first: Xiaozhe

last: Zhang

first: Jiajie

last: Pan

first: Wendong

last: Fan

first: Guifu

last: Wu


title: Mettl4 limits the proliferation and differentiation of neural stem cells via regulating RNA alternative splicing

city: Oslo

first: Xu

last: Zhao

address: Sognsvannsveien 20

department: Department of Microbiology

country: Norway

institute: Oslo University Hospital, Rikshospitalet

summary: Previous studies indicate that Mettl4 regulates gene expression and RNA splicing. However, the biological functions of METTL4 remain unclear. Here we show that the absence of Mettl4 leads to sub lethality in mouse and promotes brain function in processing learning and working memory. The neural stem cells in subgranular zone are more proliferative in Mettl4 knockout mice compared with wild type mice. The proliferation and neurogenesis of primary neural stem cells are increased in the absence of Mettl4. Meanwhile, Mettl4 knockout NSCs are more sensitive low levels of H2O2, which is known to promote proliferation and neurogenesis of neural stem cells.  The biological effects caused could be due to the altered gene expression or alternative splicing in neural stem cells when Mettl4 is absent. To investigate the effects at transcriptome level, we performed RNA sequencing.

overall_design: Comparative RNA expression and alternative splicing in wild type and Mettl4 knockout neural stem cells extracted from brains at E14.5. The primary NSCs were cultured in vitro.

first: Pierre

last: Chymkowitch

first: Arne

last: Klungland


title: 3′-seq of Saccharomyces cerevisiae grown in eight different conditions

city: New York

first: Christine

last: Mayr

state: NY

address: 417 E 68th Street

department: Cancer Biology and Genetics

institute: MSKCC

summary: Saccharomyces cerevisiae strain BY4741 (MATa his3Δ1 leu2Δ0 met15Δ0 ura3Δ0) was grown in YPD medium. For control conditions, cells were grown in mid-log phase at 30°C for 20 min or 2 hours. Stress conditions were heat shock (37°C for 20 min), diauxic shift (harvested at 2 hours after glucose depletion), DNA damage (harvested after 2 hours of treatment with 5 µg/ml 4-nitroquinoline 1-oxide (Sigma)). Moreover, four conditions of the yeast metabolic cycle were analyzed. S. cerevisiae strain CEN.PK (MATa URA3 TRP1 LEU2 HIS3 MAL2-8C SUC2) was used and cultivated as described elsewhere (Nocetti and Whitehouse, Genes Dev, 2016). The period of metabolic oscillation was approximately 3.2 h. Cells were harvested at 16, 32, 48, and 60 min from the start of each cycle for one oxidative, two reductive/building, and one reductive/charging phase points. Total RNA from each sample was extracted by the hot phenol method and used for library preparation. 3′-seq libraries were prepared as described previously (Lianoglou et al., Genes Dev, 2013). Each condition was sequenced in three biological replicates. The 27 individual libraries were pooled and subjected to sequencing with HiSeq with SR50 at the Integrated Genomics Operation (MSKCC).

overall_design: Mapping cleavage and polyadenylation sites in Saccharomyces cerevisiae by analyzing pooled 3'-seq data from various conditions.

first: Gang

last: Zhen

first: Buki

last: Kwon


title: Gene expression and functional analysis of GJB2 mutated HaCaT cell lines [G130V]

first: Li

summary:
Vohwinkel syndrome, VS (OMIM#124500), a rare autosomal dominant genetic disease, with less than 50 reported cases in the literature. Although clinical symptoms of VS are complex, which are caused by GJB2 mutation is more typical.
To explore related differential genes and signaling pathways of Vohwinkel syndrome (VS) caused by mutations of GJB2. Human Gene Expression Array of the GJB2-VS mutated (G130V) HaCaT cell lines and identified distinct classes of up- and down- regulated genes during this process.

overall_design: Using GJB2 gene mutant  p.Gly130Val(G130V) primers as templates,  the corresponding cDNA sequence of GJB2: NM_004004 (D66H) was amplified respectively, and then was ligated with GV308 vector. The recombinant plasmids G130V-GV308, and empty vector GV308 were identified and sequenced. Then GJB2-G130V and GJB2-D66H-overexpression lentiviruses. Human Gene Expression Array of the type of GJB2-VS mutated (G130V) and NC HaCaT cell lines for RNA extraction and hybridization on Affymetrix microarrays. And combined Ingenuity Pathway Analysis, combined apoptosis experiment, mRNA expression of LOR, FLG, and differentially expressed genes.

first: Keying

last: GUO


title: Hypoxic regulation of hypoxia inducible factor 1 alpha via antisense transcription

city: Kuopio

first: Minna

middle: U

last: Kaikkonen

department: A.I. Virtanen Institute, Department of Biotechnology and Molecular Medicine

country: Finland

institute: University of Eastern Finland

summary: Impaired oxygen homeostasis is a frequently encountered pathophysiological factor in multiple complex diseases, including cardiovascular disease and cancer. While the canonical hypoxia response pathway is well characterised, less is known about the role of non-coding RNAs in this process. Here, we investigated the nascent and steady-state non-coding transcriptional responses in endothelial cells and their potential roles in regulating the hypoxic response. Notably, we identify a novel antisense long non-coding RNA that convergently overlaps the majority of the HIF1A locus, which is expressed across several cell types and elevated in atherosclerotic lesions. The antisense (HIF1A-AS) is produced as a stable, unspliced and polyadenylated nuclear retained transcript. HIF1A-AS is highly induced in hypoxia by both HIF1A and HIF2A and exhibits anticorrelation with the coding HIF1a transcript. We further characterized this functional relationship by CRISPR-mediated bimodal perturbation of the HIF1A-AS promoter. We provide evidence that HIF1A-AS represses the expression of HIF1a in cis by repressing transcriptional elongation and deposition of H3K4me3, and that this mechanism is dependent on the act of antisense transcription.

overall_design: ChIP-Seq and RNA-Seq analysis of Eahy cells deleted for the HIF1a-AS promoter/TSS.

first: Nicholas

last: Downes

first: Vanesa

last: Tomas-Bosch


title: Affymetrix SNP array data for acute lymphoblastic leukemia samples

city: Kolkata

first: Mayur

last: Parihar

state: West Bengal

address: 14 Major Arterial Road (E-W)

department: Pathology

country: India

institute: Tata Medical Center

summary: In precursor B-cell acute lymphoblastic leukemia (ALL), whole chromosome uniparental isodisomy (wUPD) occurs almost uniquely in the high hyperdiploid (51-67 chromosomes) (HH) subtype. Comparison of 26 HH with wUPD with 31 with noUPD, showed a higher modal number of chromosomes and gains of 5+ in wUPD. Mutations in genes within epigenetic pathways with upregulation of genes involved in cellular response to stress and stimuli, and mutations in RAS/RTK pathways and upregulation of genes in RNA Polymerase III pathway were seen in wUPD and noUPD respectively. Though overall outcomes were similar in patient with and without wUPD, those with noUPD were more likely to have residual disease after treatment. Differential gene expression between the two groups showed upregulation of genes involved in thiopurine drug resistance in wUPD. Genome-wide differences between HH ALL with and without UPD identified plausible biological explanations for the heterogeneity in therapeutic response in HH ALL.

overall_design: Copy number analysis of Affymetrix CytoscanHD SNP arrays was performed for 56 pediatric High hyperdiploid Acute Lymphoblastic Leukemia samples.

first: Vaskar

last: Saha


title: Myeloid BAF60a Deficiency Alters Metabolic Homeostasis and Exacerbates Atherosclerosis

city: Ann Arbor

first: Yang

address: 2800 Plymouth Rd

institute: University of Michigan

overall_design: Refer to individual Series


title: The Second Generation of CAR-iMACs with Enhanced Polarization, CAR-mediated Efferocytosis and Superior Potency against Solid Tumors

first: Hua

last: Yu

address: No.18 Shilongshan Road

institute: Westlake University

summary: Chimeric antigen receptor (CAR)-T cell therapies have shown great success in treating hematologic malignancies. Nonetheless, their therapeutic effect on solid tumors remains to be improved. Recently, macrophages have attracted great attention, given their ability to infiltrate solid tumors, phagocytize tumor cells as well as their immunomodulatory capacities. The first generation of CD3ζ-based CAR-macrophages demonstrated that the CAR could stimulate macrophage phagocytosis in a tumor antigen-dependent way. Here, we genetically engineered induced pluripotent stem cell (iPSC)-derived macrophages (iMACs) with TLR4 intracellular TIR domain-containing CARs against EGFRvIII and GPC3, which yielded markedly enhanced antitumor effect in two different solid tumor models including glioblastoma, and hepatocellular carcinoma in which complete remission was achieved with CAR-iMACs alone or in combination with CD47 antibody. Moreover, the tandem CD3ζ-TIR-CAR, or the “second-generation” design of TIR-based dual signaling CAR, endowed iMACs with both target engulfment/efferocytosis capacity against antigen-expressing solid tumor cells, and potency of antigen-dependent M1 state polarization and M2 state resistance in an NF-κB dependent manner. We also illustrated a surprising mechanism of tumor cell elimination by CAR-induced efferocytosis against tumor cell apoptotic bodies. Taken together, we established the next generation CAR-iMACs equipped with orthogonal phagocytosis and polarization capacity for better antitumor functions in treating solid tumors.

overall_design: We pretreated EGFRvIII-targeting CAR-iMACs were incubated with U87MGEGFRvIII cells with IFN-γ/LPS for 24 hours. Subsequently, we performed 10×genomics single-cell RNA-sequencing  and found  that TIR-based CARs contributed to the proinflammatory state of CAR-iMACs in a CAR- and antigen-dependent manner when encountering tumor cells and improved immune activity against target tumor cells.

first: Anhua

last: Lei

first: Yuqing

last: Zhu


title: Muscle from two pig breeds with varying muscularity

city: Guwahati

first: Narayana

last: Mohan

state: Assam

address: Rani

department: ICAR-National Research Centre on Pig

institute: Indian Council of Agricultural Research

type: Methylation profiling by high throughput sequencing


title: Acetylation of EED by CBP/P300 Regulates the Targeting of PRC2 and is Required for mESC Differentiation [RNA-seq]

last: Li

overall_design: RNA sequencing of wildtype mESCs, FLAG-EED wildtype and FLAG-EED K19R mutant knock-in mESCs and EBs. All experiments were conducted in duplicates.

first: Yiqi

last: Yang

first: Yunze

last: Liu


title: Differential expression profile and diagnostic value of plasma exosomal circular RNA in hepatocellular carcinoma

city: Ningbo

first: liyun

last: fu

state: Zhejiang

address: ningbo xibei street No.41, zhejiang province

department: Department of Hepatology

institute: HwaMei Hospital, University of Chinese Academy of Sciences (Ningbo No.2 hospital)

summary: Circular RNAs (circRNAs) are recently found to be promising kind of biomarkers for tumors. However, plasma exosomal circRNAs in the diagnosis of hepatocellular carcinoma (HCC) is largely unknown. In this study, we report the application of next-generation sequencing technology for high-throughput profiling of plasma exosome general transcriptome between 3 cases of hepatocellular carcinoma patients and 3 cases of healthy control. We obtained exosomes through ultrahigh-speed centrifugation techniques and verified exosomes through immunoblotting, NanoSight NS300(Malvern Panalytical, Malvern, UK) and JEM-1230 transmission electron microscope (TEM) (JEOL, Tokyo, Japan). And then, we find that there are 41 circRNAs that have differential expression. This study discovered the differences between hepatocellular carcinoma and healthy people preliminarily.

overall_design: Plasmas exosomes from three samples of HCC patients and three samples of healthy control were extracted and compared the differential expression profile between them.

first: Liyun

last: Fu


title: Gene expression profile of breast cancer cell lines

city: Oakland

first: Ravi

last: Sachidanandam

address: 2945 Webster St

institute: Girihlet Inc.

summary: Inflammatory Breast Cancer (IBC) is the most aggressive form of breast cancer, but understanding of the unique aspects of IBC biology lags far behind that of other breast cancers. One of the key barriers in advancing understanding of the molecular determinants of IBC has been the lack of adequate models for this disease that presents with distinct clinical and histopathological features. We describe here a novel human triple-negative IBC cell line, A3250, that recapitulates key features of human IBC in a mouse orthotopic model including skin erythema, diffuse tumor growth, high stroma to tumor ratio, dermal lymphatic invasion, and extensive lymph node and distant metastases. Tumor-associated macrophages were particularly enriched, and A3250 cells expressed very high levels of CCL2, a macrophage recruiting chemokine in the absence of detectable levels of its cognate receptor. CCL2 knockdown led to a striking reduction in macrophage densities, tumor cell proliferation, and metastasis in vivo. These results identify tumor-derived CCL2 as a key factor driving macrophage expansion and indirectly, the growth of A3250 IBC cells in vivo. Comparison of the A3250 chemokine expression profile with human IBC expression data sets revealed sharing of this profile across different IBC subtypes as well as enrichment for macrophage expression. Thus, this human IBC model provides a unique opportunity to uncover novel aspects of IBC biology, and to test novel therapies for this deadly disease.

overall_design:
The gene expression profile of A3250 inflamatory breast cancer cells was compared to that of other breast cancer cell lines
RNA was derived from the indicated breast cancer cell lines
*** Submitter states raw data cannot be supplied due to loss of files.

first: Stuart

last: Aaronson

first: Luca

last: Grumolato


title: Antiepileptic medication induces homeostatic synaptic plasticity in pyramidal neurons of the adult human neocortex

city: Hannover

first: Maximilian

last: Lenz

address: Carl-Neuberg-Straße 1

department: Institute of Neuroanatomy and Cell Biology

country: Germany

institute: Hannover Medical School

summary: Homeostatic synaptic plasticity serves to maintain neuronal function within a dynamic range, compensating for perturbations in network activity. While coordinated structural and functional changes at synaptic sites play a crucial role in adaptive processes, the specific regulatory mechanisms and biological relevance of homeostatic plasticity in the human brain warrant further investigation. In this study, we investigated the effects of neural network silencing, achieved through pharmacological inhibition of voltage-gated sodium channels or glutamatergic neurotransmission – common targets of antiepileptic medication – on functional and structural properties of murine and human cortical tissue. Using mouse entorhino-hippocampal tissue cultures, acute slices of adult mice, and human brain tissue, we characterize homeostatic synaptic plasticity across models, brain regions, and species. Our findings demonstrate local homeostatic synaptic plasticity in the adult human neocortex, highlighting the potential effects of antiepileptic medication in brain regions unaffected by the primary diseases, which might represent a mechanism for neuropsychiatric effects linked to these medications and increased seizure susceptibility upon discontinuation of antiepileptic medication.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Comparative gene expression profiling analysis of RNA-seq data in different brain regions across species to decipher the effects of pharmacological activity deprivation in neural circuits.

first: Pia

last: Kruse

first: Amelie

last: Eichler

first: Jakob

last: Straehle

first: Hanna

last: Hemeling

first: Phyllis

last: Stöhr

first: Jürgen

last: Beck

first: Andreas

last: Vlachos


title: Investigating the effect of Daxx deletion on gene and repeat expression in embryonic stem cells and their differentiated neural derivatives

city: Edinburgh

first: Rob

last: Illingworth

state: Midlothian

address: Centre for Regenerative Medicine

department: Centre for regenerative Medicine

institute: The University of Edinburgh

summary: Genomes comprise a large fraction of repetitive sequences folded into constitutive heterochromatin to protect genome integrity and cell identity. De novo formation of heterochromatin during preimplantation development is essential to preserve the ground-state of pluripotency and the self-renewal capacity of embryonic stem cells (ESCs). In this study we find that DAXX, an H3.3 chaperone, is essential for ESCs maintenance in the ground-state of pluripotency. DAXX accumulates at pericentromeric regions, and recruits PML and SETDB1, thereby promoting heterochromatin formation. In the absence of DAXX, the 3D-architecture and physical properties of pericentric and peripheral heterochromatin are disrupted, resulting in derepression of major satellite DNA, transposable elements and genes associated with the nuclear lamina. Our data reveal that DAXX is crucial for the maintenance and 3D-organization of the heterochromatin compartment and protects ESCs viability.

overall_design: To investigate the function of DAXX we generated a transgenic mouse ESC line lacking a functional copy of the Daxx gene (Daxx-/-). We cultured wildtype parental (WT; E14Tg2a) and Daxx-/- ESCs in three conditions: 1). Serum + LIF (ESCs), 2). 2i + Vitamin C (ground state pluripotency) and 3). Retinoic Acid -LIF (Neural differentiated). We isolated total RNA from each of these samples across three independent replicates and, following rRNA depletion, performed RNA-seq on each sample. We performed comparative gene and repeat expression analysis between WT and Daxx-/- cells cultured under each of these three conditions.

first: Antoine

last: Canat

first: Adeline

last: Veillet

first: Renaud

last: Batrin

first: Clara

last: Dubourg

first: Robert

first: Emmanuelle

last: Fabre

first: Pierre

last: Therizols


title: Suture-anchored cutaneous tension induces persistent hypertrophic scarring in a novel murine model [miRNA]

status: Public on Sep 28 2023

city: Shanghai

first: Yashu

last: Li

address: No,1665, Kongjiang Road, Yangpu District

department: Department of Plastic Surgery

institute: Xinhua hospital, Shanghai Jiaotong University

summary: Hypertrophic scars arise from dysregulated wound healing under prolonged mechanical tension, causing disfiguring fibrosis. However, limited preclinical models replicate key features of human tension-induced scarring. We developed an innovative murine model utilizing suture anchoring to impose persistent transverse-axial stretch across healing incisions, mimicking excessive wound tension that leads to hypertrophy clinically. Dorsal paired incisions were generated in mice, with wound edges on the upper back sutured to the rib cage while leaving wound edges on the lower back relaxed. This localized anchoring restrained wound contraction, maintaining high tension throughout remodeling analogous to scars widening under stress. Stretched upper wounds developed profound fibrotic changes compared to relaxed controls. Scars induced by suture-anchored tension displayed macroscopic hypertrophy, hardness, erythema, and pruritis up to 3 months. Histologically, scars induced by suture-anchored tension were hypercellular, hypervascular, hyperproliferative with disorganized extracellular matrix deposition, and displayed molecular hallmarks of hypertrophic fibrosis. MiRNA sequencing revealed the different signature in suture-anchored tension induced hypertrophic scars compared to control normal scars.

overall_design: To identify the miRNA  differences of hypertrophic scars generated in stretched group (HS) and normal scars formed in control group (NS), miRNA sequencing was conducted upon scar samples 2 weeks post excision. Comparative gene expression profiling analysis of miRNA-seq data for HS and NS.

first: Dale

last: Xu

first: Weifeng

last: He

first: Kaiyang

last: Lv


title: Genomic annotation of a non-coding region deleted on 4q25 [CUT&RUN]

city: nantes

first: Julien

last: Barc

address: 8 quai moncousu

summary: we identified 7 families presenting a new cardiac entity associated with a heterozygous deletion of 2 CTCF binding sites on 4q25, inducing TAD fusion and chromatin conformation remodeling. The CTCF binding sites are located in a gene desert at 1 Mb from the Paired-like homeodomain transcription factor 2 gene (PITX2).

overall_design: CUT&RUN on histone marks H3K4me3, H3K4me1 and H3K27ac and on the CTCF transcription factor

first: Manon

last: Baudic

first: Estelle

last: Baron

first: Enora

last: Fremy

first: Adrien

last: Foucal

first: Jean-Jacques

last: Schott


title: Quality control recommendations for RNASeq using FFPE samples based on pre-sequencing lab metrics and post-sequencing bioinformatics metrics

city: Rochester

first: Aditya

last: Bhagwate

state: MN

address: 200 1st Street SW

institute: Mayo Clinic

summary: Formalin-fixed, paraffin-embedded (FFPE) tissues have many advantages for identification of risk biomarkers, including wide availability and potential for extended follow-up endpoints. However, RNA derived from archival FFPE samples has limited quality. Here we identified parameters that determine which FFPE samples have the potential for successful RNA extraction, library preparation, and generation of usable RNAseq data. We optimized library preparation protocols designed for use with FFPE samples using seven FFPE and Fresh Frozen replicate pairs, and tested optimized protocols using a study set of 130 FFPE biopsies from women with benign breast disease. Metrics from RNA extraction and preparation procedures were collected and compared with bioinformatics sequencing summary statistics. Finally, a decision tree model was built to learn the relationship between pre-sequencing lab metrics and qc pass/fail status as determined by bioinformatics metrics.. Samples that failed bioinformatics qc tended to have low median sample-wise correlation within the cohort (Spearman correlation < 0.75), low number of reads mapped to gene regions (< 25 million), or low number of detectable genes (11,400 # of detected genes with TPM > 4). The median RNA concentration and pre-capture library Qubit values for qc failed samples were 18.9 ng/ul and 2.08 ng/ul respectively, which were significantly lower than those of qc pass samples (40.8 ng/ul and 5.82 ng/ul). We built a decision tree model based on input RNA concentration, input library qubit values, and achieved an F score of 0.848 in predicting QC status (pass/fail) of FFPE samples. We provide a bioinformatics quality control recommendation for FFPE samples from breast tissue by evaluating bioinformatic and sample metrics. Our results suggest a minimum concentration of 25 ng/ul FFPE-extracted RNA for library preparation and 1.7 ng/ul pre-capture library output to achieve adequate RNA-seq data for downstream bioinformatics analysis.

overall_design: A study set of 130 FFPE biopsies from women with benign breast disease along with 7 additional FFPE and Fresh Frozen replicates were prepped and sequenced using Illumina’s TruSeq RNA Exome and NEBNext rRNA Depletion protocols.

first: Amy

last: Degnim

first: Chen

last: Wang

first: Yuanhang

last: Liu


title: Isolated loss of the AUTS2 long isoform, brain-wide or targeted to Calbindin-lineage cells, generates a specific suite of brain, behavioral and molecular pathologies

city: Seattle

first: Christopher

last: Seward

address: Stubbs Lab, Rm 404, 720 Broadway

institute: Pacific Northwest Research Institute

summary: Rearrangements within the AUTS2 region are associated with a rare syndromic disorder with intellectual disability, developmental delay and behavioral abnormalities as core features. In addition, smaller regional variants are linked to wide range of neuropsychiatric disorders, underscoring the gene’s essential role in brain development. Like many essential neurodevelopmental genes, AUTS2 is large and complex, generating distinct long (AUTS2-l) and short (AUTS2-s) protein isoforms from alternative promoters. Although evidence suggests unique isoform functions, the contributions of each isoform to specific AUTS2-linked phenotypes have not been clearly resolved. Furthermore, Auts2 is widely expressed across the developing brain, but cell populations most central to disease presentation have not been determined. In this study, we focused on the specific roles of AUTS2-l in brain development, behavior, and postnatal brain gene expression, showing that brain-wide AUTS2-l ablation leads to specific subsets of the recessive pathologies associated with C-terminal mutations that disrupt both isoforms. We identify downstream genes that could explain expressed phenotypes including hundreds of putative direct AUTS2-l target genes. Furthermore, in contrast to C-terminal Auts2 mutations which lead to dominant hypoactivity, AUTS2-l loss-of-function is associated with dominant hyperactivity, a phenotype exhibited by many human patients. Finally, we show that AUTS2-l ablation in Calbindin 1-expressing cell lineages is sufficient to yield learning/memory deficits and hyperactivity with abnormal dentate gyrus granule cell maturation, but not other phenotypic effects. These data provide new clues to in vivo AUTS2-l functions and novel information relevant to genotype-phenotype correlations in the human AUTS2 region.

overall_design: RNA Sequencing was performed on RNA samples collected from individual forebrain of P0 or hippocampus of P14 mice.  Two mouse strains were sampled in biological triplicate, Cmv-cre, Auts2-ex7-/-, and Nes-cre, Auts2-ex7-/-.  Paired end RNA Libraries were prepared and sequenced on a Hiseq 4000 sequencer with a read length of 2x150bp.

first: Yunshu

last: Song

first: Chih-Ying

last: Chen

first: Amber

last: LeBlanc

first: Analise

last: Leddy

first: Lisa

last: Stubbs


title: Effect of overexpression of FOXA2 on biological behavior of SGC-996

city: Fuzhou

first: Yanling

address: xinquan road

department: Hepatobiliary surgery

institute: Fujian medical university union hospital

summary: In order to investigate the role of FOXA2 in the occurrence and development, we infected gallbladder cancer cells with recombinant lentivirus to construct FOXA2 overexpression SGC-996.   We then performed gene expression profiling analysis using data obtained from RNA-seq of 6 different gallbladder cancer cells from  SGC-996-NC or SGC-996-OE groups.

overall_design: Comparative gene expression profile analysis of RNA-seq data from 6 different gallbladder carcinoma cells in the SGC-996-NC and SGC-996-OE groups.

first: Lingju

last: Hong

first: Mingyuan

first: Maotuan

last: Huang

first: Weihong

first: Xiahenazi

last: Abudukeremu

first: Feifei

last: She


title: Excess folic acid supplementation increases adiposity and impacts insulin sensitivity in female mice during pregnancy and has sex-specific effects on fetal offspring

city: Vancouver

first: Angela

last: Devlin

state: British Columbia

address: 938 W 28th Ave

institute: University of British Columbia

summary: We determined the effects of excess folic acid supplementation (5x recommendation) on maternal and fetal offspring metabolic health. Using a mouse (female C57BL/6J) model of gestational dibetes (GDM; 45% kcal fat diet) and control mice (10% kcal diet) we show that  folic acid supplementation increased weight gain and fat mass in both GDM and control mice but improved insulin sensitivity in GDM mice and worsened insulin sensitivity in control mice. We found no unmetabolized folic acid in liver from supplemented mice suggesting the metabolic effects of folic acid supplementation may not be due to unmetabolized folic acid. Male fetal (gestational day 18.5) offspring from folic acid supplemented dams (GDM and control) had greater beta cell mass and density than those from unsupplemented dams; this was not observed in female offspring. Differential sex-specific hepatic gene expression profiles were observed in the offspring from supplemented dams but this differed between GDM and controls. Our findings suggest that folic acid supplementation affects insulin sensitivity in female mice, but is dependent on their metabolic phenotype, and has sex-specific effects on offspring pancreas and liver.

overall_design: Female C57BL/6J) mouse model of GDM (45% kcal fat diet) and control mice (10% kcal diet), with and without supplemental folic acid (2 or 10 mg/kg diet folic acid). On diet for 13 weeks before breeding and throughout pregnancy. Liver tissue from male and female fetal offpsring (gestational day 18.5) of each maternal diet group was sequenced (bulk RNA-seq).

first: Ei-Xia

last: Mussai

first: Zoe

middle: A

last: Lofft

first: Ben

last: Vanderkruk

first: Nicha

last: Boonpattrawong

first: Joshua

middle: W

last: Miller

first: Teodoro

last: Bottiglieri


title: Zebrafish models of Mucopolysaccharidosis types IIIA, B, & C show hyperactivity and changes in oligodendrocyte state

city: Adelaide

first: Lachlan

last: Baer

address: North Terrace

country: Australia

institute: The University of Adelaide

summary: Sanfilippo syndrome childhood dementia, also known as mucopolysaccharidosis type III (MPS III), is a rare inherited lysosomal storage disorder. Subtypes of MPS III are caused by deficiencies in one of four enzymes required for degradation of the glycosaminoglycan heparan sulfate (HS). An inability to degrade HS leads to progressive neurodegeneration and death in the second or third decades of life. Knowledge of MPS III pathogenesis is incomplete, and no effective therapies exist. We generated the hypomorphic mutations sgshS387Lfs, nagluA603Efs and hgsnatG577Sfsin the endogenous zebrafish genes orthologous to human SGSH, NAGLU, and HGSNAT that are loci for mutations causing MPS III subtypes MPS IIIA, B and C respectively. Our models display the primary MPS III disease signature of significant brain accumulation of HS, while behavioural analyses support hyperactivity phenotypes. Brain transcriptome analysis revealed changes related to lysosomal, glycosaminoglycan, immune system and iron homeostasis biology in all three models but also distinct differences in brain transcriptome state between models. The transcriptome analysis also indicated marked disturbance of the oligodendrocyte cell state in the brains of MPS IIIA, B and C zebrafish, supporting that effects on this cell type are an early and consistent characteristic of MPS III. Overall, our zebrafish models recapture key characteristics of the human disease and phenotypes seen in mouse models. Our models will allow exploitation of the zebrafish’s extreme fecundity and accessible anatomy to dissect the pathological mechanisms both common and divergent between the MPS IIIA, B, and C subtypes.

overall_design: To compare brain transcriptomes in our three MPS III zebrafish lines, we generated families of 2nd Filial generation zebrafish originating from G0 in-crosses of parental zebrafish heterozygous for two of the MPS III mutations. We initially aimed to analyse n = 6 fish per genotype from single families of progeny from in-crosses of doubly heterozygous mutant fish. We chose n = 6 as, according to a previous power calculation, this would allow approximately 70% power to detect differentially expressed genes in zebrafish models of Alzheimer’s disease. However, due to smaller than expected clutch sizes, and questionable purified RNA quality, we were forced to use multiple families of siblings, or n<6, in some analyses. For the A and B (AB) arm of the experiment, we analysed n = 4 fish per genotype, which also included 4 sgsh S387Lfs/+ fish to explore cellular processes affected in the brains of presumably unaffected carriers of MPS III mutations. For the A and C (AC) arm, we sequenced all homozygous sgsh S387Lfs zebrafish in the family (n = 7) to account for the lower number of fish with this genotype in the AB arm. For the B and C (BC) arm, we could not obtain a large enough family in a single breeding event within our timeframe to be able to compare with the other families. Therefore, we analysed fish spawned by two pairs of parents (P1 and P2, both doubly heterozygous for the nagluA603fs and hgsnatG577fs mutations) in a total of 3 spawning events.

first: Ewan

last: Gerken

first: Syahida

last: Ahmad

first: Karissa

last: Barthelson

first: Michael

last: Lardelli


title: Exploring Food-induced Obesity and Macrophage Related Differential Genes in Subcutaneous and Visceral Fat between Obese and Normal Individuals

city: Shenyang

first: Wensu

last: Liu

state: Liaoning

address: 77 Puhe Road, Shenyang North New Area

institute: China Medical University

summary: As a serious public health problem nowadays, obesity and its complications are receiving gradually more attention, especially on how food-induced obesity causes the expansion of adipose tissue and leads to the occurrence of metabolic inflammation and infiltration of immune cells. It's still worth mentioning that numerous genes in adipose tissue which differ between obese and normal individuals in food-induced obesity were not insufficiently studied. Applying machine learning methods and natural language processing methods on transcriptome datasets and PubMed abstracts, we investigated 8 genes with differential expression and high association with macrophages in the visceral and subcutaneous adipose tissue between obese and normal individuals, and meanwhile low correlation with leptin. Compared with normal group, Quantitative Real-time Polymerase Chain Reaction, Western Blot, immunohistochemistry, immunofluorescence and other subsequent animal experiments on the 8 filtered genes were applied in the adipose tissue of  C57BL/6J and OB/OB mice, we found that the expression of glycoprotein IX (GP9), MHC class I polypeptide related sequence B (MICB), CDGSH iron sulfur domain 1 (CISD1) and Pre-mRNA-processing factor 19 (PRPF19)  in protein level were down-regulated in visceral fat of obese mice induced by high-fat diet. And these obese mice showed decreased levels of mannosidase alpha class 2B member 1 (MAN2B1) and  TYRO protein tyrosine kinase binding protein (TYROBP) in their subcutaneous adipose tissue, while phosphate and tension homology deleted on chromosome pten (PTEN) and eukaryotic translation initiation factor 4E binding protein 1 (EIF4EBP1) had been sufficiently studied. The pattern in OB/OB mouse was dramatically different with these patterns. Through double immunofluorescence staining, it was found that all genes were co-localized with CD206, which was the marker of M1 and M2 macrophage in adipose tissue. Further interacting drugs for the genes were calculated and subsequently circumin and beta-catonin were proved to be interacting with these genes and functioned in inhibiting food-induced obesity. Further knowledge gragh was constructed to illustrate the relationship between obesity, filtered genes, drugs and related concepts. To summarize our findings, we discovered some new differential genes in adipose tissue which could potentially establish a new connection between adipose tissue macrophages and food-induced obesity. Furthermore, they offered unique insights into exploring the mechanism of adipose tissue genes related to food-induced obesity and related diseases.

overall_design: In order to view the effect of beta-carotene on our filtered genes for subcutaneous and visceral fat, we performed RNA-seq for 8 groups with 5 samples for each group of C57BL/6J mouse.. The 8 gourps included 4 groups for subcutaneous fat and 4 groups for visceral fat for epididymis. The 4 group included  normal group, normal+drug group, high-fat diet group and high-fat+drug group. RNA-seq was afterwards performed on each group.

first: Yang

first: Huihui

first: Deliang

last: Wen


title: A ketogenic diet can mitigate SARS-CoV-2 induced systemic reprogramming and inflammation

city: Los Angeles

first: Feiyang

last: Ma

address: 610, Charles Young Dr East, TLSB 3000C

department: Molecular Biology Institute

institute: UCLA

summary: The ketogenic diet (KD) has demonstrated benefits in numerous clinical studies and animal models of disease in modulating the immune response and promoting a systemic antiinflammatory state. Here we investigate the effects of a KD on systemic toxicity in mice following SARS-CoV-2 infection. Our data indicate that under KD, SARS-CoV-2 reduces weight loss with overall improved animal survival. Muted multi-organ transcriptional reprogramming and metabolism rewiring suggest that a KD initiates and mitigates systemic changes induced by the virus. We observed reduced metalloproteases and increased inflammatory homeostatic protein transcription in the heart, with decreased serum pro-inflammatory cytokines (i.e., TNF-α, IL-15, IL-22, G-CSF, M-CSF, MCP-1), metabolic markers of inflammation (i.e., kynurenine/tryptophane ratio), and inflammatory prostaglandins, indicative of reduced systemic inflammation in animals infected under a KD. Taken together, these data suggest that a KD can alter the transcriptional and metabolic response in animals following SARS-CoV-2 infection with improved mice health, reduced inflammation, and restored amino acid, nucleotide, lipid, and energy currency metabolism.

overall_design: Bulk RNA sequencing comparing mouse tissues with differenet hACE2 genotype, Chow or Keto diet, and SARS-CoV-2 or Mock treatment

first: Amelia

last: Palermo

first: Shen

last: Li

first: Arjun

last: Deb


title: GiRAFR improves gRNA detection and annotation in single cell CRISPR screens

city: Leuven

first: Bernard

last: Thienpont

address: ON4 Herestraat 49

department: Department of Human Genetics

country: Belgium

institute: KU Leuven

summary: Novel single cell RNA-seq analysis combined with CRISPR screens enables the high- throughput characterization of transcriptional changes caused by genetic perturbations. Dedicated software to annotate CRISPR guide RNA (gRNA) libraries and associate them with single cell transcriptomes are however lacking. Here, we generated a CRISPR droplet sequencing dataset. We demonstrate that the current default tool fails to detect mutant gRNAs. We therefore developed GiRAFR, a pysam-based software tool to characterize intact and mutant gRNAs. We show that mutant gRNAs are dysfunctional, and failure to detect and annotate them leads to an inflated estimate of the number of untransformed cells as well as an underestimated multiplet frequency. These findings are mirrored in publicly available datasets, where we find that up to 34 % of cells are transduced with a mutant gRNA. Applying GiRAFR hence stands to improve the annotation and quality of single cell CRISPR screens.

overall_design: We transformed A549 cells expressing a tamoxifen-inducible Cas9 with a lentiviral pool for expression of 120 gRNAs at low multiplicity of infection. After stringent selection using puromycin and 2 days of Cas9 induction, cells were allowed to grow for another 5 days.

first: Qian

last: Yu

first: Paulien

last: Van Minsel

first: Eva

last: Galle


title: Longitudinal host transcriptional responses to SARS-CoV-2 infection in adults with extremely high viral load

city: Houston

first: Sarah

middle: Yvette

last: Ponce

state: TX

address: 1 Baylor Plaza

department: HGSC

institute: Baylor College of Medicine

summary: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the etiologic agent of the coronavirus disease 2019 (COVID-19) pandemic. We conducted a longitudinal study to investigate gene expression patterns during the acute SARS-CoV-2 illness. The cases included SARS-CoV-2 infected individuals with an extremely high viral load early in their illness matched to individuals who either had a low SARS-CoV-2 viral load early in their infection or were otherwise stable patients who tested negative for SARS-CoV-2 prior to their outpatient surgical or aerosol generating procedure. We detected hundreds of up-regulated genes that were highly correlated to the SARS-CoV-2 viral load. Many of these up-regulated genes were enriched in cellular pathways involved in the innate immune response, antiviral interferon and cytokine signaling, and cell death.

overall_design: Of 73 mid-turbinate (MT) swab samples originally collected from SARS-CoV-2 cases with longitudinal follow-up, 44 (60.3%) MT swab samples from 20 (66.7%) individuals were of good quality to generate RNA-sequence data to study the host response to SARS-CoV-2 infection over time. Widespread gene expression changes attributable to transcriptional host responses to SARS-CoV-2 infection were determined by correlating the expression of each gene with sample viral load (representing the inverse correlation with Ct value) across the 44 MT swab samples. **RAW DATA NOT PROVIDED DUE TO PATIENT PRIVACY CONCERNS**

first: Chad

last: Creighton

first: Yiqun

last: Zhang

first: Pedro

last: Piedra

first: Vasanthi

last: Avadhanula


title: Aberrant differentiation of epithelial progenitors is accompanied by a hypoxic microenvironment in the paraquat-injured human lung

city: Beijing

first: Yanxiao

last: Wang

address: 7 Science Park Road ZGC Life Science Park

institute: National Institute of Biological Sciences

summary: Paraquat is a toxic herbicide that can cause severe lung injury, leading to alveolar epithelial cell death, subsequent lung fibrosis and respiratory failure. Understanding the repair process following paraquat injury is critical for developing effective therapeutic strategies to treat paraquat-poisoned patients. However, the lung repair program in paraquat-injured lungs is currently unknown. In this study, we analyzed lung parenchyma samples from an 18-year-old female patient who had ingested paraquat and subsequently underwent a double-lung transplantation on the 34th day after poisoning. By the combination of single cell RNA sequencing analysis and immunostaining analysis, we reveal that the airway stem/progenitor cells and AT2 cells in the paraquat-injured lung exhibit a significantly reduced capacity to regenerate alveolar epithelium epithelial in the paraquat-injured lung. The hypoxic microenvironment may contribute to this aberrant repair program by activating the NOTCH signaling pathway, ultimately leading to an irreversible loss of gas-exchange units and an exacerbated lung tissue damage.

overall_design: Lung specimens of the paraquat poisoning patient were obtained from the lung removed at the time of lung transplantation. This patient is an 18-year-old female who had ingested paraquat and subsequently underwent a double-lung transplantation on the 34th day after poisoning. Epithelial cells, endothelial cells, immune cells, and stromal cells were harvested by fluorescence-activated cell sorting and then single-cell RNA analysis were performed to analysis the cellular and molecular changes of the paraquat-injured lung.

first: Ennan

last: Bin

first: Jie

last: Yuan

first: Man

last: Huang

first: Jingyu

last: Chen

last: Tang