RNAi Reveals Phase-Specific Global Regulators of Human Somatic Cell Reprogramming.
Incomplete knowledge of the mechanisms at work continues to hamper efforts to maximize reprogramming efficiency. Here, we present a systematic genome-wide RNAi screen to determine the global regulators during the early stages of human reprogramming. Our screen identifies functional repressors and effectors that act to impede or promote the reprogramming process. Repressors and effectors form close interacting networks in pathways, including RNA processing, G protein signaling, protein ubiquitination, and chromatin modification. Combinatorial knockdown of five repressors (SMAD3, ZMYM2, SFRS11, SAE1, and ESET) synergistically resulted in ∼85% TRA-1-60-positive cells. Removal of the novel splicing factor SFRS11 during reprogramming is accompanied by rapid acquisition of pluripotency-specific spliced forms. Mechanistically, SFRS11 regulates exon skipping and mutually exclusive splicing of transcripts in genes involved in cell differentiation, mRNA splicing, and chromatin modification. Our study provides insights into the reprogramming process, which comprises comprehensive and multi-layered transcriptional, splicing, and epigenetic machineries.
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, Singapore 117456, Singapore.
Cheng-Xu Delon
CX
Toh
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore.
Jun-Wei
JW
Chan
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore.
Zheng-Shan
ZS
Chong
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
Hao Fei
HF
Wang
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; College of Life Sciences, Nankai University, Tianjin 300071, China.
Hong Chao
HC
Guo
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore.
Sandeep
S
Satapathy
Research and Development Unit (RDU), National Heart Centre Singapore, 5(th) Hospital Drive, Singapore 169609, Singapore.
Dongrui
D
Ma
Membrane Traffic Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore.
Germaine Yen Lin
GY
Goh
Division of Cancer Genetics and Therapeutics, Laboratory of NF-κB Signaling, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore.
Ekta
E
Khattar
Department of Molecular Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Genetics, Harvard Medical School, Boston, MA 02114, USA; Howard Hughes Medical Institute, Boston, MA 02114, USA.
Lin
L
Yang
Division of Cancer Genetics and Therapeutics, Laboratory of NF-κB Signaling, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117545, Singapore.
Vinay
V
Tergaonkar
Lab of Bioimaging Probe Development, A(∗)STAR Singapore Bioimaging Consortium (SBIC), 11 Biopolis Way Helios, Singapore 138673, Singapore; Department of Chemistry and MedChem Program, Life Sciences Institute, National University of Singapore, 3 Science Drive 3, Singapore 117543, Singapore.
Young-Tae
YT
Chang
Howard Hughes Medical Institute, Boston, MA 02114, USA; Institute for Medical Engineering and Science Department of Biological Engineering, and Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115, USA.
James J
JJ
Collins
Howard Hughes Medical Institute, Boston, MA 02114, USA; Stem Cell Transplantation Program, Division of Pediatric Hematology/Oncology, Boston Children's Hospital and Dana-Farber Cancer Institute, Boston, MA 02115, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Harvard Stem Cell Institute, Boston, MA 02115, USA; Manton Center for Orphan Disease Research, Boston, MA 02115, USA.
George Q
GQ
Daley
A(∗)STAR Institute of High Performance Computing (IHPC), Connexis, Singapore 138632, Singapore; A(∗)STAR Bioinformatics Institute, Singapore 138671, Singapore.
Keng Boon
KB
Wee
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
Chadi A El
CA
Farran
Center for Individualized Medicine, Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, Rochester, MN 55905, USA. Electronic address: li.hu@mayo.edu.
Hu
H
Li
NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, Singapore 117456, Singapore; Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117545, Singapore; A(∗)STAR Bioinformatics Institute, Singapore 138671, Singapore. Electronic address: bchlyp@nus.edu.sg.
Yoon-Pin
YP
Lim
Membrane Traffic Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117545, Singapore.
Frederic A
FA
Bard
Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, Singapore 117456, Singapore; Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore. Electronic address: yhloh@imcb.a-star.edu.sg.
Yuin-Han
YH
Loh
D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000235:genetics*; D055785:Gene Knockdown Techniques; D005820:Genetic Testing; D015894:Genome, Human; D006801:Humans; D007700:Kinetics; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012326:RNA Splicing / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D000068103:Serine-Arginine Splicing Factors / Q000378:metabolism
D008869:Microtubule-Associated Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D012097:Repressor Proteins; C000595775:SRSF11 protein, human; C587673:ZNF207 protein, human; D000068103:Serine-Arginine Splicing Factors
SFRS11; ZNF207; genome-wide siRNA screen; human somatic cell reprogramming; reprogramming specific alternative splicing