Dynamic transcriptomes during neural differentiation of human embryonic stem cells revealed by short, long, and paired-end sequencing
- Jia Qian Wu a , 1 , 2 ,
- Lukas Habegger b , 1 ,
- Parinya Noisa c ,
- Anna Szekely d ,
- Caihong Qiu e ,
- Stephen Hutchison f ,
- Debasish Raha g ,
- Michael Egholm f ,
- Haifan Lin e ,
- Sherman Weissman d ,
- Wei Cui c ,
- Mark Gerstein b , h , i , and
- Michael Snyder a , 2
- aDepartment of Genetics, Stanford University School of Medicine, Stanford, CA 94305;
- bProgram in Computational Biology and Bioinformatics, Yale University, New Haven, CT 06511;
- cInstitute of Reproductive and Developmental Biology, Imperial College London, Hammersmith Hospital, W12 ONN London, United Kingdom;
- dDepartment of Genetics, Yale University, New Haven, CT 06511;
- eYale Stem Cell Center, Yale University, New Haven, CT 06509;
- f454 Life Sciences Sequencing Centre, Branford, CT 06405; and
- gDepartments ofgMolecular, Cellular and Developmental Biology,
- hMolecular Biophysics and Biochemistry, and
- iComputer Science, Yale University, New Haven, CT 06511
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Edited* by Joseph R Ecker, Salk Intstitute, La Jolla, CA, and approved January 26, 2010 (received for review December 8, 2009)
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1J.Q.W and L.H contributed equally to this work.
Abstract
To examine the fundamental mechanisms governing neural differentiation, we analyzed the transcriptome changes that occur during the differentiation of hESCs into the neural lineage. Undifferentiated hESCs as well as cells at three stages of early neural differentiation—N1 (early initiation), N2 (neural progenitor), and N3 (early glial-like)—were analyzed using a combination of single read, paired-end read, and long read RNA sequencing. The results revealed enormous complexity in gene transcription and splicing dynamics during neural cell differentiation. We found previously unannotated transcripts and spliced isoforms specific for each stage of differentiation. Interestingly, splicing isoform diversity is highest in undifferentiated hESCs and decreases upon differentiation, a phenomenon we call isoform specialization. During neural differentiation, we observed differential expression of many types of genes, including those involved in key signaling pathways, and a large number of extracellular receptors exhibit stage-specific regulation. These results provide a valuable resource for studying neural differentiation and reveal insights into the mechanisms underlying in vitro neural differentiation of hESCs, such as neural fate specification, neural progenitor cell identity maintenance, and the transition from a predominantly neuronal state into one with increased gliogenic potential.
Footnotes
- 2To whom correspondence may be addressed. E-mail: Jiaqian2009.wu{at}gmail.com or mpsnyder{at}stanford.edu.
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Author contributions: J.Q.W., S.W., and M.S. designed research; J.Q.W., L.H., P.N., A.S., C.Q., and S.H. performed research; L.H., P.N., A.S., C.Q., D.R., M.E., H.L., W.C., and M.G. contributed new reagents/analytic tools; J.Q.W. and L.H. analyzed data; and J.W., L.H., and M.S. wrote the paper.
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The authors declare no conflict of interest.
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↵*This Direct Submission article had a prearranged editor.
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Data deposition: Data were submitted to Gene Expression Omnibus (GEO; accession number GSE20301).
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This article contains supporting information online at www.pnas.org/cgi/content/full/0914114107/DCSupplemental.
