Project name: Mus musculus

Description: Murine embryonic stem cells (ESCs) are defined by continuous self-renewal and pluripotency. A diverse repertoire of protein isoforms arising from alternative splicing are expressed in ES cells without defined biological roles. Sall4, a transcription factor essential for pluripotency, exists as two isoforms (Sall4a and Sall4b). By genome-wide location analysis, we have determined that Sall4b, and not Sall4a, binds preferentially to highly expressed loci in ES cells. Sall4a and Sall4b binding sites are distinguished by both epigenetic marks at target loci and their clustering with binding sites of other pluripotency factors. When ESCs expressing a single isoform of Sall4 are generated, Sall4b alone could maintain the pluripotent state, although it could not completely suppress all differentiation markers. Sall4a and Sall4b collaborate in maintenance of the pluripotent state, but play distinct roles. Our work is novel in establishing such isoform-specific differences in ES cells.Overall design: Cells infected with an empty lentivirus (Wt) are compared with cells missing both isoforms of Sall4 (WT+shRNA) or a single isoform (Sall4aIMM+shRNA or Sall4bIMM+shRNA). ES cells expressing a single isoform of Sall4 (either Sall4a or Sall4b) were generated by using a lentivirally based shRNA (Open Biosystems TRC0000097821) that depleted both endogenous isoforms and then adding back a single isoform cDNA that was immune to the shRNA. Transcriptome analysis was performed 48 hrs after infection to identify changes in gene expression at various loci when only a single isoform of Sall4 was present.

Data type: Transcriptome or Gene expression

Sample scope: Multiisolate

Relevance: ModelOrganism

Release date: 2010-09-13

Last updated: 2010-03-24