The Role of Polycomp-like 3 and Ofd1 in Embryonic Stem Cell Maintenance and Differentiation
Embryonic stem cells (ESCs) hold tremendous promise for cell based therapeutics and as a biological tool. As differentiation of ESCs mimics early development, ESCs also serve as an excellent tool to study the underlying mechanisms of many developmental processes and cancer. The transition from ESC to somatic cell requires proper signaling as well as many intricate layers of gene regulation. Clarifying the interplay between these different processes is important for understanding the many transitions that happen during development as well as those that occur upon cancer metastasis.
, To elucidate some of the molecular mechanisms of ESC differentiation, we explored different areas of regulation within ESCs including cell signaling as well as epigenetic and transcription gene regulation. To best investigate these areas, we developed a recombinant gene technology called the Floxin system, in which we specifically and efficiently tagged genes within their endogenous loci in ESCs. The gene trap and tagged ESC lines were used to identify novel binding partners and investigate disease alleles.
Through the Floxin system, we demonstrate that Ofd1, a protein required for ciliogenesis, restrains neural differentiation in embryoid bodies (EBs). We also show that Ofd1 is essential for proper Hh and Wnt signaling and that disease alleles of Ofd1 cannot rescue proper neural differentiation. Secondly, we investigate Polycomb-like 3 (Pcl3), a protein involved in histone modifications and gene repression. We found that Pcl3 incorporates into Polycomb repressive complex 2 (PRC2) and promotes complex function. Diminishment of Pcl3 decreased PRC2 binding as well as ESC self-renewal, indicating that Pcl3 enhances PRC2 targeting and ESC maintenance. Finally, we identify novel binding partners of Sall4, a transcription factor important for ESC pluripotency and reprogramming.
Notably, Ofd1, Pcl3, and Sall4 are all associated with human conditions and cancer. Thus besides understanding how these proteins function in ESC maintenance and differentiation, we have gained insight into the mechanisms by which disruption of these proteins contributes to human disease.