Polycomb Regulation of Embryonic Stem Cell Differentiation

Polycomb group of proteins, which has been conserved during evolution, is important in regulating the expression of homeotic genes during development of the fruit fly embryo. This paper and an accompanying report in Nature, extend our knowledge of how these proteins regulate mammalian cell differentiation.

This study focused on the Polycomb Repressive Complex 2 (PRC2), which silences genes via histone H3 lysine-27 methylation. Specifically, the work examined the SUZ12 component, which is a suppressor of zeste 12 and is highly correlated with the presence of the other two PRC2 components, EED and EZH2. Mapping of the binding of SUZ12 to the genome of human embryonic stem cells shows that PRC2 is associated with about 8% of the 22,500 annotated genes studied, while RNA polymerase II is associated with 32%. Just as important, there is relatively little overlap between the two binding patterns. This, plus a finding that 95% of SUZ12 binding occurs within 1kb of a gene promoter, suggests that PRC2 acts by preventing initiation of gene transcription.

In most instances, PRC2 occupied genes were underexpressed in embryonic stem cells, relative to their expression in differentiated neuronal, muscle, immune, and reproductive tissue. In cases in which the opposite was true, SUZ12 binding was less extensive and/or RNA polymerase II was present. Thus, PRC2 appears to promote gene silencing in most of the targeted genes of the embryonic cell.

The types of genes to which SUZ12 binds include a large number that control development and transcription. Indeed, it binds to about 50% of the genes associated with controlling developmental processes, including 39 of 40 homeodomain genes that regulate neurogenesis, hematopoiesis, axial patterning, tissue patterning, organogenesis, and cell-fate specification. It was also found associated with the FOX, SOX, and TBX gene families. SUZ12 binding to these genes promoter regions is highly defined spatially, though it is extensive (in the order of 100 kb) in some instances. The PRC2 targets also include a number of genes encoding for signaling pathways. Among these are transforming growth factor–beta, bone morphogenic protein, wingless-type MMTV integration site (Wnt), and fibroblast growth factor.

An examination of gene activity in cultured stem cells lends further support to the role of PRC2 in regulating embryogenesis. During differentiation, genes previously found occupied by SUZ12 were more likely to be activated than genes not associated with this protein. Indeed, 36% of genes bound by SUZ12 showed greater than two-fold increases in expression during differentiation, versus 16% not bound by it. Moreover, SUZ12 occupied 83% of developmental regulators that were induced by more than ten-fold during embryonic stem cell differentiation. On the other hand, differentiated muscle cells showed diminished SUZ12 binding to the MYOD1 gene, which is the master regulator of muscle differentiation, while binding to regulators of nonmuscle differentiation remained unchanged (at a high level).

Finally, binding sites of SUZ12 were compared with those of three transcription factors, OCT4, SOX2, and NANOG, to gain additional support for the role of PRC2 in embryonic stem cell regulation. SUZ12 bound to a many of the genes that are under the influence of the three transcription factors.

Combined, the results of this study and the aforementioned Nature paper, which focused on mouse embryonic stem cells, establish the importance of PRC12 in the regulation of embryogenesis and in the maintenance of pluripotent stem cells in culture.

Message posted by: Keith Markey