Pleiohomeotic Can Link Polycomb to DNA and Mediate Transcriptional Repression

Abstract

Polycomb group (PcG) proteins function through cis-acting DNA elements called PcG response elements (PREs) to stably silence developmental regulators, including the homeotic genes. However, the mechanism by which they are targeted to PREs remains largely unclear. Pleiohomeotic (PHO) is a sequence-specific DNAbinding PcG protein and therefore may function to tether other PcG proteins to the DNA. Here, we show that PHO can directly bind to a Polycomb (PC)-containing complex as well as the Brahma (BRM) chromatinremodeling complex. PHO contacts the BRM complex through its zinc finger DNA-binding domain and a short N-terminal region. A distinct domain of PHO containing a conserved motif contacts the PcG proteins PC and Polyhomeotic (PH). With mobility shift assays and DNA pulldown experiments, we demonstrated that PHO is able to link PC, which lacks sequence-specific DNA-binding activity, to the DNA. Importantly, we found that the PC-binding domain of PHO can mediate transcriptional repression in transfected Drosophila Schneider cells. Concomitant overexpression of PC resulted in stronger PHO-directed repression that was dependent on its PC-binding domain. Together, these results suggest that PHO can contribute to PRE-mediated silencing by direct recruitment of a PC complex to repress transcription.

Adaptive selection and coevolution at the proteins of the Polycomb repressive complexes in Drosophila

“…In contrast, Context Mirror predicts the PHO/SFMBT interaction (Alfieri et al, 2013) and the well-characterized interactions between these two subunits of the PhoRC complex and those of PRC1: PHO/PC (Mohd-Sarip et al, 2002), PHO/PH-P (Mohd- Sarip et al, 2002) and SFMBT/SCM (Grimm et al, 2009). The bioinformatic method also predicts the well-established interactions between subunits of the PRC1 complex, that is, PH-P/PC (Strutt and Paro, 1997), PH-P/PSC (Kyba and Brock, 1998), PH-P/SCM (Peterson et al, 1997), PC/PSC (Kyba and Brock, 1998), PC/SCE and PSC/SCE (Gorfinkel et al, 2004).…”

“…Thereafter, the H3K27me3 hallmark deposited by Pcl-PRC2 would trigger the recruitment of PRC1 by means of the chromodomain of PC. Nevertheless, PHO interacts not only with subunits of the PRC2 complex (Wang et al, 2006) but also with subunits of the PRC1 complex (Mohd-Sarip et al, 2002), which could also contribute to anchor PRC1 at PRE. In addition, it has been proposed that the recruitment of PcG complexes to chromatin might require additional not yet identified proteins (Wang et al, 2010).…”

Comparing active and repressed expression states of genes controlled by the Polycomb/Trithorax group proteins

“…We observe that in particular the Pho protein depicts a distribution profile that reflects the expression state of the target gene. Several studies suggest the DNA binding protein Pho as a targeting factor for PRC1 (24)(25)(26), whereas others have shown, by immunostaining of polytene chromosomes, that the binding of Pc, Ph, and Psc to most sites is not altered in pho, pho-like double mutants (pho-like encodes a Pho-related protein, which binds to the same DNA motif) (27). So far our knowledge about Pho binding sites on chromatin has been limited to low-resolution polytene chromosome stainings and the PCR analysis of Ubx sequences and the iab-7 PRE after ChIPs (12,16,26).…”

The Pleiohomeotic Functions as a Negative Regulator of Drosophila even-skipped Gene during Embryogenesis

“…Of the molecularly characterized PcG proteins, only Pleiohomeotic (Pho) and its homolog, Pho-like (Phol), have been found to have DNA binding domains (Brown et al, 1998;. Pho binds directly to PRC1 as well as to the Brahma (BRM) chromatin remodeling complex in Drosophila embryonic nuclear extracts (Mohd-Sarip et al, 2002). Pho also directly binds to a PRC2 in the wing imaginal disc (Wang et al, 2004).…”