The Drosophila cohesin subunit Rad21 is a trithorax group (trxG) protein

Abstract

The cohesin complex is a key player in regulating cell division. Cohesin proteins SMC1, SMC3, Rad21, and stromalin (SA), along with associated proteins Nipped-B, Pds5, and EcoI, maintain sister chromatid cohesion before segregation to daughter cells during anaphase. Recent chromatin immunoprecipitation (ChIP) data reveal extensive overlap of Nipped-B and cohesin components with RNA polymerase II binding at active genes in Drosophila. These and other data strongly suggest a role for cohesion in transcription; however, there is no clear evidence for any specific mechanisms by which cohesin and associated proteins regulate transcription. We report here a link between cohesin components and trithorax group (trxG) function, thus implicating these proteins in transcription activation and/or elongation. We show that the Drosophila Rad21 protein is encoded by verthandi (vtd), a member of the trxG gene family that is also involved in regulating the hedgehog (hh) gene. In addition, mutations in the associated protein Nipped-B show similar trxG activity i.e., like vtd, they act as dominant suppressors of Pc and hh Mrt without impairing cell division. Our results provide a framework to further investigate how cohesin and associated components might regulate transcription.Hedgehog ͉ heterochromatin ͉ Nipped-B ͉ Polycomb ͉ cohesion I n eukaryotic mitosis, accurate chromosome segregation requires paired sister chromatids to attach to opposite spindle poles. Sister chromatids are held together by the cohesin protein complex, which consists of four core subunits, Rad21/SCC1, stromalin (SA) and structural maintenance of chromosome (SMC) proteins SMC1 and SMC3. A widely accepted model postulates that cohesin forms a ring-like structure via interaction of the N-and C-termini of Rad21 with a SMC1/SMC3 heterodimer. With the participation of SCC2/Nipped-B, SCC4, EcoI/Ctf7, and Pds5 proteins, sister-chromatid cohesion is maintained until the onset of mitosis. Cleavage of Rad21 and the resulting removal of cohesin then allow separation of sister chromatids in anaphase (see refs. 1-3 for reviews). Mutation of genes encoding these subunits leads to errors in chromosome segregation and aneuploidy, which are hallmarks of cancer and a leading cause of birth defects in humans (4).Given the highly conserved role for cohesin in sister chromatid cohesion, it was unexpected to discover that cohesin and associated proteins might also play a distinct, independent role in regulating gene expression. Reduction in Nipped-B expression in Drosophila affects expression of the cut and Ultrabithorax genes (5-7), and mutations in the human orthologue, NIPBL, result in Cornelia de Lange Syndrome (8). In zebrafish, mutations in rad21 or Smc3 affect embryonic runx gene transcription in heterozygous mutant animals without compromising cell division, suggesting that these proteins may have functions in transcription that are distinct from a mitotic role (9). Recently, Misulovin et al. observed extensive overlap of Nipped-B and cohesin components with RNA polymer...

Polycomb purification by in vivo biotinylation tagging reveals cohesin and Trithorax group proteins as interaction partners

“…Besides its traditional role in sister chromatid cohesion, cohesin has also been implicated in both activation and repression of transcription (28). Furthermore, mutations in the Rad21 subunit of the cohesin complex strongly enhance TrxG (29) and suppress PcG loss of function phenotypes. Pc and cohesins are not colocalized on salivary gland chromatin, and removal of cohesin does not affect Pc binding.…”

“…Besides its role in sister chromatid cohesion there is accumulating evidence from multiple organisms that cohesin and associated factors have diverse roles in gene regulation, organization of chromatin structure and development (27,28). One example is the genetic interaction of Rad21 with TrxG mutations in Drosophila (29). In addition, we show that the Rad21ex3 and (A) Western blot analysis after streptavidin PD of 7.5 mg of NE using antibodies against Rad21, Pho, dSfmbt, and Fs(1)h. In the following Co-IPs, 1 mg of NE was used as IN and 50% of the PD eluates were loaded to the gels.…”

Drosophila O -GlcNAc transferase (OGT) is encoded by the Polycomb group (PcG) gene, super sex combs ( sxc )

“…Genomic DNA was isolated from individual embryos of each strain and PCR was used (15) to identify samples originating from homozygous embryos containing sxc defects. Primers were designed to amplify the GFP transgene (330-bp fragment) and an X-linked gene used to evaluate DNA integrity, Grp84 (215-bp fragment) as previously described (34). DNA specimens for which PCR amplification detected the presence of Grp84 and the absence of the GFP transgene were used in subsequent analysis.…”

Does cohesin regulate developmental gene expression in Drosophila ?

“…Two articles published earlier this year demonstrated that cohesin mutations in Drosophila disrupt axon pruning at a postmitotic stage in development of the Drosophila nervous system by inhibiting expression of the Ecdysone-Receptor (EcR) gene (7,8). In this issue of PNAS, Hallson et al (9) suggest a considerably broader role of cohesin in gene regulation in Drosophila. Their findings indicate that mutations in the gene encoding the Rad21 cohesin subunit interact with mutations in hedgehog (hh), a signal-transduction gene with multiple developmental roles (10), and in Polycomb (Pc), which encodes a component of a chromatin complex responsible for silencing expression of multiple homeotic genes during development.…”

“…These phenotypes are characteristic of mutations in trithorax-group (trxG) genes, which encode a diverse group of chromatin proteins required to counteract Polycomb group (PcG)-mediated silencing and maintain appropriate expression of several genes important for proper identity of Drosophila segments and other developmental processes (11)(12)(13). Thus, the findings of Hallson et al (9) suggest that cohesin may play a fundamental role in developmental gene regulation in addition to its established role in chromosome segregation.…”