Drosophila TDP-43 RNA-Binding Protein Facilitates Association of Sister Chromatid Cohesion Proteins with Genes, Enhancers and Polycomb Response Elements

摘要

The cohesin protein complex mediates sister chromatid cohesion and participates in transcriptional control of genes that regulate growth and development. Substantial reduction of cohesin activity alters transcription of many genes without disrupting chromosome segregation. Drosophila Nipped-B protein loads cohesin onto chromosomes, and together Nipped-B and cohesin occupy essentially all active transcriptional enhancers and a large fraction of active genes. It is unknown why some active genes bind high levels of cohesin and some do not. Here we show that the TBPH and Lark RNA-binding proteins influence association of Nipped-B and cohesin with genes and gene regulatory sequences. In vitro, TBPH and Lark proteins specifically bind RNAs produced by genes occupied by Nipped-B and cohesin. By genomic chromatin immunoprecipitation these RNA-binding proteins also bind to chromosomes at cohesin-binding genes, enhancers, and Polycomb response elements (PREs). RNAi depletion reveals that TBPH facilitates association of Nipped-B and cohesin with genes and regulatory sequences. Lark reduces binding of Nipped-B and cohesin at many promoters and aids their association with several large enhancers. Conversely, Nipped-B facilitates TBPH and Lark association with genes and regulatory sequences, and interacts with TBPH and Lark in affinity chromatography and immunoprecipitation experiments. Blocking transcription does not ablate binding of Nipped-B and the RNA-binding proteins to chromosomes, indicating transcription is not required to maintain binding once established. These findings demonstrate that RNA-binding proteins help govern association of sister chromatid cohesion proteins with genes and enhancers. Author Summary The cohesin protein complex binds chromosomes to facilitate their proper division into two daughter cells when a cell divides. Cohesin and the Nipped-B protein that loads cohesin onto chromosomes also bind to genes and regions that regulate genes to ensure that genes produce proper amounts of RNA. Nipped-B and cohesin preferentially bind a subset of genes important for growth and development, and small changes in cohesin activity cause severe birth defects. Why cohesin binds some genes and not others is unknown. We tested the idea that proteins that specifically bind the RNA sequences being produced by genes may help determine which genes bind cohesin. We find that the Drosophila TBPH and Lark RNA-binding proteins bind RNA produced by genes that bind Nipped-B and cohesin. They also bind to these genes on chromosomes and the chromosomal regions that regulate them. TBPH ensures that Nipped-B and cohesin are present at high levels on regulatory regions and genes. Lark inhibits Nipped-B and cohesin binding to some genes and increases their binding to some regulatory regions. Nipped-B interacts with both RNA-binding proteins and promotes their binding to genes. These findings show that RNA-binding proteins help determine which genes bind Nipped-B and cohesin.