A growing body of evidence supports the coordination of pre-mRNA processing and transcriptional regulation. We demonstrate here that mammalian PRP4 kinase (PRP4K) is associated with complexes involved in both of these processes. PRP4K is implicated in pre-mRNA splicing as the homologue of the Schizosaccharomyces pombe pre-mRNA splicing kinase Prp4p, and it is enriched in SC35-containing nuclear splicing speckles. RNA interference of Caenorhabditis elegans PRP4K indicates that it is essential in metazoans. In support of a role for PRP4K in pre-mRNA splicing, we identified PRP6, SWAP, and pinin as interacting proteins and demonstrated that PRP4K is a U5 snRNP-associated kinase. In addition, BRG1 and N-CoR, components of nuclear hormone coactivator and corepressor complexes, also interact with PRP4K. PRP4K coimmunoprecipitates with N-CoR, BRG1, pinin, and PRP6, and we present data suggesting that PRP6 and BRG1 are substrates of this kinase. Lastly, PRP4K, BRG1, and PRP6 can be purified as components of the N-CoR-2 complex, and affinity-purified PRP4K/N-CoR complexes exhibit deacetylase activity. We suggest that PRP4K is an essential kinase that, in association with the both U5 snRNP and N-CoR deacetylase complexes, demonstrates a possible coordination of pre-mRNA splicing with chromatin remodeling events involved in transcriptional regulation.The regulation of eukaryotic gene expression involves the modulation of chromatin structure and the coordinated transcription and splicing of mRNA. Both histone modification (61) and chromatin remodeling modulate the access of the transcriptional apparatus to chromatin (14, 44). The SWI/SNF proteins are the archetypal family of ATPases involved in remodeling chromatin, and members of this family in both yeast (Snf2) and mammals (BRG1) have been isolated in association with the RNA polymerase II holoenzyme complex (45, 59). Once transcription has been initiated it is thought that premRNA splicing occurs cotranscriptionally (4,22). A number of recent studies have also shown links between corepressor and coactivator complexes and pre-mRNA splicing (e.g., PSF [40] and PGC1 [42], respectively), which suggests yet another level of coordination between the regulation of gene expression and pre-mRNA splicing.If chromatin remodeling and pre-mRNA splicing occur in coordination with transcription, there may be proteins that either play a direct role in both chromatin structure and splicing and/or interact with factors involved in each process. Previously, we isolated a murine gene-trap protein, designated CT143 (56), which is similar to Schizosaccharomyces pombe Prp4 kinase (17). Prp4p is essential for pre-mRNA splicing, as demonstrated by the accumulation of unspliced pre-mRNA at the restrictive temperature in yeast carrying a temperaturesensitive (ts) mutation in prp4 (2). Prp4p interacts genetically with other S. pombe splicing proteins, including the non-SR splicing proteins Prp1p (a homologue of Saccharomyces cerevisiae Prp6p) and Prp5p (51). Multiple rounds of phosphorylation and d...
Zinc-finger protein 217 (ZNF217) is a Kruppel-like zincfinger protein located at 20q13.2, within a region of recurrent maximal amplification. Here, we demonstrate that ZNF217 is a transcriptional repressor protein and report the purification and characterization of a ZNF217 complex. The purified ZNF217 complex consists of approximately six proteins and contains the transcriptional co-repressors CoREST, BHC110/LSD1, histone deacetylase (HDAC) 2 and C-terminal binding protein (CtBP1). The purified ZNF217 complex possesses deacetylase activity as well as lysine 4 histone H3-specific demethylase activity that is most likely mediated by the BHC110/LSD1 component. To determine if ZNF217 is a sequence-specific binding protein, we have made use of cyclic amplification and selection of targets (CAST) assay and identify for the first time a ZNF217 DNA consensus recognition sequence (CRS) that is highly conserved in the human E-cadherin promoter. Chromatin immunoprecipitation (ChIP) experiments demonstrate that ZNF217, as well as the other components of the ZNF217 complex, are found on the region of the proximal E-cadherin promoter that contains the identified ZNF217 CRS in vivo. Using a combination of transient transfections and small interfering RNA, we demonstrate that ZNF217 represses the E-cadherin promoter. Collectively, our results implicate ZNF217 and its associated proteins in a novel pathway that may have profound effects on cancer progression.
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