Anti-CSTF2 monoclonal antibody, clone 4E2-4B7

Pre-mRNA processing often occurs in coordination with transcription thereby coupling these two key regulatory events. As such, many proteins involved in mRNA processing associate with the transcriptional machinery and are in proximity to DNA. This proximity allows for the mapping of the genomic associations of RNA binding proteins by chromatin immunoprecipitation (ChIP) as a way of determining their sites of action on the encoded mRNA. Here, we used ChIP combined with high-density microarrays to localize on the human genome three functionally distinct RNA binding proteins: the splicing factor polypyrimidine tract binding protein (PTBP1/hnRNP I), the mRNA export factor THO complex subunit 4 (ALY/THOC4), and the 3' end cleavage stimulation factor 64 kDa (CSTF2). We observed interactions at promoters, internal exons, and 3' ends of active genes. PTBP1 had biases toward promoters and often coincided with RNA polymerase II (RNA Pol II). The 3' processing factor, CSTF2, had biases toward 3' ends but was also observed at promoters. The mRNA processing and export factor, ALY, mapped to some exons but predominantly localized to introns and did not coincide with RNA Pol II. Because the RNA binding proteins did not consistently coincide with RNA Pol II, the data support a processing mechanism driven by reorganization of transcription complexes as opposed to a scanning mechanism. In sum, we present the mapping in mammalian cells of RNA binding proteins across a portion of the genome that provides insight into the transcriptional assembly of RNA-protein complexes.

Alternative polyadenylation (APA) is how genes choose different sites for 3 ' end formation for mRNAs during transcription. APA often occurs in a tissue- or developmental stage-specific manner that can significantly affect gene activity by changing the protein product generated, the stability of the transcript, its localization within the cell, or its translatability. Despite the important regulatory effects that APA has on tissue-specific gene expression, only a few examples have been characterized mechanistically. In this 2018 update to our 2010 review, we examine mechanisms for the control of APA and update our understanding of the older mechanisms since 2010. We once postulated the existence of tissue-specific factors in APA. However, while a few tissue-specific polyadenylation factors are known, the emerging conclusion is that the majority of APA is accomplished by altering levels of core polyadenylation proteins. Examples of those core proteins include CSTF2, CPSF1, and subunits of mammalian cleavage factor I. But despite support for these mechanisms, no one has yet documented any of these proteins changing in either a tissue-specific or developmental manner. Given the profound effect that APA can have on gene expression and human health, improved understanding of tissue-specific APA could lead to numerous advances in gene activity control. This article is categorized under: RNA Processing > 3 ' End Processing RNA in Disease and Development > RNA in Development