The RNA binding protein RBM38 (RNPC1) regulates splicing during late erythroid differentiation

ABSTRACT: Alternative pre-mRNA splicing is a prevalent mechanism in mammals that promotes proteomic diversity, including expression of cell-type specific protein isoforms. We characterized a role for RBM38 (RNPC1) in regulation of alternative splicing during late erythroid differentiation. We used an affymetrix human exon junction (HJAY) splicing microarray to identify a panel of RBM38-regulated alternatively spliced transcripts. Using microarray databases, we noted high RBM38 expression levels in CD71+ erythroid cells and thus chose to examine RBM38 expression during erythroid differentiation of human hematopoietic stem cells, detecting enhanced RBM38 expression during late erythroid differentiation. In differentiated erythroid cells, we validated a subset of RBM38-regulated splicing events and determined that RBM38 regulates activation of Protein 4.1R (EPB41) exon 16 during late erythroid differentiation. Using Epb41 minigenes, Rbm38 was found to be a robust activator of exon 16 splicing. To further address the mechanism of RBM38-regulated alternative splicing, a novel mammalian protein expression system, followed by SELEX-Seq, was used to identify a GU-rich RBM38 binding motif. Lastly, using a tethering assay, we determined that RBM38 can directly activate splicing when recruited to a downstream intron. Together, our data support the role of RBM38 in regulating alternative splicing during erythroid differentiation. siRNA knockdown of RBM38 was perfomed in human MCF-7 breast cancer cells. The efficiency of RBM38 knockdown was monitored by western blot using an RBM38 antibody (Santa Cruz Biotechnology, SC-85873). We conducted HJAY exon and exon junction array profiling on RNAs from four siRBM38 treated MCF-7 samples vs. four sicontrol treated MCF-7 samples Control / knockdown comparison.

ORGANISM(S): Homo sapiens

SUBMITTER: Carstens Russ 

PROVIDER: S-ECPF-GEOD-49293 | biostudies-other |

REPOSITORIES: biostudies-other