SF2 and SRp55 regulation of CD45 exon 4 skipping during T cell activation

Abstract

CD45 is an alternatively spliced membrane phosphatase required for T cell activation. Exons 4, 5 and 6 can be included or skipped from spliced mRNA resulting in several protein isoforms that include or exclude epitopes referred to as RA, RB or RC, respectively. T cells reciprocally express CD45RA or CD45RO (lacking all three exons), corresponding to naive versus memory T cells. Overexpression of the alternative splicing regulators, SF2 or SWAP, induces skipping of CD45 exon 4 in transfected COS cells. We show here that the arginine/serine-rich domain of SWAP and the RNA recognition motifs of SF2 are required for skipping of CD45 exon 4. Unlike SWAP, SF2 specifically regulated alternative splicing of CD45 exon 4, having no effect on a non-regulated exon of CD45 (exon 9). Like SF2 and SWAP, the SR proteins SC35, SRp40 and SRp75, but not SRp55 also induced CD45 exon 4 skipping. In contrast, antisense inhibition of SRp55 induced exon 4 skipping. SF2 and SRp55 proteins were not detectable or expressed at a very low level in freshly isolated CD45RA+ and CD45RO+ T cells. Activation of CD45RA+ T cells shifted CD45 expression from CD45RA to CD45RO, and induced a large increase in expression of both SF2 and SRp55. Thus, SF2 at least in part determines splicing of CD45 exon 4 during T cell activation. SRp55, SR protein phosphorylation, or other splicing factors likely regulate CD45 splicing in CD45RO+ memory T cells. The different SR proteins expressed by memory and activated T cells emphasize the different phenotypes of these cell types that both express CD45RO.

Silencers regulate both constitutive and alternative splicing events in mammals

“…Moreover, some reports have suggested that SR protein interaction with exonic sequences can result in splice silencing. In the case of CD45 [74,75] inclusion of three alternative exons (A, B and C) is regulated upon T cell activation and depends on SR proteins; in particular, some of these proteins act as repressors, while others stimulate inclusion of the alternative exons (table 1). The cis-acting elements that mediate SR-protein binding and splicing regulation are presently unknown, although some evidence [76,77] indicates that they might be located within exonic sequences (at least in the case of exons 4 and 6).…”

“…For example, distinct SR proteins can counteract each other, exerting opposite effects on the inclusion of three alternative exons in CD45 mRNA (table 1); accordingly, changes in SR protein expression are observed upon T cell activation, i.e. when a shift in CD45 splicing pattern is observed [74]. Antagonistic effects can also be exerted by trans-acting factors that act alternatively as both repressors and activators on different target RNAs.…”

Alternative expression of TCRζ related genes in patients with chronic myeloid leukemia

“…ASF/SF2 acts early during spliceosome assembly and participates in multiple steps during constitutive splicing and the regulation of alternative splicing by interacting with the pre-mRNAs and/or other splicing factors [28,30,31]. ASF/SF2 regulates the alternative splicing of eukaryotic genes such as caspase-9 and the T cell differentiation marker CD45 [19,32,33]. Recently, the involvement of ASF/SF2 in the post-transcriptional regulation of TCRζ was described in T cells from patients with SLE.…”

Conserved Sr Protein Kinase Functions in Nuclear Import and Its Action Is Counteracted by Arginine Methylation inSaccharomyces cerevisiae

“…Regulation of SR and hnRNP proteins and the functional consequences associated with it are just beginning to be understood. Although most SR proteins are ubiquitously expressed, some are expressed in a tissue-specific manner (Ayane et al, 1991), during T cell development (Lemaire et al, 1999), or induced in response to mitogens (Diamond et al, 1993;Screaton et al, 1995). All RS domain-containing proteins are posttranslationally modified by phosphorylation.…”