Regulation of CD45 Alternative Splicing by Heterogeneous Ribonucleoprotein, hnRNPLL

Department of Pathology, Immune Disease Institute, Harvard Medical School, Boston, MA 02115, USA.
Science (Impact Factor: 33.61). 09/2008; 321(5889):686-91. DOI: 10.1126/science.1157610
Source: PubMed


The transition from naïve to activated T cells is marked by alternative splicing of pre-mRNA encoding the transmembrane phosphatase
CD45. Using a short hairpin RNA interference screen, we identified heterogeneous ribonucleoprotein L-like (hnRNPLL) as a critical
inducible regulator of CD45 alternative splicing. HnRNPLL was up-regulated in stimulated T cells, bound CD45 transcripts,
and was both necessary and sufficient for CD45 alternative splicing. Depletion or overexpression of hnRNPLL in B and T cell
lines and primary T cells resulted in reciprocal alteration of CD45RA and RO expression. Exon array analysis suggested that
hnRNPLL acts as a global regulator of alternative splicing in activated T cells. Induction of hnRNPLL during hematopoietic
cell activation and differentiation may allow cells to rapidly shift their transcriptomes to favor proliferation and inhibit
cell death.

Download full-text


Available from: Luis F. Moita, Jan 05, 2015
  • Source
    • "Silencing of Ptprc exons 4, 5 and 6 in T cells requires hnRNPLL, a protein with three RNA-recognition motif (RRM) domains whose mRNA expression correlates with Ptprc exon exclusion: it is highest in CD45RO+ activated and memory T cells that exclude exons 4 to 6, at intermediate levels in CD45RB+ naïve T cells, and at very low levels in CD45RABC+ B cells that include all three exons [15-17]. Mice homozygous for a destabilizing point mutation in the amino-terminal RRM domain, Hrnpllthu, fail to exclude exons 4, 5, and 6 in T-cell Ptprc mRNA and expression of CD45-RA and CD45-RC protein isoforms are increased 50-fold on different T-cell subsets [16]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Retention of a subset of introns in spliced polyadenylated mRNA is emerging as a frequent, unexplained finding from RNA deep sequencing in mammalian cells. Here we analyze intron retention in T lymphocytes by deep sequencing polyadenylated RNA. We show a developmentally regulated RNA-binding protein, hnRNPLL, induces retention of specific introns by sequencing RNA from T cells with an inactivating Hnrpll mutation and from B lymphocytes that physiologically downregulate Hnrpll during their differentiation. In Ptprc mRNA encoding the tyrosine phosphatase CD45, hnRNPLL induces selective retention of introns flanking exons 4 to 6; these correspond to the cassette exons containing hnRNPLL binding sites that are skipped in cells with normal, but not mutant or low, hnRNPLL. We identify similar patterns of hnRNPLL-induced differential intron retention flanking alternative exons in 14 other genes, representing novel elements of the hnRNPLL-induced splicing program in T cells. Retroviral expression of a normally spliced cDNA for one of these targets, Senp2, partially corrects the survival defect of Hnrpll-mutant T cells. We find that integrating a number of computational methods to detect genes with differentially retained introns provides a strategy to enrich for alternatively spliced exons in mammalian RNA-seq data, when complemented by RNA-seq analysis of purified cells with experimentally perturbed RNA-binding proteins. Our findings demonstrate that intron retention in mRNA is induced by specific RNA-binding proteins and suggest a biological significance for this process in marking exons that are poised for alternative splicing.
    Genome biology 01/2014; 15(1):R26. DOI:10.1186/gb-2014-15-1-r26 · 10.81 Impact Factor
  • Source
    • "Previous transcriptome-wide studies analyzing stimulus-induced TIV focused predominantly on immune cells [17], [18], [19], [20]. Likewise, hypoxic stress and androgen stimulation were shown to generate, after 24 hours, widespread TIV in endothelial and prostate cancer cells, respectively [21], [22]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Signal-induced transcript isoform variation (TIV) includes alternative promoter usage as well as alternative splicing and alternative polyadenylation of mRNA. To assess the phenotypic relevance of signal-induced TIV, we employed exon arrays and breast epithelial cells, which migrate in response to the epidermal growth factor (EGF). We show that EGF rapidly - within one hour - induces widespread TIV in a significant fraction of the transcriptome. Importantly, TIV characterizes many genes that display no differential expression upon stimulus. In addition, similar EGF-dependent changes are shared by a panel of mammary cell lines. A functional screen, which utilized isoform-specific siRNA oligonucleotides, indicated that several isoforms play essential, non-redundant roles in EGF-induced mammary cell migration. Taken together, our findings highlight the importance of TIV in the rapid evolvement of a phenotypic response to extracellular signals.
    PLoS ONE 12/2013; 8(12):e80566. DOI:10.1371/journal.pone.0080566 · 3.23 Impact Factor
  • Source
    • "Specifically, in resting T cells hnRNP L is solely responsible for repressing inclusion of CD45 exon 4, as hnRNP LL is not expressed at significant levels [7]. However, hnRNP LL expression is induced upon T cell activation leading to this protein also binding to the ESS1 to confer signal-induced hyper-repression of exon 4 [6,7,15]. Interestingly, we have previously shown that mutation of CA4 within ESS1 specifically disrupts the hnRNP L-mediated silencing in resting T cells [14]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: HnRNP (heterogeneous nuclear ribonucleoprotein) proteins are a large family of RNA-binding proteins that regulate numerous aspects of RNA processing. Interestingly, several paralogous pairs of hnRNPs exist that exhibit similar RNA-binding specificity to one another, yet have non-redundant functional targets in vivo. In this study we systematically investigate the possibility that the paralogs hnRNP L and hnRNP LL have distinct RNA binding determinants that may underlie their lack of functional redundancy. Using a combination of RNAcompete and native gel analysis we find that while both hnRNP L and hnRNP LL preferentially bind sequences that contain repeated CA dinucleotides, these proteins differ in their requirement for the spacing of the CAs. Specifically, hnRNP LL has a more stringent requirement for a two nucleotide space between CA repeats than does hnRNP L, resulting in hnRNP L binding more promiscuously than does hnRNP LL. Importantly, this differential requirement for the spacing of CA dinucleotides explains the previously observed differences in the sensitivity of hnRNP L and LL to mutations within the CD45 gene. We suggest that overlapping but divergent RNA-binding preferences, as we show here for hnRNP L and hnRNP LL, may be commonplace among other hnRNP paralogs.
    PLoS ONE 11/2013; 8(11):e80701. DOI:10.1371/journal.pone.0080701 · 3.23 Impact Factor
Show more