Induced transcription and stability of CELF2 mRNA drives widespread alternative splicing during T-cell signaling

Abstract

Studies in several cell types have highlighted dramatic and diverse changes in mRNA processing that occur upon cellular stimulation. However, the mechanisms and pathways that lead to regulated changes in mRNA processing remain poorly understood. Here we demonstrate that expression of the splicing factor CELF2 (CUGBP, Elav-like family member 2) is regulated in response to T-cell signaling through combined increases in transcription and mRNA stability. Transcriptional induction occurs within 6 h of stimulation and is dependent on activation of NF-κB. Subsequently, there is an increase in the stability of the CELF2 mRNA that correlates with a change in CELF2 3′UTR length and contributes to the total signalinduced enhancement of CELF2 expression. Importantly, we uncover dozens of splicing events in cultured T cells whose changes upon stimulation are dependent on CELF2 expression, and provide evidence that CELF2 controls a similar proportion of splicing events during human thymic T-cell development. Taken together, these findings expand the physiologic impact of CELF2 beyond that previously documented in developing neuronal and muscle cells to T-cell development and function, identify unappreciated instances of alternative splicing in the human thymus, and uncover novel mechanisms for CELF2 regulation that may broadly impact CELF2 expression across diverse cell types.alternative splicing | CELF2 | thymic development | T cells | mRNA stability

Activation-Dependent TRAF3 Exon 8 Alternative Splicing Is Controlled by CELF2 and hnRNP C Binding to an Upstream Intronic Element

“…3A and B). Consistent with previous observations (29,30), we found CELF2 expression to be increased upon T cell activation ( Fig. 3C) providing a possible explanation for increased binding in our cross-link assays.…”

“…This idea is consistent with the recent finding that overexpressing human CELF2 in mouse heart leads to substantial changes in the global splicing pattern; among the potential target exons found in this study was TRAF3 exon 8, which was not found when overexpressing CELF1 (28). It should be noted, however, that there is only little overlap between the exons regulated by CELF2 overexpression in the heart and CELF2 knockdown in T cells (28,30), which may indicate a tissue-specific regulation of CELF2 activity. One possibility is alternative splicing of CELF2 itself (27,39), which, based on reference sequences, gives rise to many different isoforms.…”

“…CELF2 has been suggested to regulate alternative splicing of many targets during T cell activation (30). We therefore investigated whether expression of CELF2 in Ramos B cells is sufficient to regulate splicing of additional exons.…”

“…We therefore investigated whether expression of CELF2 in Ramos B cells is sufficient to regulate splicing of additional exons. We chose additional targets that have been described to be responsive to T cell activation and/or CELF2 knockdown, namely, PPP1R12A, BRD8, FAM36, and REPS1 (30,34). PPP1R12A showed strongly increased exon exclusion in the CELF2-overexpressing cells, whereas the other targets showed a consistent but weaker response to CELF2 expression ( Fig.…”

“…CELF2 controls a variety of alternative exons with functional implications for differentiation and functionality of the respective tissues (28). CELF2 expression is also increased upon differentiation of primary human thymocytes and activation of a model T cell line (Jsl1), where it regulates alternative splicing of the transcription factor Lef1 and other targets (29,30).…”