The major challenge is therefore to integrate our current knowledge about individual RBPs into concepts of higher-order gene regulation that reflect the interplay of different, and ideally of all cellular RBPs.Ī prerequisite for studying higher-order post-transcriptional networks is to know the cell type-specific RBPomes that account for differential and dynamic expression of RBPs and mRNP plasticity. Such functional or physical interactions together with interdependent binding to the transcriptome create enormous regulatory potential. RBPs can also recruit additional co-factors as for example Roquin-1 binds together with Nufip2 to RNA 24, and some of them have antagonistic RBPs like HuR and TTP 25 or Regnase-1 and Arid5a 26. Intriguingly, many key factors of the immune system have acquired long 3′-UTRs enabling their regulation by multiple, and often overlapping sets of post-transcriptional regulators 23. Underscoring the relevance for the immune system, loss-of-function of these factors has often been associated with profound alterations in T cell development or functions which caused immune-related diseases 19, 20, 21, 22. Moreover, the first evidence for m6A RNA methylation in this cell type has been provided 9. Previous studies of T helper cells have focused on a small number of RNA-binding proteins, including HuR and TTP/Zfp36l1/Zfp36l2 11, 12, 13, 14, Roquin-1/2 15, 16 and Regnase-1/4 17, 18 as well as some miRNAs like miR-17–92, miR-155, miR-181, miR-125 or miR-146a 19. These RBPs, or RBPs that recognize modifications, directly affect the expression of genes by controlling mRNA stability or translation efficiency 4, 7, 8, 9, 10. Accordingly, they employ extensive post-transcriptional regulation through RBPs or miRNAs and 3′ end oligo-uridylation or m6A RNA modifications. To exit quiescence, commit to proliferation, and exert effector functions or form memory they strongly depend on programs of gene regulation 2, 3, 4, 5, 6. T lymphocytes as central entities of the adaptive immune system must be able to make critical cell fate decisions fast 1. Connecting the cellular RBPome in a post-transcriptional context, we find contributions from multiple RBPs to the prototypic regulation of mRNA targets by individual trans-acting factors. Besides Roquin-independent contributions from Rbms1 and Cpeb4 we also show Roquin-1/2-dependent and target-specific coregulation of Icos by Celf1 and Igf2bp3. Based on proximity to Roquin-1, we select ~50 RBPs for testing coregulation of Roquin-1/2 targets by induced expression in wild-type or Roquin-1/2-deficient T cells. The RBPome includes Stat1, Stat4 and Vav1 proteins suggesting unexpected functions for these transcription factors and signal transducers. Here, we identify a core RBPome of 798 mouse and 801 human T cell proteins by utilizing global RNA interactome capture (RNA-IC) and orthogonal organic phase separation (OOPS). This is evident for the Icos mRNA encoding an essential costimulatory receptor that is regulated by several RNA-binding proteins (RBP), including Roquin-1 and Roquin-2. Post-transcriptional gene regulation in T cells is dynamic and complex as targeted transcripts respond to various factors.
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