Distinct mechanisms govern the phosphorylation of different SR protein splicing factors

Long, Yunxin; Sou, Weng Hong; Yung, Kristen Wing Yu; Liu, Haizhen; Wan, Stephanie Winn Chee; Li, Qingyun; Zeng, Chuyue; Law, Carmen O. K.; Chan, Gordon Ho Ching; Lau, Terrence Chi-Kong; Ngo, Jacky Chi Ki

doi:10.1074/jbc.ra118.003392

Cited by 49 publications

(50 citation statements)

References 52 publications

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“…Our results therefore illustrate the utility of unbiased phosphoproteomics to elucidate cancer drug response. Recent work also suggests that additional kinases beyond the well-characterized SRPKs and CLKs may be involved in SR phosphorylation 55,56 . The mechanism that leads to coordinated, upregulated phosphorylation across multiple splicing factors in the context of cancer therapy, and the specific role of caspases in this process, will be an important topic for future investigation.…”

Section: Discussionmentioning

confidence: 99%

Proteasome inhibitor-induced modulation reveals the spliceosome as a specific therapeutic vulnerability in multiple myeloma

et al. 2020

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Enhancing the efficacy of proteasome inhibitors (PI) is a central goal in myeloma therapy. We proposed that signaling-level responses after PI may reveal new mechanisms of action that can be therapeutically exploited. Unbiased phosphoproteomics after treatment with the PI carfilzomib surprisingly demonstrates the most prominent phosphorylation changes on splicing related proteins. Spliceosome modulation is invisible to RNA or protein abundance alone. Transcriptome analysis after PI demonstrates broad-scale intron retention, suggestive of spliceosome interference, as well as specific alternative splicing of protein homeostasis machinery components. These findings lead us to evaluate direct spliceosome inhibition in myeloma, which synergizes with carfilzomib and shows potent anti-tumor activity. Functional genomics and exome sequencing further support the spliceosome as a specific vulnerability in myeloma. Our results propose splicing interference as an unrecognized modality of PI mechanism, reveal additional modes of spliceosome modulation, and suggest spliceosome targeting as a promising therapeutic strategy in myeloma.

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Section: Discussionmentioning

confidence: 99%

Proteasome inhibitor-induced modulation reveals the spliceosome as a specific therapeutic vulnerability in multiple myeloma

et al. 2020

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“…[ 6,8 ] The expression level dysregulation, activity, post‐translational modification, and subcellular localization of these splicing factors contributes to splicing decisions in different cells/tissues and pathological conditions. [ 8,9,16,17 ] SRSF3 is a major splicing factor in AS and has critical roles in the regulation of premRNA splicing. [ 12 ] In this study, we provide a novel additional mechanism by which splicing factor SRSF3 recognizes and binds its cis‐RNA elements to regulate AS.…”

Section: Discussionmentioning

confidence: 99%

Small Protein Hidden in lncRNA LOC90024 Promotes “Cancerous” RNA Splicing and Tumorigenesis

Meng

Chen

et al. 2020

Advanced Science

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Conventional therapies for late‐stage colorectal cancer (CRC) have limited effects because of chemoresistance, recurrence, and metastasis. The “hidden” proteins/peptides encoded by long noncoding RNAs (lncRNAs) may be a novel resource bank for therapeutic options for patients with cancer. Here, lncRNA LOC90024 is discovered to encode a small 130‐amino acid protein that interacts with several splicing regulators, such as serine‐ and arginine‐rich splicing factor 3 (SRSF3), to regulate mRNA splicing, and the protein thus is named “Splicing Regulatory Small Protein” (SRSP). SRSP, but not LOC90024 lncRNA itself, promotes CRC tumorigenesis and progression, while silencing of SRSP suppresses CRC tumorigenesis. Mechanistically, SRSP increases the binding of SRSF3 to exon 3 of transcription factor Sp4, resulting in the inclusion of Sp4 exon 3 to induce the formation of the “cancerous” long Sp4 isoform (L‐Sp4 protein) and inhibit the formation of the “noncancerous” short Sp4 isoform (S‐Sp4 peptide), which lacks the transactivation domain. The upregulated SRSP level is positively associated with malignant phenotypes and poor prognosis in patients with CRC. Collectively, the findings uncover that a lncRNA‐encoded small protein SRSP induces “cancerous” Sp4 splicing variant formation and may be a potential prognostic biomarker and therapeutic target for patients with CRC.

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“…Compared to peptides mapping to surrounding domains, BAD and RS domains have a complex combinatorial signature of PTM. As PTMs are an essential regulator of the structure and function of RBPs [68][69][70][71][72], we next sought to further characterize the frequency of these PTMs in the nucleoplasm proteome.…”

Section: Arg-rich Domains In Rbps Contain Combinatorial Ptmsmentioning

confidence: 99%

Middle-down proteomics reveals dense sites of methylation and phosphorylation in arginine-rich RNA-binding proteins

Kundinger

Bishof

Dammer

et al. 2019

Preprint

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Abbreviations:MS: mass spectrometry ETD: electron transfer dissociation PTM: post-translational modification AD: Alzheimer's disease LC: liquid chromatography RBP: RNA-binding proteins PSM : peptide spectral match Briefs:Middle-down proteomics reveals arginine-rich RNA-binding proteins contain many sites of methylation and phosphorylation. AbstractArginine (Arg)-rich RNA-binding proteins play an integral role in RNA metabolism. Posttranslational modifications (PTMs) within Arg-rich domains, such as phosphorylation and methylation, regulate multiple steps in RNA metabolism. However, the identification of PTMs within Arg-rich domains with complete trypsin digestion is extremely challenging due to the high density of Arg residues within these proteins. Here, we report a middle-down proteomic approach coupled with electron transfer dissociation (ETD) mass spectrometry to map previously unknown sites of phosphorylation and methylation within the Arg-rich domains of U1-70K and structurally similar RNA-binding proteins from nuclear extracts of HEK293 cells. Remarkably, the Arg-rich domains in RNA-binding proteins are densely modified by methylation and phosphorylation compared with the remainder of the proteome, with di-methylation and phosphorylation favoring RSRS motifs. Although they favor a common motif, analysis of combinatorial PTMs within RSRS motifs indicate that phosphorylation and methylation do not often co-occur, suggesting they may functionally oppose one another. Collectively, these findings suggest that the level of PTMs within Arg-rich domains may be among the highest in the proteome, and a possible unexplored regulator of RNA metabolism. These data also serve as a resource to facilitate future mechanistic studies of the role of PTMs in RNA-binding protein structure and function.Key proteins that carry out specialized biological processes such as RNA splicing, polyadenylation and transport, contain domains disproportionately enriched with arginine [1,2].RNA-binding proteins (RBPs) that harbor Arginine (Arg)-rich domains may be broadly classified into two subsets based on residue composition. One class of these RBPs contains highly repetitive complementary repeats of basic (K/R) and acidic (D/E) residues, that we have previously referred to as Basic Acidic Dipeptide (BAD) domains [3]. Importantly, BAD domains facilitate protein aggregation, and in the context of Alzheimer's disease (AD), facilitate interactions with pathological Tau protein [3]. A second subset, related to the BAD proteins, are the Arginine/serinerich (RS) domains that are ubiquitous in the Serine/Arginine (SR) family of proteins [2]. Upon serine phosphorylation, RS domains mimic BAD domains with a similarly alternating basic-acidic dipeptide sequence pattern. The RS domains are commonly found in splicing factors and are essential for alternative splicing, protein-protein interactions, and localization [4-9]. The functional and structural diversity of the proteome is markedly increased through posttranslational modifications (PTMs), which...

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Distinct mechanisms govern the phosphorylation of different SR protein splicing factors

Cited by 49 publications

References 52 publications

Proteasome inhibitor-induced modulation reveals the spliceosome as a specific therapeutic vulnerability in multiple myeloma

Proteasome inhibitor-induced modulation reveals the spliceosome as a specific therapeutic vulnerability in multiple myeloma

Small Protein Hidden in lncRNA LOC90024 Promotes “Cancerous” RNA Splicing and Tumorigenesis

Middle-down proteomics reveals dense sites of methylation and phosphorylation in arginine-rich RNA-binding proteins

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