{"title":"BioID 接近性图谱揭示了前接合体复合体中新型 SAP18 相互作用。","authors":"Sweta Kumari, Ankita Adhikary, Kusum Kumari Singh","doi":"10.1016/j.bbrc.2024.150944","DOIUrl":null,"url":null,"abstract":"<div><div>SAP18 protein was originally discovered in association with the SIN3 transcriptional repressor complex. Subsequent biochemical fractionation studies identified SAP18 as a component of another distinct trimeric complex termed as the apoptosis- and splicing-associated protein (ASAP) complex. The existence of SAP18 in distinct complexes highlights its dual role in transcriptional and splicing regulation. In our study, we aim to define the <em>in vivo</em> interactome of SAP18 using proximity-dependent biotin identification (BioID). Mass spectrometry analysis of streptavidin-purified biotinylated proteins revealed new SIN3-associated interactors, including RBBP4 and SAP30BP. Notably, we identified 72 spliceosomal proteins as highly enriched interactors. Additionally, a complementary immunoprecipitation assay validated novel interactions of SAP18 with the prespliceosomal components SNRNP70, SNRPA, SF3B1, U2AF1, and the SR protein SRSF1. Mutational analysis using a C-terminal SAP18 double point mutant, which is known to be deficient in ASAP-interaction, demonstrated a debilitated interaction with the prespliceosomal proteins. Altogether, our results present a refined understanding of the SAP18 interactome, uncovering its association with the prespliceosome in conjugation with ASAP components.</div></div>","PeriodicalId":8779,"journal":{"name":"Biochemical and biophysical research communications","volume":"738 ","pages":"Article 150944"},"PeriodicalIF":2.5000,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"BioID proximity mapping reveals novel SAP18 interactions in the prespliceosomal complex\",\"authors\":\"Sweta Kumari, Ankita Adhikary, Kusum Kumari Singh\",\"doi\":\"10.1016/j.bbrc.2024.150944\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>SAP18 protein was originally discovered in association with the SIN3 transcriptional repressor complex. Subsequent biochemical fractionation studies identified SAP18 as a component of another distinct trimeric complex termed as the apoptosis- and splicing-associated protein (ASAP) complex. The existence of SAP18 in distinct complexes highlights its dual role in transcriptional and splicing regulation. In our study, we aim to define the <em>in vivo</em> interactome of SAP18 using proximity-dependent biotin identification (BioID). Mass spectrometry analysis of streptavidin-purified biotinylated proteins revealed new SIN3-associated interactors, including RBBP4 and SAP30BP. Notably, we identified 72 spliceosomal proteins as highly enriched interactors. Additionally, a complementary immunoprecipitation assay validated novel interactions of SAP18 with the prespliceosomal components SNRNP70, SNRPA, SF3B1, U2AF1, and the SR protein SRSF1. Mutational analysis using a C-terminal SAP18 double point mutant, which is known to be deficient in ASAP-interaction, demonstrated a debilitated interaction with the prespliceosomal proteins. Altogether, our results present a refined understanding of the SAP18 interactome, uncovering its association with the prespliceosome in conjugation with ASAP components.</div></div>\",\"PeriodicalId\":8779,\"journal\":{\"name\":\"Biochemical and biophysical research communications\",\"volume\":\"738 \",\"pages\":\"Article 150944\"},\"PeriodicalIF\":2.5000,\"publicationDate\":\"2024-11-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Biochemical and biophysical research communications\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0006291X24014803\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biochemical and biophysical research communications","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0006291X24014803","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
BioID proximity mapping reveals novel SAP18 interactions in the prespliceosomal complex
SAP18 protein was originally discovered in association with the SIN3 transcriptional repressor complex. Subsequent biochemical fractionation studies identified SAP18 as a component of another distinct trimeric complex termed as the apoptosis- and splicing-associated protein (ASAP) complex. The existence of SAP18 in distinct complexes highlights its dual role in transcriptional and splicing regulation. In our study, we aim to define the in vivo interactome of SAP18 using proximity-dependent biotin identification (BioID). Mass spectrometry analysis of streptavidin-purified biotinylated proteins revealed new SIN3-associated interactors, including RBBP4 and SAP30BP. Notably, we identified 72 spliceosomal proteins as highly enriched interactors. Additionally, a complementary immunoprecipitation assay validated novel interactions of SAP18 with the prespliceosomal components SNRNP70, SNRPA, SF3B1, U2AF1, and the SR protein SRSF1. Mutational analysis using a C-terminal SAP18 double point mutant, which is known to be deficient in ASAP-interaction, demonstrated a debilitated interaction with the prespliceosomal proteins. Altogether, our results present a refined understanding of the SAP18 interactome, uncovering its association with the prespliceosome in conjugation with ASAP components.
期刊介绍:
Biochemical and Biophysical Research Communications is the premier international journal devoted to the very rapid dissemination of timely and significant experimental results in diverse fields of biological research. The development of the "Breakthroughs and Views" section brings the minireview format to the journal, and issues often contain collections of special interest manuscripts. BBRC is published weekly (52 issues/year).Research Areas now include: Biochemistry; biophysics; cell biology; developmental biology; immunology
; molecular biology; neurobiology; plant biology and proteomics