剪接子成分USP39在B细胞发育和分化中的作用及机制研究

成人的B细胞发育自骨髓造血干细胞开始，分化形成分泌抗体的浆细胞，经历了诸如B细胞受体基因重排、转录调控以及转录后调控等一系列的调控与筛选。剪接子介导的RNA剪接是转录后调控中重要的一环，主要负责去除前体mRNA中内含子。此外，RNA剪接也与转录、染色质塑形有关。近年来，越来越多的研究表明，RNA剪接在免疫细胞的发育分化中有着重要的调控作用。而异常的RNA剪接也与诸如免疫细胞肿瘤、免疫缺陷病等疾病的发生息息相关。本文中所关注的泛素特异性肽酶39（USP39）是剪接子中U4/U6.U5小核核糖核蛋白三聚体（tri-snRNP）的组分之一，对tri-snRNP的组装以及活化剪接子前体的形成有着重要的调控作用。近年来有诸多报道表明USP39参与了肿瘤的发生和发展，然而USP39在B细胞发育和分化中扮演怎样的角色、通过何种机制进行调控尚有待探索。

本研究首先通过Cre-lox敲除系统构建了在B细胞中特异性敲除Usp39的转基因小鼠，研究了Usp39在B细胞中的生物学功能。实验结果表明，在B细胞发育早期敲除Usp39导致了骨髓及外周淋巴器官中B细胞数量的显著减少，但并不影响成熟B细胞的分化。这些结果证明Usp39对于B细胞的发育是必须的。而对B细胞免疫反应的评价结果表明，Usp39的缺失并不影响B细胞活化，也未显著改变T细胞依赖以及非依赖的免疫反应水平。以上结果表明Usp39在B细胞发育而非免疫反应中有重要的生物学功能。

本研究进一步解析了Usp39在B细胞早期发育中的功能及机制通路。在B细胞发育更早期敲除Usp39导致小鼠外周淋巴器官中的B细胞几近消失，骨髓B细胞的发育被完全阻滞在pre-pro B细胞阶段。结合早期敲除小鼠pro B至pre B细胞发育受阻的实验结果，证明Usp39的主要功能是促进B细胞通过早期发育检查点。全转录组测序分析、B细胞受体分析以及诸多实验结果表明，Usp39这一功能并不由细胞增殖或细胞凋亡介导，也不通过转录调控实现，而是通过调控重链的重排、进而影响重链与轻链的合成得以实现。此外，通过引入重排完成的IgM以重塑B细胞受体可以使敲除小鼠骨髓B细胞重新顺利通过发育检查点，并使其外周B细胞恢复至野生型水平。以上结果表明，Usp39通过调控B细胞受体重排来促进B细胞通过发育检查点。

本研究最后明确了Usp39调控B细胞受体重排的分子机制。对B细胞中RNA剪接水平的评价结果表明，Usp39的缺失并没有从整体以及重链局部影响RNA的剪接。但突变其负责介导tri-snRNP组装的活性位点，Usp39丧失了促进B细胞发育的功能。以上结果证明Usp39调控B细胞发育的功能依赖于剪接子的组装，而非RNA的剪接。对B细胞受体重排中顺式与反式调控元件的评价结果表明，Usp39并非通过影响反式调控元件Rag重组酶的功能，而是调控了重链位点3’端顺式调控元件的相互作用，进而影响V区基因接近重排中心、参与重排反应。重链的染色质可接近性评价结果也表明，3’端染色质开放区域集中在重排中心相关位点。以上结果证明Usp39通过调控重链基因位点染色质的相互作用调节重链重排。此外，本文进一步探索了Usp39在B细胞淋巴瘤发生中的作用。在淋巴瘤模型小鼠中敲除Usp39可以显著延长小鼠的无瘤存活周期，并明显减少肿瘤样B细胞的产生。这些结果证明Usp39作为B细胞淋巴瘤治疗的靶点具有潜在的可能性。

综上所述，本研究通过免疫学、分子生物学以及多组学手段，首次揭示了Usp39在B细胞早期发育调控中的生物学功能，阐明了Usp39通过调控B细胞受体重排促进B细胞通过发育检查点的机制通路，并进一步证明了Usp39通过剪接子依赖的方式调控染色质相互作用、进而影响重链重排。基于该理论，本研究还证实了Usp39可以作为B细胞淋巴瘤治疗的潜在靶点。以上研究结果为剪接子调控B细胞发育提供了一种新的理论基础，也为通过剪接子相关途径治疗B细胞肿瘤提供了理论可能。

[1] KUROSAKI T, SHINOHARA H, BABA Y. B cell signaling and fate decision[J]. Annu Rev Immunol, 2010, 28: 21-55.
[2] NUTT S L, KEE B L. The transcriptional regulation of B cell lineage commitment[J]. Immunity, 2007, 26(6): 715-25.
[3] ZHENG Z, ZHANG L, CUI X L, et al. Control of early B cell development by the RNA N(6)-methyladenosine methylation[J]. Cell Rep, 2020, 31(13): 107819.
[4] PESTAL K, FUNK C C, SNYDER J M, et al. Isoforms of RNA-editing enzyme ADAR1 independently control nucleic acid sensor MDA5-driven autoimmunity and multi-organ development[J]. Immunity, 2015, 43(5): 933-44.
[5] WILKINSON M E, CHARENTON C, NAGAI K. RNA splicing by the spliceosome[J]. Annu Rev Biochem, 2020, 89: 359-88.
[6] HERZEL L, OTTOZ D S M, ALPERT T, et al. Splicing and transcription touch base: co-transcriptional spliceosome assembly and function[J]. Nat Rev Mol Cell Biol, 2017, 18(10): 637-50.
[7] BONNET A, GROSSO A R, ELKAOUTARI A, et al. Introns protect eukaryotic genomes from transcription-associated genetic instability[J]. Mol Cell, 2017, 67(4): 608-21 e6.
[8] BLACK K L, NAQVI A S, ASNANI M, et al. Aberrant splicing in B-cell acute lymphoblastic leukemia[J]. Nucleic Acids Res, 2018, 46(21): 11357-69.
[9] SCOTTI M M, SWANSON M S. RNA mis-splicing in disease[J]. Nat Rev Genet, 2016, 17(1): 19-32.
[10] MAKAROVA O V, MAKAROV E M, LUHRMANN R. The 65 and 110 kDa SR-related proteins of the U4/U6.U5 tri-snRNP are essential for the assembly of mature spliceosomes[J]. EMBO J, 2001, 20(10): 2553-63.
[11] HADJIVASSILIOU H, ROSENBERG O S, GUTHRIE C. The crystal structure of S. cerevisiae Sad1, a catalytically inactive deubiquitinase that is broadly required for pre-mRNA splicing[J]. RNA, 2014, 20(5): 656-69.
[12] BERTRAM K, AGAFONOV D E, DYBKOV O, et al. Cryo-EM structure of a pre-catalytic human spliceosome primed for activation[J]. Cell, 2017, 170(4): 701-13 e11.
[13] WU J, CHEN Y, GENG G, et al. USP39 regulates DNA damage response and chemo-radiation resistance by deubiquitinating and stabilizing CHK2[J]. Cancer Lett, 2019, 449: 114-24.
[14] LI X, YUAN J, SONG C, et al. Deubiquitinase USP39 and E3 ligase TRIM26 balance the level of ZEB1 ubiquitination and thereby determine the progression of hepatocellular carcinoma[J]. Cell Death Differ, 2021, 28(8): 2315-32.
[15] HUANG Y, PAN X W, LI L, et al. Overexpression of USP39 predicts poor prognosis and promotes tumorigenesis of prostate cancer via promoting EGFR mRNA maturation and transcription elongation[J]. Oncotarget, 2016, 7(16): 22016-30.
[16] MANDEL E M, GROSSCHEDL R. Transcription control of early B cell differentiation[J]. Curr Opin Immunol, 2010, 22(2): 161-7.
[17] ULLRICH S, GUIGO R. Dynamic changes in intron retention are tightly associated with regulation of splicing factors and proliferative activity during B-cell development[J]. Nucleic Acids Res, 2020, 48(3): 1327-40.
[18] TURNER M, DIAZ-MUNOZ M D. RNA-binding proteins control gene expression and cell fate in the immune system[J]. Nat Immunol, 2018, 19(2): 120-9.
[19] HARDY R R, KINCADE P W, DORSHKIND K. The protean nature of cells in the B lymphocyte lineage[J]. Immunity, 2007, 26(6): 703-14.
[20] PROUDHON C, HAO B, RAVIRAM R, et al. Long-range regulation of V(D)J recombination[J]. Adv Immunol, 2015, 128: 123-82.
[21] NUSSENZWEIG M C, ALT F W. Antibody diversity: one enzyme to rule them all[J]. Nat Med, 2004, 10(12): 1304-5.
[22] LIU C, ZHANG Y, LIU C C, et al. Structural insights into the evolution of the RAG recombinase[J]. Nat Rev Immunol, 2021.
[23] TANAKA S, BABA Y. B cell receptor signaling[J]. Adv Exp Med Biol, 2020, 1254: 23-36.
[24] RUMFELT L L, ZHOU Y, ROWLEY B M, et al. Lineage specification and plasticity in CD19-early B cell precursors[J]. J Exp Med, 2006, 203(3): 675-87.
[25] MELCHERS F. Checkpoints that control B cell development[J]. J Clin Invest, 2015, 125(6): 2203-10.
[26] CAROTTA S, DAKIC A, D'AMICO A, et al. The transcription factor PU.1 controls dendritic cell development and Flt3 cytokine receptor expression in a dose-dependent manner[J]. Immunity, 2010, 32(5): 628-41.
[27] ANDERSON K L, NELSON S L, PERKIN H B, et al. PU.1 is a lineage-specific regulator of tyrosine phosphatase CD45[J]. J Biol Chem, 2001, 276(10): 7637-42.
[28] KWON K, HUTTER C, SUN Q, et al. Instructive role of the transcription factor E2A in early B lymphopoiesis and germinal center B cell development[J]. Immunity, 2008, 28(6): 751-62.
[29] LIN Y C, JHUNJHUNWALA S, BENNER C, et al. A global network of transcription factors, involving E2A, EBF1 and Foxo1, that orchestrates B cell fate[J]. Nat Immunol, 2010, 11(7): 635-43.
[30] TREIBER T, MANDEL E M, POTT S, et al. Early B cell factor 1 regulates B cell gene networks by activation, repression, and transcription- independent poising of chromatin[J]. Immunity, 2010, 32(5): 714-25.
[31] BANERJEE A, NORTHRUP D, BOUKARABILA H, et al. Transcriptional repression of Gata3 is essential for early B cell commitment[J]. Immunity, 2013, 38(5): 930-42.
[32] FUXA M, BUSSLINGER M. Reporter gene insertions reveal a strictly B lymphoid-specific expression pattern of Pax5 in support of its B cell identity function[J]. J Immunol, 2007, 178(12): 8222-8.
[33] VERMA-GAUR J, TORKAMANI A, SCHAFFER L, et al. Noncoding transcription within the Igh distal V(H) region at PAIR elements affects the 3D structure of the Igh locus in pro-B cells[J]. Proc Natl Acad Sci U S A, 2012, 109(42): 17004-9.
[34] MCMANUS S, EBERT A, SALVAGIOTTO G, et al. The transcription factor Pax5 regulates its target genes by recruiting chromatin-modifying proteins in committed B cells[J]. EMBO J, 2011, 30(12): 2388-404.
[35] HOLMES M L, PRIDANS C, NUTT S L. The regulation of the B-cell gene expression programme by Pax5[J]. Immunol Cell Biol, 2008, 86(1): 47-53.
[36] XU L S, SOKALSKI K M, HOTKE K, et al. Regulation of B cell linker protein transcription by PU.1 and Spi-B in murine B cell acute lymphoblastic leukemia[J]. J Immunol, 2012, 189(7): 3347-54.
[37] FERREIROS-VIDAL I, CARROLL T, TAYLOR B, et al. Genome-wide identification of Ikaros targets elucidates its contribution to mouse B-cell lineage specification and pre-B-cell differentiation[J]. Blood, 2013, 121(10): 1769-82.
[38] MA S, PATHAK S, TRINH L, et al. Interferon regulatory factors 4 and 8 induce the expression of Ikaros and Aiolos to down-regulate pre-B-cell receptor and promote cell-cycle withdrawal in pre-B-cell development[J]. Blood, 2008, 111(3): 1396-403.
[39] SWAMINATHAN S, HUANG C, GENG H, et al. BACH2 mediates negative selection and p53-dependent tumor suppression at the pre-B cell receptor checkpoint[J]. Nat Med, 2013, 19(8): 1014-22.
[40] GREIG K T, DE GRAAF C A, MURPHY J M, et al. Critical roles for c-Myb in lymphoid priming and early B-cell development[J]. Blood, 2010, 115(14): 2796-805.
[41] NIEBUHR B, KRIEBITZSCH N, FISCHER M, et al. Runx1 is essential at two stages of early murine B-cell development[J]. Blood, 2013, 122(3): 413-23.
[42] OCHIAI K, MAIENSCHEIN-CLINE M, MANDAL M, et al. A self-reinforcing regulatory network triggered by limiting IL-7 activates pre-BCR signaling and differentiation[J]. Nat Immunol, 2012, 13(3): 300-7.
[43] VU L T, TSUKAHARA T. C-to-U editing and site-directed RNA editing for the correction of genetic mutations[J]. Biosci Trends, 2017, 11(3): 243-53.
[44] YABLONOVITCH A L, DENG P, JACOBSON D, et al. The evolution and adaptation of A-to-I RNA editing[J]. PLoS Genet, 2017, 13(11): e1007064.
[45] EISENBERG E, LEVANON E Y. A-to-I RNA editing - immune protector and transcriptome diversifier[J]. Nat Rev Genet, 2018, 19(8): 473-90.
[46] MARCU-MALINA V, GOLDBERG S, VAX E, et al. ADAR1 is vital for B cell lineage development in the mouse bone marrow[J]. Oncotarget, 2016, 7(34): 54370-9.
[47] ZHAO B S, ROUNDTREE I A, HE C. Post-transcriptional gene regulation by mRNA modifications[J]. Nat Rev Mol Cell Biol, 2017, 18(1): 31-42.
[48] DENG L J, DENG W Q, FAN S R, et al. m6A modification: recent advances, anticancer targeted drug discovery and beyond[J]. Mol Cancer, 2022, 21(1): 52.
[49] GRENOV A C, MOSS L, EDELHEIT S, et al. The germinal center reaction depends on RNA methylation and divergent functions of specific methyl readers[J]. J Exp Med, 2021, 218(10).
[50] NAIR L, ZHANG W, LAFFLEUR B, et al. Mechanism of noncoding RNA-associated N(6)-methyladenosine recognition by an RNA processing complex during IgH DNA recombination[J]. Mol Cell, 2021, 81(19): 3949-64 e7.
[51] CHENG Y, FU Y, WANG Y, et al. The m6A methyltransferase METTL3 is functionally implicated in DLBCL development by regulating m6A modification in PEDF[J]. Front Genet, 2020, 11: 955.
[52] HAN H, FAN G, SONG S, et al. piRNA-30473 contributes to tumorigenesis and poor prognosis by regulating m6A RNA methylation in DLBCL[J]. Blood, 2021, 137(12): 1603-14.
[53] SCHOENBERG D R, MAQUAT L E. Regulation of cytoplasmic mRNA decay[J]. Nat Rev Genet, 2012, 13(4): 246-59.
[54] LABNO A, TOMECKI R, DZIEMBOWSKI A. Cytoplasmic RNA decay pathways-Enzymes and mechanisms[J]. Biochim Biophys Acta, 2016, 1863(12): 3125-47.
[55] NASIF S, CONTU L, MUHLEMANN O. Beyond quality control: The role of nonsense-mediated mRNA decay (NMD) in regulating gene expression[J]. Semin Cell Dev Biol, 2018, 75: 78-87.
[56] INOUE T, MORITA M, HIJIKATA A, et al. CNOT3 contributes to early B cell development by controlling Igh rearrangement and p53 mRNA stability[J]. J Exp Med, 2015, 212(9): 1465-79.
[57] YANG C Y, RAMAMOORTHY S, BOLLER S, et al. Interaction of CCR4-NOT with EBF1 regulates gene-specific transcription and mRNA stability in B lymphopoiesis[J]. Genes Dev, 2016, 30(20): 2310-24.
[58] GALLOWAY A, SAVELIEV A, LUKASIAK S, et al. RNA-binding proteins ZFP36L1 and ZFP36L2 promote cell quiescence[J]. Science, 2016, 352(6284): 453-9.
[59] ZHANG L, REYNOLDS T L, SHAN X, et al. Coupling of V(D)J recombination to the cell cycle suppresses genomic instability and lymphoid tumorigenesis[J]. Immunity, 2011, 34(2): 163-74.
[60] NEWMAN R, AHLFORS H, SAVELIEV A, et al. Maintenance of the marginal-zone B cell compartment specifically requires the RNA-binding protein ZFP36L1[J]. Nat Immunol, 2017, 18(6): 683-93.
[61] HUANG H, ZHANG G, RUAN G X, et al. Mettl14-mediated m6A modification is essential for germinal center B cell response[J]. J Immunol, 2022, 208(8): 1924-36.
[62] SHI Y. Mechanistic insights into precursor messenger RNA splicing by the spliceosome[J]. Nat Rev Mol Cell Biol, 2017, 18(11): 655-70.
[63] PAN Q, SHAI O, LEE L J, et al. Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing[J]. Nat Genet, 2008, 40(12): 1413-5.
[64] WANG E T, SANDBERG R, LUO S, et al. Alternative isoform regulation in human tissue transcriptomes[J]. Nature, 2008, 456(7221): 470-6.
[65] KIM M S, PINTO S M, GETNET D, et al. A draft map of the human proteome[J]. Nature, 2014, 509(7502): 575-81.
[66] SHI Y. The Spliceosome: A protein-directed metalloribozyme[J]. J Mol Biol, 2017, 429(17): 2640-53.
[67] MATERA A G, WANG Z. A day in the life of the spliceosome[J]. Nat Rev Mol Cell Biol, 2014, 15(2): 108-21.
[68] PLASCHKA C, LIN P C, CHARENTON C, et al. Prespliceosome structure provides insights into spliceosome assembly and regulation[J]. Nature, 2018, 559(7714): 419-22.
[69] ZHANG X, YAN C, HANG J, et al. An atomic structure of the human spliceosome[J]. Cell, 2017, 169(5): 918-29 e14.
[70] RAUHUT R, FABRIZIO P, DYBKOV O, et al. Molecular architecture of the Saccharomyces cerevisiae activated spliceosome[J]. Science, 2016, 353(6306): 1399-405.
[71] OHRT T, ODENWALDER P, DANNENBERG J, et al. Molecular dissection of step 2 catalysis of yeast pre-mRNA splicing investigated in a purified system[J]. RNA, 2013, 19(7): 902-15.
[72] SCHWER B. A conformational rearrangement in the spliceosome sets the stage for Prp22-dependent mRNA release[J]. Mol Cell, 2008, 30(6): 743-54.
[73] ULE J, BLENCOWE B J. Alternative splicing regulatory networks: functions, mechanisms, and evolution[J]. Mol Cell, 2019, 76(2): 329-45.
[74] SHEPARD P J, HERTEL K J. The SR protein family[J]. Genome Biol, 2009, 10(10): 242.
[75] GEUENS T, BOUHY D, TIMMERMAN V. The hnRNP family: insights into their role in health and disease[J]. Hum Genet, 2016, 135(8): 851-67.
[76] SHARMA S, MARIS C, ALLAIN F H, et al. U1 snRNA directly interacts with polypyrimidine tract-binding protein during splicing repression[J]. Mol Cell, 2011, 41(5): 579-88.
[77] BONNAL S, MARTINEZ C, FORCH P, et al. RBM5/Luca-15/H37 regulates Fas alternative splice site pairing after exon definition[J]. Mol Cell, 2008, 32(1): 81-95.
[78] CHEN C D, KOBAYASHI R, HELFMAN D M. Binding of hnRNP H to an exonic splicing silencer is involved in the regulation of alternative splicing of the rat beta-tropomyosin gene[J]. Genes Dev, 1999, 13(5): 593-606.
[79] BURATTI E, STUANI C, DE PRATO G, et al. SR protein-mediated inhibition of CFTR exon 9 inclusion: molecular characterization of the intronic splicing silencer[J]. Nucleic Acids Res, 2007, 35(13): 4359-68.
[80] CHIOU N T, SHANKARLING G, LYNCH K W. hnRNP L and hnRNP A1 induce extended U1 snRNA interactions with an exon to repress spliceosome assembly[J]. Mol Cell, 2013, 49(5): 972-82.
[81] KOSTER T, STAIGER D. RNA-binding protein immunoprecipitation and high-throughput sequencing[J]. Methods Mol Biol, 2021, 2200: 453-61.
[82] STERNBURG E L, KARGINOV F V. Global approaches in studying RNA-binding protein interaction networks[J]. Trends Biochem Sci, 2020, 45(7): 593-603.
[83] KWAK H, LIS J T. Control of transcriptional elongation[J]. Annu Rev Genet, 2013, 47: 483-508.
[84] DAS R, DUFU K, ROMNEY B, et al. Functional coupling of RNAP II transcription to spliceosome assembly[J]. Genes Dev, 2006, 20(9): 1100-9.
[85] WARF M B, BERGLUND J A. Role of RNA structure in regulating pre-mRNA splicing[J]. Trends Biochem Sci, 2010, 35(3): 169-78.
[86] SPILUTTINI B, GU B, BELAGAL P, et al. Splicing-independent recruitment of U1 snRNP to a transcription unit in living cells[J]. J Cell Sci, 2010, 123(Pt 12): 2085-93.
[87] BRODY Y, NEUFELD N, BIEBERSTEIN N, et al. The in vivo kinetics of RNA polymerase II elongation during co-transcriptional splicing[J]. PLoS Biol, 2011, 9(1): e1000573.
[88] ALMADA A E, WU X, KRIZ A J, et al. Promoter directionality is controlled by U1 snRNP and polyadenylation signals[J]. Nature, 2013, 499(7458): 360-3.
[89] CHIU A C, SUZUKI H I, WU X, et al. Transcriptional pause sites delineate stable nucleosome-associated premature polyadenylation suppressed by U1 snRNP[J]. Mol Cell, 2018, 69(4): 648-63 e7.
[90] REED R, HURT E. A conserved mRNA export machinery coupled to pre-mRNA splicing[J]. Cell, 2002, 108(4): 523-31.
[91] MOORE M J, PROUDFOOT N J. Pre-mRNA processing reaches back to transcription and ahead to translation[J]. Cell, 2009, 136(4): 688-700.
[92] BATSCHE E, YANIV M, MUCHARDT C. The human SWI/SNF subunit Brm is a regulator of alternative splicing[J]. Nat Struct Mol Biol, 2006, 13(1): 22-9.
[93] CAVELLAN E, ASP P, PERCIPALLE P, et al. The WSTF-SNF2h chromatin remodeling complex interacts with several nuclear proteins in transcription[J]. J Biol Chem, 2006, 281(24): 16264-71.
[94] KFIR N, LEV-MAOR G, GLAICH O, et al. SF3B1 association with chromatin determines splicing outcomes[J]. Cell Rep, 2015, 11(4): 618-29.
[95] LUCO R F, PAN Q, TOMINAGA K, et al. Regulation of alternative splicing by histone modifications[J]. Science, 2010, 327(5968): 996-1000.
[96] ZHOU H L, LUO G, WISE J A, et al. Regulation of alternative splicing by local histone modifications: potential roles for RNA-guided mechanisms[J]. Nucleic Acids Res, 2014, 42(2): 701-13.
[97] GELFMAN S, COHEN N, YEARIM A, et al. DNA-methylation effect on cotranscriptional splicing is dependent on GC architecture of the exon-intron structure[J]. Genome Res, 2013, 23(5): 789-99.
[98] ZHOU H L, HINMAN M N, BARRON V A, et al. Hu proteins regulate alternative splicing by inducing localized histone hyperacetylation in an RNA-dependent manner[J]. Proc Natl Acad Sci U S A, 2011, 108(36): E627-35.
[99] KIM S, KIM H, FONG N, et al. Pre-mRNA splicing is a determinant of histone H3K36 methylation[J]. Proc Natl Acad Sci U S A, 2011, 108(33): 13564-9.
[100] DE ALMEIDA S F, GROSSO A R, KOCH F, et al. Splicing enhances recruitment of methyltransferase HYPB/Setd2 and methylation of histone H3 Lys36[J]. Nat Struct Mol Biol, 2011, 18(9): 977-83.
[101] YUAN W, XIE J, LONG C, et al. Heterogeneous nuclear ribonucleoprotein L Is a subunit of human KMT3a/Set2 complex required for H3 Lys-36 trimethylation activity in vivo[J]. J Biol Chem, 2009, 284(23): 15701-7.
[102] CONVERTINI P, SHEN M, POTTER P M, et al. Sudemycin E influences alternative splicing and changes chromatin modifications[J]. Nucleic Acids Res, 2014, 42(8): 4947-61.
[103] YABAS M, ELLIOTT H, HOYNE G F. The Role of alternative splicing in the control of immune homeostasis and cellular differentiation[J]. Int J Mol Sci, 2015, 17(1).
[104] CHABOT B, SHKRETA L. Defective control of pre-messenger RNA splicing in human disease[J]. J Cell Biol, 2016, 212(1): 13-27.
[105] LYGEROU Z, CHRISTOPHIDES G, SERAPHIN B. A novel genetic screen for snRNP assembly factors in yeast identifies a conserved protein, Sad1p, also required for pre-mRNA splicing[J]. Mol Cell Biol, 1999, 19(3): 2008-20.
[106] PLASCHKA C, LIN P C, NAGAI K. Structure of a pre-catalytic spliceosome[J]. Nature, 2017, 546(7660): 617-21.
[107] HUANG Y H, CHUNG C S, KAO D I, et al. Sad1 counteracts Brr2-mediated dissociation of U4/U6.U5 in tri-snRNP homeostasis[J]. Mol Cell Biol, 2014, 34(2): 210-20.
[108] SOWA M E, BENNETT E J, GYGI S P, et al. Defining the human deubiquitinating enzyme interaction landscape[J]. Cell, 2009, 138(2): 389-403.
[109] JERONIMO C, FORGET D, BOUCHARD A, et al. Systematic analysis of the protein interaction network for the human transcription machinery reveals the identity of the 7SK capping enzyme[J]. Mol Cell, 2007, 27(2): 262-74.
[110] VAN LEUKEN R J, LUNA-VARGAS M P, SIXMA T K, et al. Usp39 is essential for mitotic spindle checkpoint integrity and controls mRNA-levels of aurora B[J]. Cell Cycle, 2008, 7(17): 2710-9.
[111] ZHAO Y, ZHANG B, LEI Y, et al. Knockdown of USP39 induces cell cycle arrest and apoptosis in melanoma[J]. Tumour Biol, 2016, 37(10): 13167-76.
[112] YAN C, YUAN J, XU J, et al. Ubiquitin-specific peptidase 39 regulates the process of proliferation and migration of human ovarian cancer via p53/p21 pathway and EMT[J]. Med Oncol, 2019, 36(11): 95.
[113] DING K, JI J, ZHANG X, et al. RNA splicing factor USP39 promotes glioma progression by inducing TAZ mRNA maturation[J]. Oncogene, 2019, 38(37): 6414-28.
[114] YUAN J, ZHANG G, LI X, et al. Knocking down USP39 inhibits the growth and metastasis of non-small-cell lung cancer cells through activating the p53 pathway[J]. Int J Mol Sci, 2020, 21(23).
[115] DONG X, LIU Z, ZHANG E, et al. USP39 promotes tumorigenesis by stabilizing and deubiquitinating SP1 protein in hepatocellular carcinoma[J]. Cell Signal, 2021, 85: 110068.
[116] YUAN J, LI X, ZHANG G, et al. USP39 mediates p21-dependent proliferation and neoplasia of colon cancer cells by regulating the p53/p21/CDC2/cyclin B1 axis[J]. Mol Carcinog, 2021, 60(4): 265-78.
[117] YUAN J, LI X, ZHANG Y, et al. USP39 attenuates the antitumor activity of cisplatin on colon cancer cells dependent on p53[J]. Cell Biol Toxicol, 2021.
[118] WANG S, WANG Z, LI J, et al. Splicing factor USP39 promotes ovarian cancer malignancy through maintaining efficient splicing of oncogenic HMGA2[J]. Cell Death Dis, 2021, 12(4): 294.
[119] PAN X W, XU D, CHEN W J, et al. USP39 promotes malignant proliferation and angiogenesis of renal cell carcinoma by inhibiting VEGF-A165b alternative splicing via regulating SRSF1 and SRPK1[J]. Cancer Cell Int, 2021, 21(1): 486.
[120] FRAILE J M, MANCHADO E, LUJAMBIO A, et al. USP39 deubiquitinase is essential for KRAS oncogene-driven cancer[J]. J Biol Chem, 2017, 292(10): 4164-75.
[121] PENG Y, GUO J, SUN T, et al. USP39 serves as a deubiquitinase to stabilize STAT1 and sustains type I IFN-induced antiviral immunity[J]. J Immunol, 2020, 205(11): 3167-78.
[122] KIM J J, LEE S Y, HWANG Y, et al. USP39 promotes non-homologous end-joining repair by poly(ADP-ribose)-induced liquid demixing[J]. Nucleic Acids Res, 2021, 49(19): 11083-102.
[123] RIOS Y, MELMED S, LIN S, et al. Zebrafish usp39 mutation leads to rb1 mRNA splicing defect and pituitary lineage expansion[J]. PLoS Genet, 2011, 7(1): e1001271.
[124] ERGUN A, DORAN G, COSTELLO J C, et al. Differential splicing across immune system lineages[J]. Proc Natl Acad Sci U S A, 2013, 110(35): 14324-9.
[125] TOUNG J M, MORLEY M, LI M, et al. RNA-sequence analysis of human B-cells[J]. Genome Res, 2011, 21(6): 991-8.
[126] HERMISTON M L, XU Z, WEISS A. CD45: a critical regulator of signaling thresholds in immune cells[J]. Annu Rev Immunol, 2003, 21: 107-37.
[127] HATHCOCK K S, HIRANO H, MURAKAMI S, et al. CD45 expression by B cells. Expression of different CD45 isoforms by subpopulations of activated B cells[J]. J Immunol, 1992, 149(7): 2286-94.
[128] MCNEILL L, CASSADY R L, SARKARDEI S, et al. CD45 isoforms in T cell signalling and development[J]. Immunol Lett, 2004, 92(1-2): 125-34.
[129] BEVERLEY P C, DASER A, MICHIE C A, et al. Functional subsets of T cells defined by isoforms of CD45[J]. Biochem Soc Trans, 1992, 20(1): 184-7.
[130] PREUSSNER M, SCHREINER S, HUNG L H, et al. HnRNP L and L-like cooperate in multiple-exon regulation of CD45 alternative splicing[J]. Nucleic Acids Res, 2012, 40(12): 5666-78.
[131] PIOLI P D, DEBNATH I, WEIS J J, et al. Zfp318 regulates IgD expression by abrogating transcription termination within the Ighm/Ighd locus[J]. J Immunol, 2014, 193(5): 2546-53.
[132] ENDERS A, SHORT A, MIOSGE L A, et al. Zinc-finger protein ZFP318 is essential for expression of IgD, the alternatively spliced Igh product made by mature B lymphocytes[J]. Proc Natl Acad Sci U S A, 2014, 111(12): 4513-8.
[133] XU Y, ZHOU H, POST G, et al. Rad52 mediates class-switch DNA recombination to IgD[J]. Nat Commun, 2022, 13(1): 980.
[134] MA J, GUNDERSON S I, PHILLIPS C. Non-snRNP U1A levels decrease during mammalian B-cell differentiation and release the IgM secretory poly(A) site from repression[J]. RNA, 2006, 12(1): 122-32.
[135] ANAND S, BATISTA F D, TKACH T, et al. Multiple transcripts of the murine immunoglobulin epsilon membrane locus are generated by alternative splicing and differential usage of two polyadenylation sites[J]. Mol Immunol, 1997, 34(2): 175-83.
[136] BENSON M J, AIJO T, CHANG X, et al. Heterogeneous nuclear ribonucleoprotein L-like (hnRNPLL) and elongation factor, RNA polymerase II, 2 (ELL2) are regulators of mRNA processing in plasma cells[J]. Proc Natl Acad Sci U S A, 2012, 109(40): 16252-7.
[137] BEGUM N A, HAQUE F, STANLIE A, et al. Phf5a regulates DNA repair in class switch recombination via p400 and histone H2A variant deposition[J]. EMBO J, 2021, 40(12): e106393.
[138] KANEHIRO Y, TODO K, NEGISHI M, et al. Activation-induced cytidine deaminase (AID)-dependent somatic hypermutation requires a splice isoform of the serine/arginine-rich (SR) protein SRSF1[J]. Proc Natl Acad Sci U S A, 2012, 109(4): 1216-21.
[139] SINGH A K, TAMRAKAR A, JAISWAL A, et al. SRSF1-3, a splicing and somatic hypermutation regulator, controls transcription of IgV genes via chromatin regulators SATB2, UBN1 and histone variant H3.3[J]. Mol Immunol, 2020, 119: 69-82.
[140] KUMAR SINGH A, TAMRAKAR A, JAISWAL A, et al. Splicing regulator SRSF1-3 that controls somatic hypermutation of IgV genes interacts with topoisomerase 1 and AID[J]. Mol Immunol, 2019, 116: 63-72.
[141] NOWAK U, MATTHEWS A J, ZHENG S, et al. The splicing regulator PTBP2 interacts with the cytidine deaminase AID and promotes binding of AID to switch-region DNA[J]. Nat Immunol, 2011, 12(2): 160-6.
[142] JIN W, NIU Z, XU D, et al. RBM5 promotes exon 4 skipping of AID pre-mRNA by competing with the binding of U2AF65 to the polypyrimidine tract[J]. FEBS Lett, 2012, 586(21): 3852-7.
[143] MONZON-CASANOVA E, MATHESON L S, TABBADA K, et al. Polypyrimidine tract-binding proteins are essential for B cell development[J]. Elife, 2020, 9.
[144] MONZON-CASANOVA E, BATES K J, SMITH C W J, et al. Essential requirement for polypyrimidine tract binding proteins 1 and 3 in the maturation and maintenance of mature B cells in mice[J]. Eur J Immunol, 2021, 51(9): 2266-73.
[145] MONZON-CASANOVA E, SCREEN M, DIAZ-MUNOZ M D, et al. The RNA-binding protein PTBP1 is necessary for B cell selection in germinal centers[J]. Nat Immunol, 2018, 19(3): 267-78.
[146] DIAZ-MUNOZ M D, BELL S E, FAIRFAX K, et al. The RNA-binding protein HuR is essential for the B cell antibody response[J]. Nat Immunol, 2015, 16(4): 415-25.
[147] CHANG X, LI B, RAO A. RNA-binding protein hnRNPLL regulates mRNA splicing and stability during B-cell to plasma-cell differentiation[J]. Proc Natl Acad Sci U S A, 2015, 112(15): E1888-97.
[148] DIAZ-MUNOZ M D, MONZON-CASANOVA E, TURNER M. Characterization of the B cell transcriptome bound by RNA-binding proteins with iCLIP[J]. Methods Mol Biol, 2017, 1623: 159-79.
[149] MALCOVATI L, PAPAEMMANUIL E, BOWEN D T, et al. Clinical significance of SF3B1 mutations in myelodysplastic syndromes and myelodysplastic/myeloproliferative neoplasms[J]. Blood, 2011, 118(24): 6239-46.
[150] GRAUBERT T A, SHEN D, DING L, et al. Recurrent mutations in the U2AF1 splicing factor in myelodysplastic syndromes[J]. Nat Genet, 2011, 44(1): 53-7.
[151] YOSHIDA K, SANADA M, SHIRAISHI Y, et al. Frequent pathway mutations of splicing machinery in myelodysplasia[J]. Nature, 2011, 478(7367): 64-9.
[152] FANG J, BOLANOS L C, CHOI K, et al. Ubiquitination of hnRNPA1 by TRAF6 links chronic innate immune signaling with myelodysplasia[J]. Nat Immunol, 2017, 18(2): 236-45.
[153] CHEN L, TOVAR-CORONA J M, URRUTIA A O. Increased levels of noisy splicing in cancers, but not for oncogene-derived transcripts[J]. Hum Mol Genet, 2011, 20(22): 4422-9.
[154] DVINGE H, BRADLEY R K. Widespread intron retention diversifies most cancer transcriptomes[J]. Genome Med, 2015, 7(1): 45.
[155] PUENTE X S, BEA S, VALDES-MAS R, et al. Non-coding recurrent mutations in chronic lymphocytic leukaemia[J]. Nature, 2015, 526(7574): 519-24.
[156] TAYLOR J, LEE S C. Mutations in spliceosome genes and therapeutic opportunities in myeloid malignancies[J]. Genes Chromosomes Cancer, 2019, 58(12): 889-902.
[157] LEE S C, ABDEL-WAHAB O. Therapeutic targeting of splicing in cancer[J]. Nat Med, 2016, 22(9): 976-86.
[158] WANG L, LAWRENCE M S, WAN Y, et al. SF3B1 and other novel cancer genes in chronic lymphocytic leukemia[J]. N Engl J Med, 2011, 365(26): 2497-506.
[159] ROSSI D, BRUSCAGGIN A, SPINA V, et al. Mutations of the SF3B1 splicing factor in chronic lymphocytic leukemia: association with progression and fludarabine-refractoriness[J]. Blood, 2011, 118(26): 6904-8.
[160] SOTILLO E, BARRETT D M, BLACK K L, et al. Convergence of acquired mutations and alternative splicing of CD19 enables resistance to CART-19 immunotherapy[J]. Cancer Discov, 2015, 5(12): 1282-95.
[161] WU X, DARCE J R, CHANG S K, et al. Alternative splicing regulates activation-induced cytidine deaminase (AID): implications for suppression of AID mutagenic activity in normal and malignant B cells[J]. Blood, 2008, 112(12): 4675-82.
[162] GALLARDO M, MALANEY P, AITKEN M J L, et al. Uncovering the role of RNA-binding protein hnRNP K in B-cell lymphomas[J]. J Natl Cancer Inst, 2020, 112(1): 95-106.
[163] KOZYREV S V, ABELSON A K, WOJCIK J, et al. Functional variants in the B-cell gene BANK1 are associated with systemic lupus erythematosus[J]. Nat Genet, 2008, 40(2): 211-6.
[164] MI H, MURUGANUJAN A, HUANG X, et al. Protocol update for large-scale genome and gene function analysis with the PANTHER classification system (v.14.0)[J]. Nat Protoc, 2019, 14(3): 703-21.
[165] BOLOTIN D A, POSLAVSKY S, DAVYDOV A N, et al. Antigen receptor repertoire profiling from RNA-seq data[J]. Nat Biotechnol, 2017, 35(10): 908-11.
[166] SHUGAY M, BAGAEV D V, TURCHANINOVA M A, et al. VDJtools: unifying post-analysis of T cell receptor repertoires[J]. PLoS Comput Biol, 2015, 11(11): e1004503.
[167] SUBRAHMANYAM R, DU H, IVANOVA I, et al. Localized epigenetic changes induced by DH recombination restricts recombinase to DJH junctions[J]. Nat Immunol, 2012, 13(12): 1205-12.
[168] GUO C, YOON H S, FRANKLIN A, et al. CTCF-binding elements mediate control of V(D)J recombination[J]. Nature, 2011, 477(7365): 424-30.
[169] YEO G, BURGE C B. Maximum entropy modeling of short sequence motifs with applications to RNA splicing signals[J]. J Comput Biol, 2004, 11(2-3): 377-94.
[170] SHEN S, PARK J W, LU Z X, et al. rMATS: robust and flexible detection of differential alternative splicing from replicate RNA-Seq data[J]. Proc Natl Acad Sci U S A, 2014, 111(51): E5593-601.
[171] BOLLAND D J, WOOD A L, AFSHAR R, et al. Antisense intergenic transcription precedes Igh D-to-J recombination and is controlled by the intronic enhancer Emu[J]. Mol Cell Biol, 2007, 27(15): 5523-33.
[172] TRANCOSO I, BONNET M, GARDNER R, et al. A Novel quantitative fluorescent reporter assay for RAG targets and RAG activity[J]. Front Immunol, 2013, 4: 110.
[173] HOLMES R, ZUNIGA-PFLUCKER J C. The OP9-DL1 system: generation of T-lymphocytes from embryonic or hematopoietic stem cells in vitro[J]. Cold Spring Harb Protoc, 2009, 2009(2): pdb prot5156.
[174] RAO S S, HUNTLEY M H, DURAND N C, et al. A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping[J]. Cell, 2014, 159(7): 1665-80.
[175] NIU L, SHEN W, HUANG Y, et al. Amplification-free library preparation with SAFE Hi-C uses ligation products for deep sequencing to improve traditional Hi-C analysis[J]. Commun Biol, 2019, 2: 267.
[176] SEBINA I, PEPPER M. Humoral immune responses to infection: common mechanisms and unique strategies to combat pathogen immune evasion tactics[J]. Curr Opin Immunol, 2018, 51: 46-54.
[177] HOBEIKA E, THIEMANN S, STORCH B, et al. Testing gene function early in the B cell lineage in mb1-cre mice[J]. Proc Natl Acad Sci U S A, 2006, 103(37): 13789-94.
[178] WANG L, CHEN T, LI X, et al. USP39 promotes ovarian cancer malignant phenotypes and carboplatin chemoresistance[J]. Int J Oncol, 2019, 55(1): 277-88.
[179] KITAMURA T, KOSHINO Y, SHIBATA F, et al. Retrovirus-mediated gene transfer and expression cloning: powerful tools in functional genomics[J]. Exp Hematol, 2003, 31(11): 1007-14.
[180] MASON D Y, JONES M, GOODNOW C C. Development and follicular localization of tolerant B lymphocytes in lysozyme/anti-lysozyme IgM/IgD transgenic mice[J]. Int Immunol, 1992, 4(2): 163-75.
[181] TENG G, SCHATZ D G. Regulation and evolution of the RAG recombinase[J]. Adv Immunol, 2015, 128: 1-39.
[182] KOTAKE Y, SAGANE K, OWA T, et al. Splicing factor SF3b as a target of the antitumor natural product pladienolide[J]. Nat Chem Biol, 2007, 3(9): 570-5.
[183] EBERT A, HILL L, BUSSLINGER M. Spatial Regulation of V-(D)J Recombination at Antigen Receptor Loci[J]. Adv Immunol, 2015, 128: 93-121.
[184] MARSHALL A J, WU G E, PAIGE G J. Frequency of VH81x usage during B cell development: initial decline in usage is independent of Ig heavy chain cell surface expression[J]. J Immunol, 1996, 156(6): 2077-84.
[185] BURGER J A, WIESTNER A. Targeting B cell receptor signalling in cancer: preclinical and clinical advances[J]. Nat Rev Cancer, 2018, 18(3): 148-67.
[186] RUSTAD E H, MISUND K, BERNARD E, et al. Stability and uniqueness of clonal immunoglobulin CDR3 sequences for MRD tracking in multiple myeloma[J]. Am J Hematol, 2019, 94(12): 1364-73.
[187] AGATHANGELIDIS A, CHATZIDIMITRIOU A, GEMENETZI K, et al. Higher-order connections between stereotyped subsets: implications for improved patient classification in CLL[J]. Blood, 2021, 137(10): 1365-76.
[188] ADAMS J M, HARRIS A W, PINKERT C A, et al. The c-myc oncogene driven by immunoglobulin enhancers induces lymphoid malignancy in transgenic mice[J]. Nature, 1985, 318(6046): 533-8.
[189] KOH C M, BEZZI M, LOW D H, et al. MYC regulates the core pre-mRNA splicing machinery as an essential step in lymphomagenesis[J]. Nature, 2015, 523(7558): 96-100.