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研究生:高維娟
研究生(外文):Wei-Chang Kao
論文名稱:剪接調節分子RBM4與其結合蛋白之功能探討
論文名稱(外文):Functional study of RBM4-interacting proteins
指導教授:譚婉玉
指導教授(外文):Woan-Yuh Tarn
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:解剖暨細胞生物學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
中文關鍵詞:剪接調節分子
外文關鍵詞:RBM4
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在真核細胞中遺傳訊息DNA轉錄出來的precursor mRNA,需要經過剪接 (splicing) 才能將基因適當的表達出來,而選擇性剪接 (alternative splicing) 的方式大量造就基因表達的複雜度及多樣性。許多的研究結果已知,SR蛋白家族不僅調控 precursor mRNA splicing 也參與 alternative splicing 的機制,而這些SR蛋白除了有 RNA recongination motif (RRM) 之外,都有一段arginine/serine重複的胺基酸序列 (RS domain),而 RS domain 的功能在於和其它蛋白分子之間產生交互作用。
我們以前的研究發現一個蛋白分子 RNA binding motif protein 4 (RBM4),它和 SR 蛋白經由相同的路徑從細胞質進入到細胞核,且和某些 SR 蛋白相同,皆游移在細胞核與質當中。其結構上,RBM4 的 N端有兩個RRM,C端有三處 alanine-rich 的序列, 稱為 CAD 。 RRM 和 CAD 在 alternative splicing 上都扮演著決定性的角色。
我們預測 RBM4 的 CAD 會和其它蛋白分子產生交互作用。我們經由酵母菌雙雜合篩選法得到可能和 RBM4 結合的蛋白。其中一個蛋白為 hnRNP M,hnRNP M 屬於 hnRNP 家族,此家族成員參與核醣核酸合成及代謝過程的許多步驟,例如剪接、運輸和降解。經由 in vitro binding assay 得知 RBM4 和hnRNP M 兩者之間有專一性的結合,從免疫螢光染色的結果也看到 RBM4 和hnRNP M,同時位於細胞核內。另外藉由 yeast two-hybrid assay進ㄧ步發現hnRNP M 是藉由本身的 methionine/arginie 區域 (M/R-rich motif) 與RBM4 結合。最後,由in vivo splicing assay結果得知,hnRNP M 對於不同的pre-mRNA,會有不同剪接的影響。而我們更發現 hnRNP M 的 M/R-rich motif 能替代 CAD 在 alternative splicing 機制中,修飾剪接點及選擇exon 的功能。
Alternative splicing of pre-mRNA is a regulatory mechanism of gene expression in eukaryotic cells. This pathway can generate different mRNA isoforms, and thereby increases the coding capacity of genes. SR (Ser/Arg dipeptide-rich) proteins and hnRNP proteins play important roles in constitutive and/or regulated splicing. RBM4 is a non-SR splicing regulatory protein, and shares a same import pathway with SR proteins to the nucleus. RBM4 possesses two RNA recognition motifs (RRM) and an alanine-rich C-terminal domain (CAD). Both RRM and CAD are critical for alternative splicing. We postulate that the CAD of RBM4 acts as a module for protein-protein interaction. In this study, we employed a yeast two-hybrid system to search for proteins that interact with the CAD. The screens yielded five positive clones. The candidate protein hnRNP M belongs to the subfamily of hnRNPs that participate in various steps of RNA metabolism, including pre-mRNA splicing. The interaction of RBM4 with hnRNP M was confirmed by in vitro binding assays. We demonstrated that hnRNP M binds to RBM4 through its methionine/argenine-rich region (M/R-rich motif). Transient expression of hnRNP M in HEK293 cells can differentially affect alternative splice site or exon selection. More interestingly, the M/R-rich motif can replace the CAD of RBM4 in modulation in alternative splicing.
參考文獻:
Auboeuf D, Honig A, Berget SM, O''Malley BW. (2002). Coordinate Regulation of transcription and Splicing by Steroid Receptor Coregulators. Science 298, 416-419.
Allemand E, Dokudovskaya S, Bordonne R, Tazi J. (2002). A Conserved Drosophila Transportin-Serine/Arginine-rich (SR) Protein Permits Nuclear Import of Drosophila SR Protein Splicing Factors and Their Antagonist Repressor Splicing Factor 1. Mol. Cell. Biol. 13, 2436-2447.
Ahn MY, Huang G, Bae SC, Wee HJ, Kim WY, Ito Y. (1998). Negative regulation of granulocytic differentiation in the myeloid precursor cell line 32Dcl3 by ear-2, a mammalian homolog of Drosophila seven-up, and a chimeric leukemogenic gene, AML1/ETO(MTG8). Proc Natl Acad Sci U S A. 95, 1812.
Black DL. (1998). Splicing in the inner ear: a familiar tune, but what are the instruments? Neuron. 20,165-168.
Blencowe BJ. (2000). Exonic splicing enhancers: mechanism of action, diversity and role in human genetic diseases. Trends Biochem Sci. 25, 106-110.
Barnett SF, Friedman DL, LeStourgeon WM. (1989). The C proteins of HeLa 40S nuclear ribonucleoprotein particles exist as anisotropic tetramers of (C1)3 C2. Mol. Cell. Biol. 9, 492-498.
Braddock DT, Baber JL, Levens D, Clore GM. (2002). Molecular basis of sequence-specific single-stranded DNA recognition by KH domains: solution structure of a complex between hnRNP K KH3 and single-stranded DNA. EMBO J. 21, 3476-3485.
Burd CG, Dreyfuss G. (1994). RNA binding specificity of hnRNP A1: significance of hnRNP A1 high-affinity binding sites in pre-mRNA splicing. EMBO J. 13, 1197-1204.
Barnard DC, Patton JG. (2000). Identification and Characterization of a Novel Serine-Arginine-Rich Splicing Regulatory Protein. Mol. Cell. Biol. 20, 3049-3057.
Chu PH, Ruiz-Lozano P, Zhou Q, Cai C, Chen J. (2000). Expression patterns of FHL/SLIM family members suggest important functional roles in skeletal muscle and cardiovascular system. Mech. Dev. 95, 259-265.
Caceres JF, Krainer AR. (1993). Functional analysis of pre-mRNA splicing factor SF2/ASF structural domains. EMBO 12, 4715-4726.
Carter KC, Taneja KL, Lawrence JB. (1991). Discrete nuclear domains of poly(A) RNA and their relationship to the functional organization of the nucleus. J Cell Biol. 115, 1191-1202.
Cindy L Will and Reinhard Luhrmann (1997). Protein functions in pre-mRNA splicing. Curr. Opin. Cell Biol. 9, 320-328.
Cobianchi F, Karpel RL, Williams KR, Notario V, Wilson SH. (1988). Mammalian heterogeneous nuclear ribonucleoprotein complex protein A1. Large-scale overproduction in Escherichia coli and cooperative binding to single-stranded nucleic acids. J. Biol. Chem. 263, 1063-1071.
Chiodi I, Biggiogera M, Denegri M, Corioni M, Weighardt F, Cobianchi F, Riva S, Biamonti G. (2000). Structure and dynamics of hnRNP-labelled nuclear bodies induced by stress treatments. J. Cell Sci. 113, 4043-4053.
Cronshaw JM, Krutchinsky AN, Zhang W, Chait BT, Matunis MJ. (2002). Proteomic analysis of the mammalian nuclear pore complex. J. Cell Biol. 158, 915-927.
Chan KK, Tsui SK, Lee SM, Luk SC, Liew CC, Fung KP, Waye MM, Lee CY. (1998). Molecular cloning and characterization of FHL2, a novel LIM domain protein preferentially expressed in human heart. Gene 210, 345-350.
Cartegni L, Maconi M, Morandi E, Cobianchi F, Riva S, Biamonti G. (1996). hnRNP A1 Selectively Interacts Through its Gly-rich Domain with Different RNA-binding Proteins. J. Mol. Biol. 259, 337-348.
Calvo O, Manley JL. (2001). Evolutionarily Conserved Interaction between CstF-64 and PC4 Links Transcription, Polyadenylation, and Termination. Mol Cell. 7, 1013-1023.
Duncan PI, Stojdl DF, Marius RM, Scheit KH, Bell JC. (1998). The Clk2 and Clk3 Dual-Specificity Protein Kinases Regulate the Intranuclear Distribution of SR Proteins and Influence Pre-mRNA Splicing. Exp. Cell Res. 241, 300-308.
Dye BT, Patton JG. (2001). An RNA Recognition Motif (RRM) Is Required for the Localization of PTB-Associated Splicing Factor (PSF) to Subnuclear Speckles. Exp. Cell Res. 263, 131-144.
Graveley BR. (2000). Sorting out the complexity of SR protein functions. RNA 6, 1197-1211.
Ge H, Manley JL. (1990). A protein factor, ASF, controls cell-specific alternative splicing of SV40 early pre-mRNA in vitro. Cell 62, 25-34.
Gattoni R, Mahe D, Mahl P, Fischer N, Mattei MG, Stevenin J, Fuchs JP. (1996). The human hnRNP-M proteins: structure and relation with early heat shock-induced splicing arrest and chromosome mapping. Nucl. Acids. Res. 24, 2535-2542.
Gattoni R, Schmitt P, Stevenin J. (1988). In vitro splicing of adenovirus E1A transcripts: characterization of novel reactions and of multiple branch points abnormally far from the 3'' splice site. Nucl. Acids. Res. 16, 2389-2409.
Hastings ML, Krainer AR. (2001). Pre-mRNA splicing in the new millennium. Curr. Opin. Cell Biol. 13, 302-309.
Jurica MS, Moore MJ. (2003). Pre-mRNA Splicing: Awash in a Sea of Proteins. Mol Cell. 12, 5-14.
Krecic AM, Swanson MS. (1999). hnRNP complexes: composition, structure, and function. Curr. Opin. Cell Biol. 11, 363-371.
Kafasla P, Patrinou-Georgoula M, Lewis JD, Guialis A. (2002). Association of the 72/74-kDa proteins, members of the heterogeneous nuclear ribonucleoprotein M group, with the pre-mRNA at early stages of spliceosome assembly. Biochem. J. 363, 793-799.
Konig H, Ponta H, Herrlich P. (1998). Coupling of signal transduction to alternative pre-mRNA splicing by a composite splice regulator. EMBO J. 17, 2904-2913.
Kiledjian M, Dreyfuss G. (1992). Primary structure and binding activity of the hnRNP U protein: binding RNA through RGG box. EMBO J. 11, 2655-2664
Kim JH, Hahm B, Kim YK, Choi M, Jang SK. (2000). Protein-protein interaction among hnRNPs shuttling between nucleus and cytoplasm. J. Mol. Biol. 298, 395-405.
Kim YJ, Zuo P, Manley JL, Baker BS. (1992). The Drosophila RNA-binding protein RBP1 is localized to transcriptionally active sites of chromosomes and shows a functional similarity to human splicing factor ASF/SF2. Genes Dev. 6, 2569-2579.
Li H and Bingham PM (1991). Arginine/serine-rich domains of the su(wa) and tra RNA processing regulators target proteins to a subnuclear compartment implicated in splicing. Cell 67, 335-342.
Liu Z, Luyten I, Bottomley MJ, Messias AC, Houngninou-Molango S, Sprangers R, Zanier K, Kramer A, Sattler M. (2001). Structural basis for recognition of the intron branch site RNA by splicing factor 1. Science 294, 1098-1102.
Lai MC, Kuo HW, Chang WC, Tarn WY. (2003). A novel splicing regulator shares a nuclear import pathway with SR proteins. EMBO J. 22, 1359-1369.
Li J, Hawkins IC, Harvey CD, Jennings JL, Link AJ, Patton JG. (2003). Regulation of alternative Splicing by SRrp86 and Its Interacting Proteins. Mol. Cell. Biol. 23, 7437-7447.
Lau CK, Diem MD, Dreyfuss G, Van Duyne GD. (2003). Structure of the Y14-Magoh Core of the Exon Junction Complex. Curr. Biol. 13, 933-941.
Lindberg RA, Quinn AM, Hunter T. (1992). Dual-specificity protein kinases: Will any hydroxyl do. Trends Biochem. Sci. 17, 114-119.
Lee SM, Tsui SK, Chan KK, Garcia-Barcelo M, Waye MM, Fung KP, Liew CC, Lee CY. (1998). Chromosomal mapping, tissue distribution and cDNA sequence of Four-and-a-half LIM domain protein 1 (FHL1). Gene 216, 163-170.
Melchior F, Schergaut M, Pichler A. (2003). SUMO: ligases, isopeptidases and nuclear pores. Trends Biochem Sci. 28, 612-618.
Menegay H, Moeslein F, Landreth G. (1999). The Dual Specificity Protein Kinase CLK3 Is Abundantly Expressed in Mature Mouse Spermatozoa. Exp. Cell Res. 253, 463-473.
Muller JM, Isele U, Metzger E, Rempel A, Moser M, Pscherer A, Breyer T, Holubarsch C, Buettner R, Schule R. (2000). FHL2, a novel tissue-specific coactivator of the androgen receptor. EMBO J. 19, 359-369.
Muller JM, Metzger E, Greschik H, Bosserhoff AK, Mercep L, Buettner R, Schule R. (2002). The transcriptional coactivator FHL2 transmits Rho signals from the cell membrane into the nucleus. EMBO J. 21, 736-748.
Miau LH, Chang CJ, Shen BJ, Tsai WH, Lee SH. (1998). Identification of Heterogeneous Nuclear Ribonucleoprotein K (hnRNP K) as a Repressor of C/EBP-mediated Gene Activation. J Biol Chem. 273, 10784-10791.
Michelsen JW, Schmeichel KL, Beckerle MC, Winge DR. (1993). The LIM motif defines a specific zinc-binding protein domain. Proc. Natl. Acad. Sci. USA 90, 4404-4408.
Morgan MJ, Madgwick AJ. (1996). SLIM defines a novel family of LIM-proteins expressed in skeletal muscle. Biochem. Biophys. Res. Commun. 225, 632-638.
Morgan MJ, Madgwick AJ, Charleston B, Pell JM, Loughna PT. (1995). The developmental regulation of a novel muscle LIM-protein. Biochem. Biophys. Res. Commun. 212, 840-846.
Myers MP, Murphy MB, Landreth G. (1994). The dual-specificity CLK kinase induces neuronal differentiation of PC12 cells. Mol. Cell. Biol. 14, 6954-6961.
Mata M de la, Alonso CR, Kadener S, Fededa JP, Blaustein M, Pelisch F, Cramer P, Bentley D, Kornblihtt AR. (2003). A Slow RNA Polymerase II Affects Alternative Splicing In Vivo. Mol Cell. 12, 525-532.
Nayler O, Stamm S, Ullrich A. (1997). Characterization and comparison of four serine- and arginine-rich (SR) protein kinases. Biochem. J. 326, 693-700.
Nakielny S and Dreyfuss G (1997). Nuclear export of proteins and RNAs. Curr. Opin. Cell Biol. 9, 420-429.
Oh YL, Hahm B, Kim YK, Lee HK, Lee JW, Song O, Tsukiyama-Kohara K, Kohara M, Nomoto A, Jang SK. (2003). Surface expression of heterogeneous nuclear RNA binding protein M4 on Kupffer cell relates to its function as a carcinoembryonic antigen receptor. Exp. Cell Res. 291, 228-241.
Perez I, McAfee JG, Patton JG. (1997). Multiple RRMs contribute to RNA binding specificity and affinity for polypyrimidine tract binding protein. Biochemistry 36, 11881-11890.
Perez Canadillas JM, Varani G. (2003). Recognition of GU-rich polyadenylation regulatory elements by human CstF-64 protein. EMBO J. 22, 2821-2830.
Pinol-Roma S, Choi YD, Matunis MJ, Dreyfuss G. (1988). Immunopurification of heterogeneous nuclear ribonucleoprotein particles reveals an assortment of RNA-binding proteins. Genes Dev. 2, 215-227.
Shi H, Xu RM. (2003). Crystal structure of the Drosophila Mago nashi-Y14 complex. Genes Dev. 17, 971-976.
Smith CW, Valcarcel J. (2000). Alternative pre-mRNA splicing: the logic of combinatorial control. Trends Biochem Sci. 25, 381-388.
Sakashita E, Tatsumi S, Werner D, Endo H, Mayeda A. (2004). Human RNPS1 and Its Associated Factors: a Versatile Alternative Pre-mRNA Splicing Regulator In Vivo. Mol. Cell. Biol. 24, 1174-1187.
Sleeman JE, Lamond AI. (1999). Nuclear organization of pre-mRNA splicing factors. Curr. Opin. Cell Biol. 11, 372-377.
Siomi H, Matunis MJ, Michael WM, Dreyfuss G. (1993). The pre-mRNA binding K protein contains a novel evolutionarily conserved motif. Nucleic Acids Res. 21, 1193-1198.
Shav-Tal Y, Cohen M, Lapter S, Dye B, Patton JG, Vandekerckhove J, Zipori D. (2004). Nuclear Relocalization of the Pre-mRNA Splicing Factor PSF during Apoptosis Involves hyperphosphorylation, Masking of Antigenic Epitopes, and Changes in Protein Interactions. Mol. Biol. Cell 12, 2340.
Takagaki Y, Manley JL. (2000). Complex Protein Interactions within the Human Polyadenylation Machinery Identify a Novel Component. Mol. Cell. Biol. 20, 1515-1525.
Takagaki Y, Seipelt RL, Peterson ML, Manley JL. (1996). The Polyadenylation Factor CstF-64 Regulates Alternative Processing of IgM Heavy Chain Pre-mRNA during B Cell Differentiation. Cell 87, 941-952.
Tolnay M, Vereshchagina LA, Tsokos GC. (1999). Heterogeneous nuclear ribonucleoprotein D0B is a sequence-specificDNA-binding protein. Biochem. J. 338, 417-425.
Tacke R, Manley JL. (1999). Determinants of SR protein specificity. Curr. Opin. Cell Biol. 11, 358-362.
Valcarcel J, Green MR. (1996). The SR protein family: pleiotropic functions in pre-mRNA splicing. Trends Biochem Sci. 21, 296-301.
Vassileva MT, Matunis MJ. (2004). SUMO Modification of Heterogeneous Nuclear Ribonucleoproteins. Mol. Cell. Biol. 24, 3623-3632
Visa N, Puvion-Dutilleul F, Harper F, Bachellerie JP, Puvion E. (1993). Intranuclear distribution of poly(A) RNA determined by electron microscope in situ hybridization. Exp. Cell Res. 208, 19-34.
Wu JY, Maniatis T. (1993). Specific interactions between proteins implicated in splice site selection and regulated alternative splicing. Cell 75, 1061-1070.
Young L, Hahm B, Kim YK, Lee HK, Lee JW, Song O, Tsukiyama-Kohara K, Kohara M, Nomoto A, Jang SK.(1998). Determination of functional domains in polypyrimidine-tract-binding protein. Biochem. J. 331, 169-175.
Zhu XG, Park KS, Kaneshige M, Bhat MK, Zhu Q, Mariash CN, McPhie P, Cheng SY. (2000). The Orphan Nuclear Receptor Ear-2 Is a Negative Coregulator for Thyroid Hormone Nuclear Receptor Function. Mol. Cell Biol. 20, 2604-2618.
Zahler AM, Neugebauer KM, Lane WS, Roth MB. (1993). Distinct functions of SR proteins in alternative pre-mRNA splicing. Science 260, 219-222.
Zu K, Sikes ML, Beyer AL. (1998). Separable roles in vivo for the two RNA binding domains of Drosophila A1-hnRNP homolog. RNA. 4, 1585-1598.
Zuo P, Manley JL. (1993). Functional domains of the human splicing factor ASF/SF2. EMBO J. 12, 4727-4737.
Zahler AM, Roth MB. (1995). Distinct Functions of SR Proteins in Recruitment ofU1 Small Nuclear Ribonucleoprotein to Alternative 5'' Splice Sites. Proc Natl Acad Sci U S A. 92, 2642-2646.
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