A BIPARTITE NUCLEAR-LOCALIZATION SIGNAL IN THE RETINOBLASTOMA GENE-PRODUCT AND ITS IMPORTANCE FOR BIOLOGICAL-ACTIVITY

被引:97
作者
ZACKSENHAUS, E
BREMNER, R
PHILLIPS, RA
GALLIE, BL
机构
[1] Div. of Immunol. and Cancer Research, Research Institute, Hospital for Sick Children, Toronto, Ont. M5G 1X8
关键词
D O I
10.1128/MCB.13.8.4588
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The retinoblastoma gene product, p110RB1, appears to regulate cell growth by modulating the activities of nuclear transcription factors. The elements that specify the transport of p110RB1 into the nucleus have not yet been explored. We now report the identification of a basic region, KRSAEGGNPPKPLKKLR, in the C terminus of p110RB1, which has sequence similarity to known bipartite nuclear localization signals (NLSs). A two-amino-acid mutation introduced into this putative NLS [to give mutant NLS(NQ)] or deletion of the entire NLS (DELTANLS) abrogated exclusive nuclear localization, yielding proteins which were distributed either equally throughout the cell or predominantly in the cytoplasm. A mutant protein [NLS(NQ)/DELTA22] containing both the mutated NLS and a deletion of exon 22, previously shown to disrupt the interaction of p110RB1 with several cellular transcription factors and oncoproteins, accumulated only in the cytoplasm. When fused to the C terminus of Escherichia coli 13-galactosidase, the RB1 NLS directed this protein to the nucleus, indicating that the motif is not only necessary but also sufficient for nuclear transport. Neither NLS(NQ) nor DELTANLS was hyperphosphorylated in vivo, but both retained their abilities to interact, in vitro, with simian virus 40 large T antigen, adenovirus E1a, and the cellular transcription factor E2F. When transfected at multiple copy number, the NLS mutant alleles displayed reduced biological activity, measured by inhibition of growth of the osteogenic sarcoma cell line Saos-2, which has no wild-type RB1. Naturally occurring mutations and deletions in exon 25 of RB1 which disrupt the NLS may lead to partial or complete inactivation of p110RB1 and may be responsible for some retinoblastoma and other tumors.
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页码:4588 / 4599
页数:12
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共 60 条
  • [1] ADDISON C, 1990, ONCOGENE, V5, P423
  • [2] SEQUENCE-SPECIFIC TRANSCRIPTIONAL ACTIVATION BY MYC AND REPRESSION BY MAX
    AMIN, C
    WAGNER, AJ
    HAY, N
    [J]. MOLECULAR AND CELLULAR BIOLOGY, 1993, 13 (01) : 383 - 390
  • [3] RETINOBLASTOMA-REPRESSION OF E2F-DEPENDENT TRANSCRIPTION DEPENDS ON THE ABILITY OF THE RETINOBLASTOMA PROTEIN TO INTERACT WITH E2F AND IS ABROGATED BY THE ADENOVIRUS E1A ONCOPROTEIN
    ARROYO, M
    RAYCHAUDHURI, P
    [J]. NUCLEIC ACIDS RESEARCH, 1992, 20 (22) : 5947 - 5954
  • [4] STRUCTURE AND EXPRESSION OF THE MURINE RETINOBLASTOMA GENE AND CHARACTERIZATION OF ITS ENCODED PROTEIN
    BERNARDS, R
    SCHACKLEFORD, GM
    GERBER, MR
    HOROWITZ, JM
    FRIEND, SH
    SCHARTL, M
    BOGENMANN, E
    RAPAPORT, JM
    MCGEE, T
    DRYJA, TP
    WEINBERG, RA
    [J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1989, 86 (17) : 6474 - 6478
  • [5] THE RETINOBLASTOMA PROTEIN IS PHOSPHORYLATED DURING SPECIFIC PHASES OF THE CELL-CYCLE
    BUCHKOVICH, K
    DUFFY, LA
    HARLOW, E
    [J]. CELL, 1989, 58 (06) : 1097 - 1105
  • [6] THE E2F TRANSCRIPTION FACTOR IS A CELLULAR TARGET FOR THE RB PROTEIN
    CHELLAPPAN, SP
    HIEBERT, S
    MUDRYJ, M
    HOROWITZ, JM
    NEVINS, JR
    [J]. CELL, 1991, 65 (06) : 1053 - 1061
  • [7] PHOSPHORYLATION OF THE RETINOBLASTOMA GENE-PRODUCT IS MODULATED DURING THE CELL-CYCLE AND CELLULAR-DIFFERENTIATION
    CHEN, PL
    SCULLY, P
    SHEW, JY
    WANG, JYJ
    LEE, WH
    [J]. CELL, 1989, 58 (06) : 1193 - 1198
  • [8] IDENTIFICATION OF THE HUMAN C-MYC PROTEIN NUCLEAR TRANSLOCATION SIGNAL
    DANG, CV
    LEE, WMF
    [J]. MOLECULAR AND CELLULAR BIOLOGY, 1988, 8 (10) : 4048 - 4054
  • [9] DAVY J, 1985, CELL, V40, P667
  • [10] SV40 LARGE TUMOR-ANTIGEN FORMS A SPECIFIC COMPLEX WITH THE PRODUCT OF THE RETINOBLASTOMA SUSCEPTIBILITY GENE
    DECAPRIO, JA
    LUDLOW, JW
    FIGGE, J
    SHEW, JY
    HUANG, CM
    LEE, WH
    MARSILIO, E
    PAUCHA, E
    LIVINGSTON, DM
    [J]. CELL, 1988, 54 (02) : 275 - 283