Pericentrin, a conserved centrosomal component, provides the structural scaffold to anchor numerous centrosomal proteins, and thus plays an essential role in the organization and function of the centrosome and the mit...Pericentrin, a conserved centrosomal component, provides the structural scaffold to anchor numerous centrosomal proteins, and thus plays an essential role in the organization and function of the centrosome and the mitotic spindle. Although pericentrin was shown to localize in the cytoplasm and reported to be sensitive to leptomycin B （LMB）, a specific inhibitor of Crml, the regions within pericentrin that serve as signals for transporting in and out of the nucleus have not yet been identified. In this study, we identified five novel nuclear export signals （NESs） in pericentrin with diverse export activities. All of the five NESs could bind to Crml in a LMB-sensitive way when mediating the nuclear export of pericentrin. We also demonstrated that the region of amino acids 8-42 in pericentrin contains a tripartite nuclear localization signal （NLS） consisting of three clusters of basic amino acids. The NLS of pericentrin binds to importin β directly or via the adaptor importin α to form the import complex, which could be disrupted by RanQ69L, a dominant-negative Ran GTPase possessing high affinity for importin β. Furthermore, we found that mutation of the NESs in full-length pericentrin results in both nuclear and cytoplasmic localization, and mutation of the NLS abolishes the nuclear import of pericentrin. On the basis of these results, we suggest that the NESs and NLS of pericentrin are essential for its subcellular localization and nucleocytoplasmic trafficking during the cell cycle.展开更多
Transcription factor TFIIA is controlled by complex regulatory networks including proteolysis by the protease Taspase 1, though the full impact of cleavage remains elusive. Here, we demonstrate that in contrast to the...Transcription factor TFIIA is controlled by complex regulatory networks including proteolysis by the protease Taspase 1, though the full impact of cleavage remains elusive. Here, we demonstrate that in contrast to the general assumption, de novo produced TFIIA is rapidly confined to the cytoplasm via an evolutionary conserved nuclear export signal （NES, amino acids ^21VINDVRDIFL^30）, interacting with the nuclear export receptor Exportin-1/chromosomal region maintenance 1 （Crml）. Chemical export inhibition or genetic inactivation of the NES not only promotes TFIIA＇s nuclear localization but also affects its transcrip- tional activity. Notably, Taspase 1 processing promotes TFIIA＇s nuclear accumulation by NES masking, and modulates its tran- scriptional activity. Moreover, TFIIA complex formation with the TATA box binding protein （TBP） is cooperatively enhanced by inhibition of proteolysis and nuclear export, leading to an increase of the ceil cycle inhibitor p16INK, which is counteracted by pre- vention of TBP binding. We here identified a novel mechanism how proteolysis and nuclear transport cooperatively fine-tune tran- scriptional programs.展开更多
文摘Pericentrin, a conserved centrosomal component, provides the structural scaffold to anchor numerous centrosomal proteins, and thus plays an essential role in the organization and function of the centrosome and the mitotic spindle. Although pericentrin was shown to localize in the cytoplasm and reported to be sensitive to leptomycin B （LMB）, a specific inhibitor of Crml, the regions within pericentrin that serve as signals for transporting in and out of the nucleus have not yet been identified. In this study, we identified five novel nuclear export signals （NESs） in pericentrin with diverse export activities. All of the five NESs could bind to Crml in a LMB-sensitive way when mediating the nuclear export of pericentrin. We also demonstrated that the region of amino acids 8-42 in pericentrin contains a tripartite nuclear localization signal （NLS） consisting of three clusters of basic amino acids. The NLS of pericentrin binds to importin β directly or via the adaptor importin α to form the import complex, which could be disrupted by RanQ69L, a dominant-negative Ran GTPase possessing high affinity for importin β. Furthermore, we found that mutation of the NESs in full-length pericentrin results in both nuclear and cytoplasmic localization, and mutation of the NLS abolishes the nuclear import of pericentrin. On the basis of these results, we suggest that the NESs and NLS of pericentrin are essential for its subcellular localization and nucleocytoplasmic trafficking during the cell cycle.
文摘Transcription factor TFIIA is controlled by complex regulatory networks including proteolysis by the protease Taspase 1, though the full impact of cleavage remains elusive. Here, we demonstrate that in contrast to the general assumption, de novo produced TFIIA is rapidly confined to the cytoplasm via an evolutionary conserved nuclear export signal （NES, amino acids ^21VINDVRDIFL^30）, interacting with the nuclear export receptor Exportin-1/chromosomal region maintenance 1 （Crml）. Chemical export inhibition or genetic inactivation of the NES not only promotes TFIIA＇s nuclear localization but also affects its transcrip- tional activity. Notably, Taspase 1 processing promotes TFIIA＇s nuclear accumulation by NES masking, and modulates its tran- scriptional activity. Moreover, TFIIA complex formation with the TATA box binding protein （TBP） is cooperatively enhanced by inhibition of proteolysis and nuclear export, leading to an increase of the ceil cycle inhibitor p16INK, which is counteracted by pre- vention of TBP binding. We here identified a novel mechanism how proteolysis and nuclear transport cooperatively fine-tune tran- scriptional programs.
