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The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen
We have previously demonstrated [Rihs, H.‐P. and Peters, R. (1989) EMBO J., 8, 1479–1484] that the nuclear transport of recombinant proteins in which short fragments of the SV40 T‐antigen are fused to the amino terminus of Escherichia coli beta‐galactosidase is dependent on both the nuclear localiza...
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Published in: | The EMBO journal 1991-03, Vol.10 (3), p.633-639 |
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description | We have previously demonstrated [Rihs, H.‐P. and Peters, R. (1989) EMBO J., 8, 1479–1484] that the nuclear transport of recombinant proteins in which short fragments of the SV40 T‐antigen are fused to the amino terminus of Escherichia coli beta‐galactosidase is dependent on both the nuclear localization sequence (NLS, T‐antigen residues 126–132) and a phosphorylation‐site‐containing sequence (T‐antigen residues 111–125). While the NLS determines the specificity, the rate of transport is controlled by the phosphorylation‐site‐containing sequence. The present study furthers this observation and examines the role of the various phosphorylation sites. Purified, fluorescently labeled recombinant proteins were injected into the cytoplasm of Vero or hepatoma (HTC) cells and the kinetics of nuclear transport measured by laser microfluorimetry. By replacing serine and threonine residues known to be phosphorylated in vivo, we identified the casein kinase II (CK‐II) site S111/S112 to be the determining factor in the enhancement of the transport. Either of the residues 111 or 112 was sufficient to elicit the maximum transport enhancement. The other phosphorylation sites (S120, S123, T124) had no influence on the transport rate. Examination of the literature suggested that many proteins harboring a nuclear localization sequence also contain putative CK‐II sites at a distance of approximately 10–30 amino acid residues from the NLS. CK‐II has been previously implicated in the transmission of growth signals to the nucleus. Our results suggest that CK‐II may exert this role by controlling the rate of nuclear protein transport. |
doi_str_mv | 10.1002/j.1460-2075.1991.tb07991.x |
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P. ; Jans, D. A. ; Fan, H. ; Peters, R.</creator><creatorcontrib>Rihs, H. P. ; Jans, D. A. ; Fan, H. ; Peters, R.</creatorcontrib><description>We have previously demonstrated [Rihs, H.‐P. and Peters, R. (1989) EMBO J., 8, 1479–1484] that the nuclear transport of recombinant proteins in which short fragments of the SV40 T‐antigen are fused to the amino terminus of Escherichia coli beta‐galactosidase is dependent on both the nuclear localization sequence (NLS, T‐antigen residues 126–132) and a phosphorylation‐site‐containing sequence (T‐antigen residues 111–125). While the NLS determines the specificity, the rate of transport is controlled by the phosphorylation‐site‐containing sequence. The present study furthers this observation and examines the role of the various phosphorylation sites. Purified, fluorescently labeled recombinant proteins were injected into the cytoplasm of Vero or hepatoma (HTC) cells and the kinetics of nuclear transport measured by laser microfluorimetry. By replacing serine and threonine residues known to be phosphorylated in vivo, we identified the casein kinase II (CK‐II) site S111/S112 to be the determining factor in the enhancement of the transport. Either of the residues 111 or 112 was sufficient to elicit the maximum transport enhancement. The other phosphorylation sites (S120, S123, T124) had no influence on the transport rate. Examination of the literature suggested that many proteins harboring a nuclear localization sequence also contain putative CK‐II sites at a distance of approximately 10–30 amino acid residues from the NLS. CK‐II has been previously implicated in the transmission of growth signals to the nucleus. Our results suggest that CK‐II may exert this role by controlling the rate of nuclear protein transport.</description><identifier>ISSN: 0261-4189</identifier><identifier>EISSN: 1460-2075</identifier><identifier>DOI: 10.1002/j.1460-2075.1991.tb07991.x</identifier><identifier>PMID: 1848177</identifier><language>eng</language><publisher>England</publisher><subject>Amino Acid Sequence ; Animals ; Antigens, Polyomavirus Transforming - genetics ; beta-Galactosidase - genetics ; casein kinase II ; Casein Kinases ; Cell Nucleus - metabolism ; Cell Transformation, Viral ; Cytoplasm - metabolism ; Humans ; Kinetics ; Molecular Sequence Data ; Phosphorylation ; Plasmids ; Protein Kinases - genetics ; proteins ; Recombinant Fusion Proteins - analysis ; Restriction Mapping ; Sequence Homology, Nucleic Acid ; Simian virus 40 - enzymology ; Simian virus 40 - genetics ; Simian virus 40 - immunology ; Vero Cells</subject><ispartof>The EMBO journal, 1991-03, Vol.10 (3), p.633-639</ispartof><rights>1991 European Molecular Biology Organization</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c5091-645e3a43802900ff08733526730a511c58d51e95dcd265eccbd424954170c5de3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC452694/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC452694/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,723,776,780,881,27903,27904,53769,53771</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/1848177$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Rihs, H. P.</creatorcontrib><creatorcontrib>Jans, D. A.</creatorcontrib><creatorcontrib>Fan, H.</creatorcontrib><creatorcontrib>Peters, R.</creatorcontrib><title>The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen</title><title>The EMBO journal</title><addtitle>EMBO J</addtitle><description>We have previously demonstrated [Rihs, H.‐P. and Peters, R. (1989) EMBO J., 8, 1479–1484] that the nuclear transport of recombinant proteins in which short fragments of the SV40 T‐antigen are fused to the amino terminus of Escherichia coli beta‐galactosidase is dependent on both the nuclear localization sequence (NLS, T‐antigen residues 126–132) and a phosphorylation‐site‐containing sequence (T‐antigen residues 111–125). While the NLS determines the specificity, the rate of transport is controlled by the phosphorylation‐site‐containing sequence. The present study furthers this observation and examines the role of the various phosphorylation sites. Purified, fluorescently labeled recombinant proteins were injected into the cytoplasm of Vero or hepatoma (HTC) cells and the kinetics of nuclear transport measured by laser microfluorimetry. By replacing serine and threonine residues known to be phosphorylated in vivo, we identified the casein kinase II (CK‐II) site S111/S112 to be the determining factor in the enhancement of the transport. Either of the residues 111 or 112 was sufficient to elicit the maximum transport enhancement. The other phosphorylation sites (S120, S123, T124) had no influence on the transport rate. Examination of the literature suggested that many proteins harboring a nuclear localization sequence also contain putative CK‐II sites at a distance of approximately 10–30 amino acid residues from the NLS. CK‐II has been previously implicated in the transmission of growth signals to the nucleus. Our results suggest that CK‐II may exert this role by controlling the rate of nuclear protein transport.</description><subject>Amino Acid Sequence</subject><subject>Animals</subject><subject>Antigens, Polyomavirus Transforming - genetics</subject><subject>beta-Galactosidase - genetics</subject><subject>casein kinase II</subject><subject>Casein Kinases</subject><subject>Cell Nucleus - metabolism</subject><subject>Cell Transformation, Viral</subject><subject>Cytoplasm - metabolism</subject><subject>Humans</subject><subject>Kinetics</subject><subject>Molecular Sequence Data</subject><subject>Phosphorylation</subject><subject>Plasmids</subject><subject>Protein Kinases - genetics</subject><subject>proteins</subject><subject>Recombinant Fusion Proteins - analysis</subject><subject>Restriction Mapping</subject><subject>Sequence Homology, Nucleic Acid</subject><subject>Simian virus 40 - enzymology</subject><subject>Simian virus 40 - genetics</subject><subject>Simian virus 40 - immunology</subject><subject>Vero Cells</subject><issn>0261-4189</issn><issn>1460-2075</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1991</creationdate><recordtype>article</recordtype><recordid>eNqVUc2O0zAQthBoKQuPgGRx4JYyTuz8IHFYVgsULeJA4Wq5zqTrktjBdmHLiUfgDXg3ngSnLQscOc3I38_M-CPkEYM5A8ifbOaMl5DlUIk5axo2jyuopnp9i8xuoNtkBnnJMs7q5i65F8IGAERdsRNywmpes6qakR_LK6ReRaSuo3are1Se6l10Y6_CYDQdvYtoLI1e2TA6H6kJtMWIfjAWW7ra0ZgstAoT66OxqaGLBQ0meXa9sulpvaf8du-dVr35qqJxlgb8tEWr9-Mn0rsPHOjy57fvykazRnuf3OlUH_DBsZ6S9y8uluevssu3LxfnZ5eZFtCwrOQCC8WLGvIGoOugropC5GVVgBKMaVG3gmEjWt3mpUCtVy3PeSM4q0CLFotT8uzgO25XA7YabTq4l6M3g_I76ZSR_yLWXMm1-yx5mtLwpH981HuXLgpRDiZo7NMHoNsGyUQDXLAiEZ8eiNq7EDx2NzMYyClduZFThHKKUE7pymO68jqJH_695R_pIc6Enx3wL6bH3X84y4s3z1_v--IXqs-56A</recordid><startdate>199103</startdate><enddate>199103</enddate><creator>Rihs, H. P.</creator><creator>Jans, D. A.</creator><creator>Fan, H.</creator><creator>Peters, R.</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>8FD</scope><scope>FR3</scope><scope>M7Z</scope><scope>P64</scope><scope>5PM</scope></search><sort><creationdate>199103</creationdate><title>The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen</title><author>Rihs, H. P. ; Jans, D. A. ; Fan, H. ; Peters, R.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c5091-645e3a43802900ff08733526730a511c58d51e95dcd265eccbd424954170c5de3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1991</creationdate><topic>Amino Acid Sequence</topic><topic>Animals</topic><topic>Antigens, Polyomavirus Transforming - genetics</topic><topic>beta-Galactosidase - genetics</topic><topic>casein kinase II</topic><topic>Casein Kinases</topic><topic>Cell Nucleus - metabolism</topic><topic>Cell Transformation, Viral</topic><topic>Cytoplasm - metabolism</topic><topic>Humans</topic><topic>Kinetics</topic><topic>Molecular Sequence Data</topic><topic>Phosphorylation</topic><topic>Plasmids</topic><topic>Protein Kinases - genetics</topic><topic>proteins</topic><topic>Recombinant Fusion Proteins - analysis</topic><topic>Restriction Mapping</topic><topic>Sequence Homology, Nucleic Acid</topic><topic>Simian virus 40 - enzymology</topic><topic>Simian virus 40 - genetics</topic><topic>Simian virus 40 - immunology</topic><topic>Vero Cells</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Rihs, H. P.</creatorcontrib><creatorcontrib>Jans, D. A.</creatorcontrib><creatorcontrib>Fan, H.</creatorcontrib><creatorcontrib>Peters, R.</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biochemistry Abstracts 1</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>The EMBO journal</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Rihs, H. P.</au><au>Jans, D. A.</au><au>Fan, H.</au><au>Peters, R.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen</atitle><jtitle>The EMBO journal</jtitle><addtitle>EMBO J</addtitle><date>1991-03</date><risdate>1991</risdate><volume>10</volume><issue>3</issue><spage>633</spage><epage>639</epage><pages>633-639</pages><issn>0261-4189</issn><eissn>1460-2075</eissn><abstract>We have previously demonstrated [Rihs, H.‐P. and Peters, R. (1989) EMBO J., 8, 1479–1484] that the nuclear transport of recombinant proteins in which short fragments of the SV40 T‐antigen are fused to the amino terminus of Escherichia coli beta‐galactosidase is dependent on both the nuclear localization sequence (NLS, T‐antigen residues 126–132) and a phosphorylation‐site‐containing sequence (T‐antigen residues 111–125). While the NLS determines the specificity, the rate of transport is controlled by the phosphorylation‐site‐containing sequence. The present study furthers this observation and examines the role of the various phosphorylation sites. Purified, fluorescently labeled recombinant proteins were injected into the cytoplasm of Vero or hepatoma (HTC) cells and the kinetics of nuclear transport measured by laser microfluorimetry. By replacing serine and threonine residues known to be phosphorylated in vivo, we identified the casein kinase II (CK‐II) site S111/S112 to be the determining factor in the enhancement of the transport. Either of the residues 111 or 112 was sufficient to elicit the maximum transport enhancement. The other phosphorylation sites (S120, S123, T124) had no influence on the transport rate. Examination of the literature suggested that many proteins harboring a nuclear localization sequence also contain putative CK‐II sites at a distance of approximately 10–30 amino acid residues from the NLS. CK‐II has been previously implicated in the transmission of growth signals to the nucleus. Our results suggest that CK‐II may exert this role by controlling the rate of nuclear protein transport.</abstract><cop>England</cop><pmid>1848177</pmid><doi>10.1002/j.1460-2075.1991.tb07991.x</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Amino Acid Sequence Animals Antigens, Polyomavirus Transforming - genetics beta-Galactosidase - genetics casein kinase II Casein Kinases Cell Nucleus - metabolism Cell Transformation, Viral Cytoplasm - metabolism Humans Kinetics Molecular Sequence Data Phosphorylation Plasmids Protein Kinases - genetics proteins Recombinant Fusion Proteins - analysis Restriction Mapping Sequence Homology, Nucleic Acid Simian virus 40 - enzymology Simian virus 40 - genetics Simian virus 40 - immunology Vero Cells |
title | The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen |
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