Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa
Modulation of the intracellular calcium concentration within mammalian spermatozoa is important in several pre-fertilization events including hyperactivated motility and the acrosome reaction. To identify calcium binding proteins (CBP) potentially regulating these processes, a 45Ca overlay technique...
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| Veröffentlicht in: | Molecular human reproduction 2001-10, Vol.7 (10), p.923-933 |
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| creator | Naaby-Hansen, Soren Wolkowicz, Michael J. Klotz, Ken Bush, Leigh Ann Westbrook, V.Anne Shibahara, Hiroaki Shetty, Jagathpala Coonrod, Scott A. Reddi, P.Prabhakara Shannon, John Kinter, Michael Sherman, Nicholas E. Fox, Jay Flickinger, Charles J. Herr, John C. |
| description | Modulation of the intracellular calcium concentration within mammalian spermatozoa is important in several pre-fertilization events including hyperactivated motility and the acrosome reaction. To identify calcium binding proteins (CBP) potentially regulating these processes, a 45Ca overlay technique was employed on 2-D blots of human sperm extracts. Microsequencing by Edman degradation and CAD mass spectrometry identified a relatively abundant 60.5 kDa CBP with a pI of 4.2 as calreticulin (CRT). Immunofluorescent labelling with anti-CRT antibodies localized CRT to the acrosome, with highest fluorescence in the equatorial segment, and in the cytoplasmic droplets of 94 and 48% of human spermatozoa respectively. Double immunolabelling experiments demonstrated co-localization of CRT and the inositol 1,4,5-trisphosphate receptor (IP3R) in the acrosome, in the equatorial segment, and vesicular structures in the cytoplasmic droplets of the neck region. Electron microscopic immunogold labelling localized CRT to the equatorial segment of acrosome-reacted spermatozoa and to membrane-enclosed vesicles within the cytoplasmic droplet of both acrosome-intact and acrosome-reacted spermatozoa. Localization of the IP3 receptor to the CRT-containing vesicles, in the sperm neck and to the acrosome, suggests that capacitative calcium entry in human spermatozoa may be regulated from these putative calcium storage sites. |
| doi_str_mv | 10.1093/molehr/7.10.923 |
| format | Article |
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To identify calcium binding proteins (CBP) potentially regulating these processes, a 45Ca overlay technique was employed on 2-D blots of human sperm extracts. Microsequencing by Edman degradation and CAD mass spectrometry identified a relatively abundant 60.5 kDa CBP with a pI of 4.2 as calreticulin (CRT). Immunofluorescent labelling with anti-CRT antibodies localized CRT to the acrosome, with highest fluorescence in the equatorial segment, and in the cytoplasmic droplets of 94 and 48% of human spermatozoa respectively. Double immunolabelling experiments demonstrated co-localization of CRT and the inositol 1,4,5-trisphosphate receptor (IP3R) in the acrosome, in the equatorial segment, and vesicular structures in the cytoplasmic droplets of the neck region. Electron microscopic immunogold labelling localized CRT to the equatorial segment of acrosome-reacted spermatozoa and to membrane-enclosed vesicles within the cytoplasmic droplet of both acrosome-intact and acrosome-reacted spermatozoa. Localization of the IP3 receptor to the CRT-containing vesicles, in the sperm neck and to the acrosome, suggests that capacitative calcium entry in human spermatozoa may be regulated from these putative calcium storage sites.</description><identifier>ISSN: 1360-9947</identifier><identifier>ISSN: 1460-2407</identifier><identifier>EISSN: 1460-2407</identifier><identifier>DOI: 10.1093/molehr/7.10.923</identifier><identifier>PMID: 11574661</identifier><language>eng</language><publisher>Oxford: Oxford University Press</publisher><subject>Acrosome - metabolism ; Biological and medical sciences ; Blotting, Northern ; Calcium - metabolism ; Calcium Channels - immunology ; Calcium Channels - metabolism ; calcium regulation ; calcium storage ; Calcium-Binding Proteins - genetics ; Calcium-Binding Proteins - immunology ; Calcium-Binding Proteins - metabolism ; Calreticulin ; Cell Membrane - metabolism ; Cytoplasmic Vesicles - metabolism ; Fundamental and applied biological sciences. Psychology ; Hormone metabolism and regulation ; human spermatozoa ; Humans ; Inositol 1,4,5-Trisphosphate Receptors ; inositol trisphosphate receptor ; Male ; Mammalian male genital system ; Microscopy, Immunoelectron ; Organ Specificity ; Receptors, Cytoplasmic and Nuclear - immunology ; Receptors, Cytoplasmic and Nuclear - metabolism ; Ribonucleoproteins - genetics ; Ribonucleoproteins - immunology ; Ribonucleoproteins - metabolism ; Vertebrates: reproduction</subject><ispartof>Molecular human reproduction, 2001-10, Vol.7 (10), p.923-933</ispartof><rights>2002 INIST-CNRS</rights><rights>Copyright Oxford University Press(England) Oct 2001</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c485t-348e8e86becb92742d103b82be306b42941cde5865cefdf42a392bcedcbd7fb13</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=14077036$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/11574661$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Naaby-Hansen, Soren</creatorcontrib><creatorcontrib>Wolkowicz, Michael J.</creatorcontrib><creatorcontrib>Klotz, Ken</creatorcontrib><creatorcontrib>Bush, Leigh Ann</creatorcontrib><creatorcontrib>Westbrook, V.Anne</creatorcontrib><creatorcontrib>Shibahara, Hiroaki</creatorcontrib><creatorcontrib>Shetty, Jagathpala</creatorcontrib><creatorcontrib>Coonrod, Scott A.</creatorcontrib><creatorcontrib>Reddi, P.Prabhakara</creatorcontrib><creatorcontrib>Shannon, John</creatorcontrib><creatorcontrib>Kinter, Michael</creatorcontrib><creatorcontrib>Sherman, Nicholas E.</creatorcontrib><creatorcontrib>Fox, Jay</creatorcontrib><creatorcontrib>Flickinger, Charles J.</creatorcontrib><creatorcontrib>Herr, John C.</creatorcontrib><title>Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa</title><title>Molecular human reproduction</title><addtitle>Mol. Hum. Reprod</addtitle><description>Modulation of the intracellular calcium concentration within mammalian spermatozoa is important in several pre-fertilization events including hyperactivated motility and the acrosome reaction. To identify calcium binding proteins (CBP) potentially regulating these processes, a 45Ca overlay technique was employed on 2-D blots of human sperm extracts. Microsequencing by Edman degradation and CAD mass spectrometry identified a relatively abundant 60.5 kDa CBP with a pI of 4.2 as calreticulin (CRT). Immunofluorescent labelling with anti-CRT antibodies localized CRT to the acrosome, with highest fluorescence in the equatorial segment, and in the cytoplasmic droplets of 94 and 48% of human spermatozoa respectively. Double immunolabelling experiments demonstrated co-localization of CRT and the inositol 1,4,5-trisphosphate receptor (IP3R) in the acrosome, in the equatorial segment, and vesicular structures in the cytoplasmic droplets of the neck region. Electron microscopic immunogold labelling localized CRT to the equatorial segment of acrosome-reacted spermatozoa and to membrane-enclosed vesicles within the cytoplasmic droplet of both acrosome-intact and acrosome-reacted spermatozoa. Localization of the IP3 receptor to the CRT-containing vesicles, in the sperm neck and to the acrosome, suggests that capacitative calcium entry in human spermatozoa may be regulated from these putative calcium storage sites.</description><subject>Acrosome - metabolism</subject><subject>Biological and medical sciences</subject><subject>Blotting, Northern</subject><subject>Calcium - metabolism</subject><subject>Calcium Channels - immunology</subject><subject>Calcium Channels - metabolism</subject><subject>calcium regulation</subject><subject>calcium storage</subject><subject>Calcium-Binding Proteins - genetics</subject><subject>Calcium-Binding Proteins - immunology</subject><subject>Calcium-Binding Proteins - metabolism</subject><subject>Calreticulin</subject><subject>Cell Membrane - metabolism</subject><subject>Cytoplasmic Vesicles - metabolism</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Hormone metabolism and regulation</subject><subject>human spermatozoa</subject><subject>Humans</subject><subject>Inositol 1,4,5-Trisphosphate Receptors</subject><subject>inositol trisphosphate receptor</subject><subject>Male</subject><subject>Mammalian male genital system</subject><subject>Microscopy, Immunoelectron</subject><subject>Organ Specificity</subject><subject>Receptors, Cytoplasmic and Nuclear - immunology</subject><subject>Receptors, Cytoplasmic and Nuclear - metabolism</subject><subject>Ribonucleoproteins - genetics</subject><subject>Ribonucleoproteins - immunology</subject><subject>Ribonucleoproteins - metabolism</subject><subject>Vertebrates: reproduction</subject><issn>1360-9947</issn><issn>1460-2407</issn><issn>1460-2407</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2001</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkl9rFDEUxQdRbF199k2CoE-dbv7NZOexLNoqC4JUkL6EJHPHTZ2ZTJOM2H4yP553u6sFXySE3OT-ziHhpCheMnrKaCOWQ-hhG5cKd6cNF4-KYyZrWnJJ1WOsBdZNI9VR8Syla0qZ4tXqaXHEWKVkXbPj4tc6lH1wpvd3JvswktCRvAXix5B8Dj1hJ_KkKnP0adoGnCYDieBgyiESM7YEtRGyd3PvR5Tdq-FmNtj3picJvg0w5nsUuwMMNpoRiA3z2EJLfkDyrof0R-puc5h6kwbvSBuxhLy703YezEjSBHFA57tgnhdPOtMneHFYF8WX9-8u1xfl5tP5h_XZpnRyVeVSyBXgqC0423AlecuosCtuQdDaSt5I5lqoVnXloGs7yY1ouHXQOtuqzjKxKN7ufacYbmZIWQ8-Oeh7fESYk1aMNRWn4r8gpxXGwhWCr_8Br8McR3yE5hytVIXJLorlHnIxpBSh01P0g4m3mlG9i17vo9dqd4DRo-LVwXa2A7QP_CFrBN4cAJMwtA5jcD49cPhnFBU1cuWe8ynDz799E7_rWglV6YuvV3p9-Xmz_siu9Ln4DaB_y_0</recordid><startdate>20011001</startdate><enddate>20011001</enddate><creator>Naaby-Hansen, Soren</creator><creator>Wolkowicz, Michael J.</creator><creator>Klotz, Ken</creator><creator>Bush, Leigh Ann</creator><creator>Westbrook, V.Anne</creator><creator>Shibahara, Hiroaki</creator><creator>Shetty, Jagathpala</creator><creator>Coonrod, Scott A.</creator><creator>Reddi, P.Prabhakara</creator><creator>Shannon, John</creator><creator>Kinter, Michael</creator><creator>Sherman, Nicholas E.</creator><creator>Fox, Jay</creator><creator>Flickinger, Charles J.</creator><creator>Herr, John C.</creator><general>Oxford University Press</general><general>Oxford Publishing Limited (England)</general><scope>BSCLL</scope><scope>IQODW</scope><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>7QP</scope><scope>7TM</scope><scope>8FD</scope><scope>FR3</scope><scope>K9.</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>20011001</creationdate><title>Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa</title><author>Naaby-Hansen, Soren ; Wolkowicz, Michael J. ; Klotz, Ken ; Bush, Leigh Ann ; Westbrook, V.Anne ; Shibahara, Hiroaki ; Shetty, Jagathpala ; Coonrod, Scott A. ; Reddi, P.Prabhakara ; Shannon, John ; Kinter, Michael ; Sherman, Nicholas E. ; Fox, Jay ; Flickinger, Charles J. ; Herr, John C.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c485t-348e8e86becb92742d103b82be306b42941cde5865cefdf42a392bcedcbd7fb13</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2001</creationdate><topic>Acrosome - metabolism</topic><topic>Biological and medical sciences</topic><topic>Blotting, Northern</topic><topic>Calcium - metabolism</topic><topic>Calcium Channels - immunology</topic><topic>Calcium Channels - metabolism</topic><topic>calcium regulation</topic><topic>calcium storage</topic><topic>Calcium-Binding Proteins - genetics</topic><topic>Calcium-Binding Proteins - immunology</topic><topic>Calcium-Binding Proteins - metabolism</topic><topic>Calreticulin</topic><topic>Cell Membrane - metabolism</topic><topic>Cytoplasmic Vesicles - metabolism</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Hormone metabolism and regulation</topic><topic>human spermatozoa</topic><topic>Humans</topic><topic>Inositol 1,4,5-Trisphosphate Receptors</topic><topic>inositol trisphosphate receptor</topic><topic>Male</topic><topic>Mammalian male genital system</topic><topic>Microscopy, Immunoelectron</topic><topic>Organ Specificity</topic><topic>Receptors, Cytoplasmic and Nuclear - immunology</topic><topic>Receptors, Cytoplasmic and Nuclear - metabolism</topic><topic>Ribonucleoproteins - genetics</topic><topic>Ribonucleoproteins - immunology</topic><topic>Ribonucleoproteins - metabolism</topic><topic>Vertebrates: reproduction</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Naaby-Hansen, Soren</creatorcontrib><creatorcontrib>Wolkowicz, Michael J.</creatorcontrib><creatorcontrib>Klotz, Ken</creatorcontrib><creatorcontrib>Bush, Leigh Ann</creatorcontrib><creatorcontrib>Westbrook, V.Anne</creatorcontrib><creatorcontrib>Shibahara, Hiroaki</creatorcontrib><creatorcontrib>Shetty, Jagathpala</creatorcontrib><creatorcontrib>Coonrod, Scott A.</creatorcontrib><creatorcontrib>Reddi, P.Prabhakara</creatorcontrib><creatorcontrib>Shannon, John</creatorcontrib><creatorcontrib>Kinter, Michael</creatorcontrib><creatorcontrib>Sherman, Nicholas E.</creatorcontrib><creatorcontrib>Fox, Jay</creatorcontrib><creatorcontrib>Flickinger, Charles J.</creatorcontrib><creatorcontrib>Herr, John C.</creatorcontrib><collection>Istex</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Calcium & Calcified Tissue Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Molecular human reproduction</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Naaby-Hansen, Soren</au><au>Wolkowicz, Michael J.</au><au>Klotz, Ken</au><au>Bush, Leigh Ann</au><au>Westbrook, V.Anne</au><au>Shibahara, Hiroaki</au><au>Shetty, Jagathpala</au><au>Coonrod, Scott A.</au><au>Reddi, P.Prabhakara</au><au>Shannon, John</au><au>Kinter, Michael</au><au>Sherman, Nicholas E.</au><au>Fox, Jay</au><au>Flickinger, Charles J.</au><au>Herr, John C.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa</atitle><jtitle>Molecular human reproduction</jtitle><addtitle>Mol. Hum. Reprod</addtitle><date>2001-10-01</date><risdate>2001</risdate><volume>7</volume><issue>10</issue><spage>923</spage><epage>933</epage><pages>923-933</pages><issn>1360-9947</issn><issn>1460-2407</issn><eissn>1460-2407</eissn><abstract>Modulation of the intracellular calcium concentration within mammalian spermatozoa is important in several pre-fertilization events including hyperactivated motility and the acrosome reaction. To identify calcium binding proteins (CBP) potentially regulating these processes, a 45Ca overlay technique was employed on 2-D blots of human sperm extracts. Microsequencing by Edman degradation and CAD mass spectrometry identified a relatively abundant 60.5 kDa CBP with a pI of 4.2 as calreticulin (CRT). Immunofluorescent labelling with anti-CRT antibodies localized CRT to the acrosome, with highest fluorescence in the equatorial segment, and in the cytoplasmic droplets of 94 and 48% of human spermatozoa respectively. Double immunolabelling experiments demonstrated co-localization of CRT and the inositol 1,4,5-trisphosphate receptor (IP3R) in the acrosome, in the equatorial segment, and vesicular structures in the cytoplasmic droplets of the neck region. Electron microscopic immunogold labelling localized CRT to the equatorial segment of acrosome-reacted spermatozoa and to membrane-enclosed vesicles within the cytoplasmic droplet of both acrosome-intact and acrosome-reacted spermatozoa. Localization of the IP3 receptor to the CRT-containing vesicles, in the sperm neck and to the acrosome, suggests that capacitative calcium entry in human spermatozoa may be regulated from these putative calcium storage sites.</abstract><cop>Oxford</cop><pub>Oxford University Press</pub><pmid>11574661</pmid><doi>10.1093/molehr/7.10.923</doi><tpages>11</tpages></addata></record> |
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| subjects | Acrosome - metabolism Biological and medical sciences Blotting, Northern Calcium - metabolism Calcium Channels - immunology Calcium Channels - metabolism calcium regulation calcium storage Calcium-Binding Proteins - genetics Calcium-Binding Proteins - immunology Calcium-Binding Proteins - metabolism Calreticulin Cell Membrane - metabolism Cytoplasmic Vesicles - metabolism Fundamental and applied biological sciences. Psychology Hormone metabolism and regulation human spermatozoa Humans Inositol 1,4,5-Trisphosphate Receptors inositol trisphosphate receptor Male Mammalian male genital system Microscopy, Immunoelectron Organ Specificity Receptors, Cytoplasmic and Nuclear - immunology Receptors, Cytoplasmic and Nuclear - metabolism Ribonucleoproteins - genetics Ribonucleoproteins - immunology Ribonucleoproteins - metabolism Vertebrates: reproduction |
| title | Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa |
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