Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells
It is well known that in fMLP‐stimulated neutrophils, phosphatidyl inositol 3,4,5‐trisphosphate [PI(3,4,5)P3] localizes at the leading edge of the cells. However, no effort has been made to study the PI 4,5‐bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(...
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| Veröffentlicht in: | Journal of leukocyte biology 2008-08, Vol.84 (2), p.440-447 |
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| creator | Sharma, Ved P. DesMarais, Vera Sumners, Colin Shaw, Gerry Narang, Atul |
| description | It is well known that in fMLP‐stimulated neutrophils, phosphatidyl inositol 3,4,5‐trisphosphate [PI(3,4,5)P3] localizes at the leading edge of the cells. However, no effort has been made to study the PI 4,5‐bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(4,5)P2 is unlikely to localize, as its basal level is orders of magnitude higher than that of PI(3,4,5)P3. We developed an optimized immunostaining protocol for studying the endogenous distribution of PI(4,5)P2 in neutrophil‐like HL‐60 cells. We show that PI(4,5)P2 localizes sharply at the leading edge with an intensity gradient similar to that for PI(3,4,5)P3. The enzymes for the production of PI(4,5)P2, namely, PI5KIα and PI5KIγ, were also found to localize at the leading edge, further supporting our finding that PI(4,5)P2 localizes at the leading edge. These results imply that complementary regulation of PI3K and phosphate and tensin homolog (PTEN) is not the sole or dominant mechanism of PI(3,4,5)P3 polarization in HL‐60 cells. |
| doi_str_mv | 10.1189/jlb.0907636 |
| format | Article |
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However, no effort has been made to study the PI 4,5‐bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(4,5)P2 is unlikely to localize, as its basal level is orders of magnitude higher than that of PI(3,4,5)P3. We developed an optimized immunostaining protocol for studying the endogenous distribution of PI(4,5)P2 in neutrophil‐like HL‐60 cells. We show that PI(4,5)P2 localizes sharply at the leading edge with an intensity gradient similar to that for PI(3,4,5)P3. The enzymes for the production of PI(4,5)P2, namely, PI5KIα and PI5KIγ, were also found to localize at the leading edge, further supporting our finding that PI(4,5)P2 localizes at the leading edge. These results imply that complementary regulation of PI3K and phosphate and tensin homolog (PTEN) is not the sole or dominant mechanism of PI(3,4,5)P3 polarization in HL‐60 cells.</description><identifier>ISSN: 0741-5400</identifier><identifier>EISSN: 1938-3673</identifier><identifier>DOI: 10.1189/jlb.0907636</identifier><identifier>PMID: 18477691</identifier><language>eng</language><publisher>United States: Society for Leukocyte Biology</publisher><subject>Biomarkers - analysis ; Cell Culture Techniques ; Cell Development, Growth, Differentiation, and Function ; Cell Differentiation ; Cell Membrane - ultrastructure ; chemotaxis ; Enzyme-Linked Immunosorbent Assay ; gradient sensing ; HL-60 Cells - cytology ; Humans ; Immunohistochemistry ; Phosphatidylinositol 4,5-Diphosphate - metabolism ; Phosphatidylinositol Phosphates - metabolism ; phosphoinositides</subject><ispartof>Journal of leukocyte biology, 2008-08, Vol.84 (2), p.440-447</ispartof><rights>2008 Society for Leukocyte Biology</rights><rights>Society for Leukocyte Biology 2008</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4310-22053b4bf95d87d4018fe44e0650ceb3399a4ab98b96aae99414a654a99c83f03</citedby><cites>FETCH-LOGICAL-c4310-22053b4bf95d87d4018fe44e0650ceb3399a4ab98b96aae99414a654a99c83f03</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1189%2Fjlb.0907636$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1189%2Fjlb.0907636$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>230,314,780,784,885,1417,27924,27925,45574,45575</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/18477691$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Sharma, Ved P.</creatorcontrib><creatorcontrib>DesMarais, Vera</creatorcontrib><creatorcontrib>Sumners, Colin</creatorcontrib><creatorcontrib>Shaw, Gerry</creatorcontrib><creatorcontrib>Narang, Atul</creatorcontrib><title>Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells</title><title>Journal of leukocyte biology</title><addtitle>J Leukoc Biol</addtitle><description>It is well known that in fMLP‐stimulated neutrophils, phosphatidyl inositol 3,4,5‐trisphosphate [PI(3,4,5)P3] localizes at the leading edge of the cells. However, no effort has been made to study the PI 4,5‐bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(4,5)P2 is unlikely to localize, as its basal level is orders of magnitude higher than that of PI(3,4,5)P3. We developed an optimized immunostaining protocol for studying the endogenous distribution of PI(4,5)P2 in neutrophil‐like HL‐60 cells. We show that PI(4,5)P2 localizes sharply at the leading edge with an intensity gradient similar to that for PI(3,4,5)P3. The enzymes for the production of PI(4,5)P2, namely, PI5KIα and PI5KIγ, were also found to localize at the leading edge, further supporting our finding that PI(4,5)P2 localizes at the leading edge. These results imply that complementary regulation of PI3K and phosphate and tensin homolog (PTEN) is not the sole or dominant mechanism of PI(3,4,5)P3 polarization in HL‐60 cells.</description><subject>Biomarkers - analysis</subject><subject>Cell Culture Techniques</subject><subject>Cell Development, Growth, Differentiation, and Function</subject><subject>Cell Differentiation</subject><subject>Cell Membrane - ultrastructure</subject><subject>chemotaxis</subject><subject>Enzyme-Linked Immunosorbent Assay</subject><subject>gradient sensing</subject><subject>HL-60 Cells - cytology</subject><subject>Humans</subject><subject>Immunohistochemistry</subject><subject>Phosphatidylinositol 4,5-Diphosphate - metabolism</subject><subject>Phosphatidylinositol Phosphates - metabolism</subject><subject>phosphoinositides</subject><issn>0741-5400</issn><issn>1938-3673</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kc1v1DAQxSMEotvCiTvyBQSClHHsOPYFiVZAF61ED3C2HGeyceXE29hpVP56suyKjwunOcxv3sybl2XPKJxTKtW7G1-fg4JKMPEgW1HFZM5ExR5mK6g4zUsOcJKdxngDAKwQ8Dg7oZJXlVB0le3WfT8NISbjBjdsCd65BgeLpA0juV6_4m_L19cF8cEa736Y5MJATCKpQ-LRNL9Gmi2S0BLbYR9MSqOxyQwpj8n1kzcJG3K1yQUQi97HJ9mj1viIT4_1LPv-6eO3y6t88_Xz-vLDJrecUciLAkpW87pVZSOrhgOVLXKOIEqwWDOmlOGmVrJWwhhUilNuRMmNUlayFthZ9v6gu5vqHhuLw3KY17vR9Wa818E4_W9ncJ3ehjtdVFRK4IvAy6PAGG4njEn3Lu4tmAHDFLVYHq0KphbwzQG0Y4hxxPb3Egp6n5BeEtLHhBb6-d93_WGPkSwAHIDZebz_n5b-srkAzvdeXxxGOrftZjeijr3xftlQ6HmeJdeF3nM_AZrmqN8</recordid><startdate>200808</startdate><enddate>200808</enddate><creator>Sharma, Ved P.</creator><creator>DesMarais, Vera</creator><creator>Sumners, Colin</creator><creator>Shaw, Gerry</creator><creator>Narang, Atul</creator><general>Society for Leukocyte Biology</general><general>The Society for Leukocyte Biology</general><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>7X8</scope><scope>5PM</scope></search><sort><creationdate>200808</creationdate><title>Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells</title><author>Sharma, Ved P. ; DesMarais, Vera ; Sumners, Colin ; Shaw, Gerry ; Narang, Atul</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c4310-22053b4bf95d87d4018fe44e0650ceb3399a4ab98b96aae99414a654a99c83f03</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2008</creationdate><topic>Biomarkers - analysis</topic><topic>Cell Culture Techniques</topic><topic>Cell Development, Growth, Differentiation, and Function</topic><topic>Cell Differentiation</topic><topic>Cell Membrane - ultrastructure</topic><topic>chemotaxis</topic><topic>Enzyme-Linked Immunosorbent Assay</topic><topic>gradient sensing</topic><topic>HL-60 Cells - cytology</topic><topic>Humans</topic><topic>Immunohistochemistry</topic><topic>Phosphatidylinositol 4,5-Diphosphate - metabolism</topic><topic>Phosphatidylinositol Phosphates - metabolism</topic><topic>phosphoinositides</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Sharma, Ved P.</creatorcontrib><creatorcontrib>DesMarais, Vera</creatorcontrib><creatorcontrib>Sumners, Colin</creatorcontrib><creatorcontrib>Shaw, Gerry</creatorcontrib><creatorcontrib>Narang, Atul</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Journal of leukocyte biology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Sharma, Ved P.</au><au>DesMarais, Vera</au><au>Sumners, Colin</au><au>Shaw, Gerry</au><au>Narang, Atul</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells</atitle><jtitle>Journal of leukocyte biology</jtitle><addtitle>J Leukoc Biol</addtitle><date>2008-08</date><risdate>2008</risdate><volume>84</volume><issue>2</issue><spage>440</spage><epage>447</epage><pages>440-447</pages><issn>0741-5400</issn><eissn>1938-3673</eissn><abstract>It is well known that in fMLP‐stimulated neutrophils, phosphatidyl inositol 3,4,5‐trisphosphate [PI(3,4,5)P3] localizes at the leading edge of the cells. However, no effort has been made to study the PI 4,5‐bisphosphate [PI(4,5)P2] distribution in these cells. In fact, it has been suggested that PI(4,5)P2 is unlikely to localize, as its basal level is orders of magnitude higher than that of PI(3,4,5)P3. We developed an optimized immunostaining protocol for studying the endogenous distribution of PI(4,5)P2 in neutrophil‐like HL‐60 cells. We show that PI(4,5)P2 localizes sharply at the leading edge with an intensity gradient similar to that for PI(3,4,5)P3. The enzymes for the production of PI(4,5)P2, namely, PI5KIα and PI5KIγ, were also found to localize at the leading edge, further supporting our finding that PI(4,5)P2 localizes at the leading edge. These results imply that complementary regulation of PI3K and phosphate and tensin homolog (PTEN) is not the sole or dominant mechanism of PI(3,4,5)P3 polarization in HL‐60 cells.</abstract><cop>United States</cop><pub>Society for Leukocyte Biology</pub><pmid>18477691</pmid><doi>10.1189/jlb.0907636</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Biomarkers - analysis Cell Culture Techniques Cell Development, Growth, Differentiation, and Function Cell Differentiation Cell Membrane - ultrastructure chemotaxis Enzyme-Linked Immunosorbent Assay gradient sensing HL-60 Cells - cytology Humans Immunohistochemistry Phosphatidylinositol 4,5-Diphosphate - metabolism Phosphatidylinositol Phosphates - metabolism phosphoinositides |
| title | Immunostaining evidence for PI(4,5)P2 localization at the leading edge of chemoattractant-stimulated HL-60 cells |
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