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Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region

Phospholipase C-zeta (PLCζ) is a strong candidate for the mammalian sperm-derived factor that triggers the Ca(2+) oscillations required for egg activation at fertilization. PLCζ lacks a PH domain, which targets PLCδ1 to the phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) substrate in the pla...

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Autores principales: Nomikos, Michail, Elgmati, Khalil, Theodoridou, Maria, Calver, Brian L., Nounesis, George, Swann, Karl, Lai, F. Anthony
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Company of Biologists 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3138701/
https://www.ncbi.nlm.nih.gov/pubmed/21730019
http://dx.doi.org/10.1242/jcs.083485
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author Nomikos, Michail
Elgmati, Khalil
Theodoridou, Maria
Calver, Brian L.
Nounesis, George
Swann, Karl
Lai, F. Anthony
author_facet Nomikos, Michail
Elgmati, Khalil
Theodoridou, Maria
Calver, Brian L.
Nounesis, George
Swann, Karl
Lai, F. Anthony
author_sort Nomikos, Michail
collection PubMed
description Phospholipase C-zeta (PLCζ) is a strong candidate for the mammalian sperm-derived factor that triggers the Ca(2+) oscillations required for egg activation at fertilization. PLCζ lacks a PH domain, which targets PLCδ1 to the phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) substrate in the plasma membrane. Previous studies failed to detect PLCζ in the plasma membrane, hence the means of PLCζ binding to PtdIns(4,5)P(2) is unclear. We find that the PLCζ XY linker, but not the C2 domain, exhibits robust binding to PtdIns(4,5)P(2) or to liposomes containing near-physiological levels of PtdIns(4,5)P(2). The role of positively charged residues within the XY linker was addressed by sequentially substituting alanines for three lysine residues, K374, K375 and K377. Microinjection of these mutants into mouse eggs enabled their Ca(2+) oscillation-inducing activities to be compared with wild-type PLCζ. The XY-linker mutant proteins were purified and the in vitro PtdIns(4,5)P(2) hydrolysis and binding properties were monitored. Successive reduction of net positive charge within the PLCζ XY linker significantly affects both in vivo Ca(2+)-oscillation-inducing activity and in vitro PtdIns(4,5)P(2) interaction of mouse PLCζ. Our data suggest that positively charged residues within the XY linker play an important role in the PLCζ interaction with PtdIns(4,5)P(2), a crucial step in generating the Ca(2+) activation signal that is essential for fertilization in mammals.
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spelling pubmed-31387012011-08-01 Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region Nomikos, Michail Elgmati, Khalil Theodoridou, Maria Calver, Brian L. Nounesis, George Swann, Karl Lai, F. Anthony J Cell Sci Research Articles Phospholipase C-zeta (PLCζ) is a strong candidate for the mammalian sperm-derived factor that triggers the Ca(2+) oscillations required for egg activation at fertilization. PLCζ lacks a PH domain, which targets PLCδ1 to the phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)) substrate in the plasma membrane. Previous studies failed to detect PLCζ in the plasma membrane, hence the means of PLCζ binding to PtdIns(4,5)P(2) is unclear. We find that the PLCζ XY linker, but not the C2 domain, exhibits robust binding to PtdIns(4,5)P(2) or to liposomes containing near-physiological levels of PtdIns(4,5)P(2). The role of positively charged residues within the XY linker was addressed by sequentially substituting alanines for three lysine residues, K374, K375 and K377. Microinjection of these mutants into mouse eggs enabled their Ca(2+) oscillation-inducing activities to be compared with wild-type PLCζ. The XY-linker mutant proteins were purified and the in vitro PtdIns(4,5)P(2) hydrolysis and binding properties were monitored. Successive reduction of net positive charge within the PLCζ XY linker significantly affects both in vivo Ca(2+)-oscillation-inducing activity and in vitro PtdIns(4,5)P(2) interaction of mouse PLCζ. Our data suggest that positively charged residues within the XY linker play an important role in the PLCζ interaction with PtdIns(4,5)P(2), a crucial step in generating the Ca(2+) activation signal that is essential for fertilization in mammals. Company of Biologists 2011-08-01 /pmc/articles/PMC3138701/ /pubmed/21730019 http://dx.doi.org/10.1242/jcs.083485 Text en © 2011. http://creativecommons.org/licenses/by-nc-sa/3.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Share Alike License (http://creativecommons.org/licenses/by-nc-sa/3.0), which permits unrestricted non-commercial use, distribution and reproduction in any medium provided that the original work is properly cited and all further distributions of the work or adaptation are subject to the same Creative Commons License terms.
spellingShingle Research Articles
Nomikos, Michail
Elgmati, Khalil
Theodoridou, Maria
Calver, Brian L.
Nounesis, George
Swann, Karl
Lai, F. Anthony
Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title_full Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title_fullStr Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title_full_unstemmed Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title_short Phospholipase Cζ binding to PtdIns(4,5)P(2) requires the XY-linker region
title_sort phospholipase cζ binding to ptdins(4,5)p(2) requires the xy-linker region
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3138701/
https://www.ncbi.nlm.nih.gov/pubmed/21730019
http://dx.doi.org/10.1242/jcs.083485
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