Enkurin is a novel calmodulin and TRPC channel binding protein in sperm.
The TRPC cation channel family has been implicated in receptor- or phospholipase C (PLC)-mediated Ca2+ entry into animal cells. These channels are present in mammalian sperm and are assigned a role in ZP3-evoked Ca2+ influx that drives acrosome reactions. However, the mechanisms controlling channel activity and coupling Ca2+ entry through these channels to cellular responses are not well understood. A yeast two-hybrid screen was carried out to identify TRPC-interacting proteins that would be candidate regulators or effectors. We identified a novel protein, enkurin, that is expressed at high levels in the testis and vomeronasal organ and at lower levels in selected other tissues. Enkurin interacts with several TRPC proteins (TRPC1, TRPC2, TRPC5, but not TRPC3) and colocalizes with these channels in sperm. Three protein-protein interaction domains were identified in enkurin: a C-terminal region is essential for channel interaction; an IQ motif binds the Ca2+ sensor, calmodulin, in a Ca2+-dependent manner; and a proline-rich N-terminal region contains predicted ligand sequences for SH3 domain proteins, including the SH3 domain of the p85 regulatory subunit of 1-phosphatidylinositol-3-kinase. We suggest that enkurin is an adaptor that functions to localize a Ca2+ sensitive signal transduction machinery in sperm to a Ca2+-permeable ion channel.
Sutton
Keith A
KA
Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01655, USA.
Jungnickel
Melissa K
MK
Wang
Yanli
Y
Cullen
Kay
K
Lambert
Stephen
S
Florman
Harvey M
HM
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology; D002147:Calmodulin / Q000378:metabolism*; D002148:Calmodulin-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029589:Seminal Plasma Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D013094:Spermatozoa / Q000166:cytology / Q000378:metabolism*; D050052:TRPC Cation Channels; D014018:Tissue Distribution; D020798:Two-Hybrid System Techniques
D048868:Adaptor Proteins, Signal Transducing; D015220:Calcium Channels; D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; C493290:ENKUR protein, human; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D029589:Seminal Plasma Proteins; D050052:TRPC Cation Channels; C493289:enkurin protein, mouse; D019869:Phosphatidylinositol 3-Kinases; D002118:Calcium