Protein tyrosine phosphatases in Chaetopterus egg activation.
Changes in protein tyrosine phosphorylation are an essential aspect of egg activation after fertilization. Such changes result from the net contributions of both tyrosine kinases and phosphatases (PTP). This study was conducted to determine what role(s) PTP may have in egg activation. We identified four novel PTP in Chaetopterus pergamentaceus oocytes, cpPTPNT6, cpPTPNT7, cpPTPR2B, and cpPTPR2A, that have significant homology to, respectively, human PTPsigma, -rho, -D2 and -BAS. The first two are cytosolic and the latter two are transmembrane. Several PTP inhibitors were tested to see if they would affect Chaetopterus pergamentaceus fertilization. Eggs treated with beta-bromo-4-hydroxyacetophenone (PTP inhibitor 1) exhibited microvillar elongation, which is a sign of cortical changes resulting from activation. Those treated with Na3VO4 underwent full parthenogenetic activation, including polar body formation and pseudocleavage and did so independently of extracellular Ca2+, which is required for the Ca2+ oscillations that initiate development after fertilization. Fluorescence microscopy identified phosphotyrosine-containing proteins in the cortex and around the nucleus of vanadate-activated eggs, whereas in fertilized eggs they were concentrated only in the cortex. Immunoblots of vanadate-activated and fertilized eggs showed tyrosine hyperphosphorylation of approximately 140 kDa protein. These results suggest that PTP most likely maintain the egg in an inactive state by dephosphorylation of proteins independent of the Ca2+ oscillations in the activation process.
Hinton
Shantá D
SD
Department of Biology, Howard University, Washington, DC 20059, USA.
Yang
Dazhi
D
Eckberg
William R
WR
D000098:Acetophenones / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D000845:Annelida; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005306:Fertilization; D015151:Immunoblotting; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism*; D009991:Oscillometry; D010766:Phosphorylation; D010802:Phylogeny; D017027:Protein Tyrosine Phosphatases / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014443:Tyrosine / Q000378:metabolism
D000098:Acetophenones; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D014443:Tyrosine; D017027:Protein Tyrosine Phosphatases; C031335:4-hydroxyacetophenone; D002118:Calcium