Immobilizing chemically synthesized analogues of PI(3,4,5)P3 onto Affi-10 beads and incorporating them into liposomes allowed their use as affinity absorbents in the comprehensive analysis of the phosphoinositide interactome using cytosolic cell extracts of the LIM1215 colon cancer cell line. This led to the identification of 282 proteins that either interact with PI(3,4,5)P3 or are indirectly captured as part of a complex containing a PI(3,4,5)P3 binding partner. Identification of the proteins was achieved using affinity/LC-MS/MS experiments.
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Activation of Src and release of intracellular calcium by phosphatidic acid during Xenopus laevis fertilization.
Dev Biol
February 2014
University of Colorado Denver, Denver, CO 80217-3364, USA. Electronic address:
We report a new step in the fertilization in Xenopus laevis which has been found to involve activation of Src tyrosine kinase to stimulate phospholipase C-γ (PLC-γ) which increases inositol 1,4,5-trisphosphate (IP3) to release intracellular calcium ([Ca](i)). Molecular species analysis and mass measurements suggested that sperm activate phospholipase D (PLD) to elevate phosphatidic acid (PA). We now report that PA mass increased 2.
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