Yoon, Gyeong Mee , Guo, Feng , McCubbin, Andrew G .
Characterization of PiSCP1, a novel substrate of a CDPK that regulates pollen tube growth polarity.
Calcium plays a pivotal role in pollen tube growth, and we have previously demonstrated that a calcium-dependent protein kinase isoform from Petunia inflata (PiCDPK1) is critical to the regulation of growth polarity. To further characterize PiCDPK1 mediated pathways we have sought to identify its substrates by yeast 2-hybrid library screening. One of the PiCDPK1 interacting cDNAs identified in the library screen, termed Small CDPK-interacting Protein 1 (PiSCP1) encodes a novel polypeptide of 103 amino acids. Confirmation of the interaction of PiSCP1 and PiCDPK1 was confirmed using a pull-down assay and an in vitro phosphorylation assay. The Arabidopsis genome contains two PiSCP1 homologs, which share extensive sequence identity in their N- and C-termini, but are otherwise highly divergent. PiSCP1 is capable of rescuing loss of growth polarity in pollen tubes caused by over-expressing PiCDPK1, suggesting that it is a bona fide in vivo substrate. In an attempt to gain insight into the function of PiSCP1 we have expressed wildtype, truncated, and phospho-mutant constructs in pollen tubes. While over-expression of the wild type protein does not discernibly affect, expression of truncated or phospho-mutant constructs severely inhibits pollen tube growth. These experiments have provided evidence for phosphorylation sites at both the N- and C-termini, which are conserved between PiSCP1 and its Arabidopsis homologs. The N-termini share 32% sequence identity with the A domain of neuronal synapsin, including a regulatory phosphorylation site. Synapsin plays a key role in neurotransmitter release by acting as a phospho-reversible tether between vesicles and the actin cytoskeleton. We are currently investigating whether PiSCP1 plays an analogous role in pollen tubes.
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1 - Washington State University, School of Biological Sciences and Center of Reproductive Biology, Pullman, Wa, 99164-4236, USA
2 - Washington State University, School of Biological Sciences and Center of Reproductive Biology
pollen tube growth
Presentation Type: Plant Biology Abstract
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM