Dissertation
Investigation into the functions of the pollen specific genes PiVAMP721 and PiSCP1 in pollen tube growth
Washington State University
Doctor of Philosophy (PhD), Washington State University
12/2008
DOI:
https://doi.org/10.7273/000005793
Abstract
PiVAMP721 is a member of the plant unique VAMP7 proteins, which is expressed specifically in pollen. The VAMP7 (longin) proteins are involved in endosomal trafficking across species. PiVAMP721 was colocalized with an endocytic marker, FM4-64 and it's over-expression was found to be inhibitory to pollen tube growth. PiVAMP721/C, a truncated constitutively active form of this protein, generated bulging pollen tube tips at a frequency 3 fold higher than the than wild type protein, and lead to increased growth inhibition (mean pollen tube length 26% shorter than tubes expressing the wild type protein). In contrast expression of the truncated N-terminal "longin" domain, PiVAMP721/N, in pollen tubes did not generate a discernable phenotype. Interestingly
PiVAMP721/N was found to be able to partially rescue inhibition caused by expressing PiVAMP721/C, suggesting that PiVAMP721/N has a negative regulatory function and that the two regions of this protein are able to interact and function "normal" even when not translationally fused. Although the small novel protein, PiSCP1, has two homologs in Arabidopsis, neither has a known function. We showed that PiSCP1 is a peroxisomal protein by colocalization with a peroxisome marker. Overexpression of PiSCP1 did not generate a significant phenotype. However, the protein encoded by this gene interacted with CDPKs in yeast two hybrid assays and mutations of two potential sites of serine phosphorylation lead to proteins that inhibited pollen tube growth. These results suggest that phosphorylation of either of these two residues is important to the overall function of PiSCP1. The conserved N-terminal "A" and C-terminal "C" domains both retained the ability to localizes to peroxisomes and may still retain the ability to interact with CDPK(s). The divergent central B domain was found to be largely cytosolic and not to interact with CDPK(s). Here, we further showed that PiCDPK2 is also a peroxisomal protein. PiCDPK2 was shown to colocalize with PiSCP1 and interact with this protein in a yeast two hybrid assay, suggesting that PiCDPK2 is likely the CDPK isoform that interacts with PiSCP1 in vivo. Neither PiSCP1 nor PiCDPK2 have classic peroxisome targeting signals, and the potential targeting/sorting signals of these two proteins are discussed.
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Details
- Title
- Investigation into the functions of the pollen specific genes PiVAMP721 and PiSCP1 in pollen tube growth
- Creators
- Feng Guo
- Contributors
- Andrew McCubbin (Chair) - Washington State University, School of Biological SciencesMechthild Tegeder (Committee Member) - Washington State University, School of Biological SciencesThomas W Okita (Committee Member) - Washington State University, Institute of Biological Chemistry
- Awarding Institution
- Washington State University
- Academic Unit
- School of Biological Sciences
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Publisher
- Washington State University
- Number of pages
- 187
- Identifiers
- 99901055140401842
- Language
- English
- Resource Type
- Dissertation