Functional analysis of novel protein-protein interactions involving ROP GTPases in Arabidopsis thaliana and Populus trichocarpa

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dc.contributor.authorJia, Xiaoyanen
dc.contributor.committeechairBrunner, Amy M.en
dc.contributor.committeechairBeers, Eric P.en
dc.contributor.committeememberZhao, Bingyuen
dc.contributor.committeememberHolliday, Jason A.en
dc.contributor.departmentForest Resources and Environmental Conservationen
dc.date.accessioned2015-04-25T06:00:39Zen
dc.date.available2015-04-25T06:00:39Zen
dc.date.issued2013-09-02en
dc.description.abstractWe are using the yeast two-hybrid (Y2H) system to identify novel protein-protein interactions (PPI) relevant to wood formation. Bait proteins for Y2H binary assays and screening against a xylem cDNA prey library were selected from approximately 400 Populus trichocarpa genes that are at least 8-fold more highly expressed in differentiating secondary xylem versus phloem-cambium, and designated here as poplar biomass (PB) genes. Here we report some of the interactions involving selected PB proteins and efforts to characterize their functions in Populus and Arabidopsis. Members of the ROP GTPase family, PB15 in poplar and ROP11 in Arabidopsis, interact with the domain of unknown function (DUF) 620 (DUF620) proteins (e.g., PB129 in poplar). Ectopic co-expression of PB15 and PB129 in Arabidopsis caused outgrowths at the base of flower pedicels and altered leaf morphology. Interestingly, the co-expression phenotype could not be observed in transgenic plants that are only expressing either one of the interacting partners separately. Transgenics altered in expression of PB15 and/or PB129 were prepared in Populus and characterization of transgenic trees will be performed in greenhouse and field. In addition to DUF620 family proteins, ROP11 also interacts with the COP9 subunit CSN5A in Arabidopsis. We confirmed the interaction of ROP11 and CSN5A in Y2H and employed available mutants for ROP11 and CSN5A in Arabidopsis to genetically characterize this interaction. Surprisingly, loss of ROP11 was found to rescue the csn5a-2 pleiotropic phenotype. Ectopic expression of a ROP11 dominant negative mutant in the csn5a-2 background also complemented the stunted growth phenotype. Transcript analysis and gel blot assays showed that CSN5A transcript levels remained unchanged in all rescue lines, whereas CSN5A protein levels increased relative to WT. Taken together, we concluded that ROP11 negatively regulate CSN5A protein level in plant by some as yet unknown mechanism.en
dc.description.degreePh. D.en
dc.format.mediumETDen
dc.identifier.othervt_gsexam:1568en
dc.identifier.urihttp://hdl.handle.net/10919/51802en
dc.publisherVirginia Techen
dc.rightsIn Copyrighten
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/en
dc.subjectprotein-protein interactionen
dc.subjectyeast two-hybriden
dc.subjectROP GTPaseen
dc.subjectROP11en
dc.subjectCSN5en
dc.subjectArabidopsis thalianaen
dc.subjectPopulus trichocarpaen
dc.titleFunctional analysis of novel protein-protein interactions involving ROP GTPases in Arabidopsis thaliana and Populus trichocarpaen
dc.typeDissertationen
thesis.degree.disciplineForestryen
thesis.degree.grantorVirginia Polytechnic Institute and State Universityen
thesis.degree.leveldoctoralen
thesis.degree.namePh. D.en

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