Proteomics Guided Gene Discovery for Plant Growth and Defense
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Research carried out in this dissertation aims to discover and validate novel gene functions in plant defense and growth. Plants deploy many mechanisms to respond and overcome biological, physical, and environmental challenges. By taking advantage of proteomics, we can discover dynamic changes of the proteome during the plant response. Nonetheless, the complexity of the proteome and the effect of high abundant proteins hinder the discovery of proteins with vital functions at low abundance. I developed a new sample preparation method, dubbed Polyethyleneimine Assisted Rubisco Cleanup (PARC) to fractionate and deplete an abundant protein, namely Rubisco, from plant protein samples. The new approach was applied to investigate mechanisms for plant defense against herbivorous insects. My results indicated that PARC can effectively remove Rubisco and almost two times more differentially regulated proteins were identified. Over-expression of jacalin-like and cupin-like genes was carried out to validate their role in insect resistance in rice. The results further highlighted that PARC can serve as an effective strategy for gene discovery. Furthermore, I integrated a rapid sample preparation method and bioinformatics classification system for comparative analysis of plant responses to two plant hormones, zeatin and brassinosteroid (BR). My data showed the metabolic pathways in sucrose and starch biosynthesis and utilization were significantly changed in zeatin treated plants, yet the lipid biosynthesis remained unchanged. For brassinosteroid treated plants, lipid biosynthesis and β-oxidation were both down-regulated, yet the changes in sucrose and starch metabolism were minor. Finally, a prohibitin (PHB8) involved in plant growth regulation was discovered by bioinformatics and proteomics methods. The over-expression of PHB8 in Arabidopsis increased the plant height, stem diameter, branch number and seed yield significantly. Downstream proteomics revealed that due to PHB8 over-expression, an ATPase beta subunit is significantly up-regulated. Further genetic study showed that over-expression of ATPase led to a similar phenotype as PHB8 over-expression lines. Pull-down assays revealed that PHB8 interacts with PHB9 and PHB16 to regulate ATPase level and energy metabolism. Overall, my research shows that the latest proteomics platform and appropriate sample preparation methods can facilitate the discovery of genes involved in plant defense and growth regulation.
Zhang, Yixiang (2014). Proteomics Guided Gene Discovery for Plant Growth and Defense. Doctoral dissertation, Texas A&M University. Available electronically from