טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentCarp Nov Marie Jeanne
SubjectGenomics and Proteomics of Post Harvest Leaf Senescence
DepartmentDepartment of Biotechnology
Supervisor Professor Emeritus Shimon Gepstein


Abstract

Leaf senescence, the final stage of leaf development, is characterized by drastic catabolic changes resulting in extensive cellular dysfunction and disintegration and has an important impact on agriculture, affecting crop yield and the shelf life of leafy vegetables. Leaf senescence is a highly regulated and active process, characterized by differential and sequential changes in almost every sub cellular compartment controlled and executed by a specific set of genes and proteins. We have used a suppression subtraction hybridization approach to identify transcripts preferentially expressed during post harvest leaf senescence as compared to developmental senescence. The study identified common and unique transcripts for post harvest and developmental senescence. In addition we show different expression patterns; early and late upregulated senescence genes. Functional classification revealed genes involved in macromolecular degradation, stress response, hormonal and transcriptional regulation. T-DNA insertion lines for the identified genes were obtained and the knock-out transgenic plants did not show any phenotypic alterations during developmental and senescence. Since transcript levels are not necessarily indicative of the levels of the corresponding proteins, we have conducted a comparative proteomics analysis of the senescence syndrome in Arabidopsis leaves during age-dependent and dark-induced, detached leaf senescence. Protein separation by 2D gel electrophoresis of leaf extracts from different progressive senescence stages followed by MS analysis showed several temporal expression profiles of differentially expressed proteins. The identified senescence-upregulated proteins fall into several functional categories: metabolism, respiration, stress responses, cell wall degradation etc. Comparison between the pattern changes of the senescence-associated proteins and the corresponding mRNAs indicated only partial correlation. These results suggest several regulation levels such as transcription, post transcription, translation and post-translation as well as different turnover of individual proteins. Finally, we analyzed the changes in global protein modification patterns as a result of oxidation. We present a subset of oxidized proteins appeared during the course of leaf development and senescence. The thesis presents a data set of identified upregulated genes and proteins as well as oxidized proteins during leaf senescence. This data may better indicate candidates for reverse genetic manipulation that will enable the production of plants with extended shelf life and delayed leaf senescence.