|M.Sc Thesis||Department of Biology|
|Supervisor:||Prof. Schuster Gadi|
|Full Thesis text|
mRNA degradation in E. coli starts with an endonucleolitic cleavage of the trarget molecule by the enzyme Ribonuclease E ( RNase E ). Homologous genes of RNase E were detected in many prokaryotes and in the nuclear genome of higher plants.
The evolutionary origin of the chloroplast was an endosymbiotic event of photosynthetic bacteria and the pro-eukaryotic cell. During evolution, many genes were transferred from the chloroplast genome to the nucleus, and acquired an N-terminal extension that serves as a transit peptide, which targets the proteins to the chloroplast. The prokaryotic origin of RNase E led us to hypothesize that RNase E homologous in higher plants participates in mRNA degradation in the chloroplast. This key player in mRNA degradation has not yet been characterized in higher plants.
By manufacturing transgenic tobacco (Nicotiana tabacum) that expressese tomato (Lycopersicon esculentum) or Arabidopsis (Arabidopsis thaliana) RNase E, or parts of its different domains, we intend to study the influence of RNase E on the Tobbaco plant.
The transgenic Tobacco plants would enabled us to investigate in vivo the effect of over-expessing RNase E or one of its different domains: The non catalytic domain, deletion of the non catalytic domain, deletion of the transit peptide and inactive RNase E.
Tomato and Arabidopsis RNase E and different domains of the enzymes were cloned into the plant’s expression vector pPZP111. Transformations were performed to introduce the gene and produce the recombinant proteins in tobacco plants. Only the Arabidopsis non catalytic domain was expressed in the tobacco plants.
The transgenic plants were significantly smaller than wild type plants.