Screening And Identification Of The Candidate Genes In The Pathways Of Mitochondrial Retrograde Regulation In Plants

Mitochondrial retrograde signaling is an important pathway of communication from mitochondrion to the nucleus. In order to adapt to a variety of stress conditions, mitochondrion will transmit the stress signals to the nucleus by some signaling molecules and signal transduction proteins, then nucleus gene expression will be changed, thus affecting the metabolic activity within the cell to maintain normal life activities. Antimycin A, which is known as an inhibitor of mitochondrial electron respiratory chain, will induce AOX1a expression, AOX1a always acts as a reporter gene in MRR studies, it can be induced to express by majority of stress conditions. Therefore, this study take the mechanism of RTG in yeast and the generation mechanism of alternative respiratory pathway in plants as our main theoretical basis, some T-DNA insertion mutants of the candidate genes, which has a high similarity with RTG regulators, were screened from TAIR and a EMS mutagenesis mutant of Col:LUC in Arabidopsis thaliana was made to identify the genes involved in mitochondrial retrograde regulation. Meanwhile, we use male sterile line (CMS) and its maintain system (MF) from Brassica juncea to study the regulation mechanism of BjAOX1a. The main conclusions are as follows:1GRF3is a negative factor in regulation of MRR in plantsGRF3(General regulatory factor3) has a high homology with Bmhlp, which is a negative regulatory factor in RTG pathways. When treated with50μM AA for3hours, the content of H2O2in grf3was increased and the transcription level of AOX1a was significantly up regulated. When treated with20mM H2O2, AOX1a was rapidly up regulated, the fold-change of transcription level was not as significant as that in AA treatment.The expression trends of NDB2was consistent with that of AOX1a. These results suggest GRF3is a negative regulatory factor in the plant mitochondrial retrograde regulation pathway.2EMS2is a positive regulator in regulation of MRR in plantsems2was obtained by EMS mutagenesis of transgenic Col:LUC, the mutant containing a reporter gene LUCIFERASE (LUC). When treated with AA or H2O2, AOX1a and NDB2 expression was induced less in ems2. We identified EMS2as a positive regulatory factor in the plant mitochondria retrograde regulation pathway.3BjAOX1a is hardly induced by AA treatment in CMS line, BjCDKEl, which is positive regulator of BjAOX1a is also hardly induced by AA treatmentBjAOX1a were induced to express in MF and CMS after AA treatment, but there was a significantly lower expression level of BjAOX1a in CMS than MF. BjCDKE1gene expression levels were lower in CMS than MF in normal conditions. However, BjCDKE1expression was down regulated by AA in MF, while it was down-regulated in the CMS. The mechanism of regulation of BjAOX1a needs further investigated.