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Isolation and characterization of high chlorophyll fluorescence mutants of Arabidopsis thaliana

University of British Columbia

Photosynthetic electron transport mutants of Arabidopsis thaflana were isolated using the high chlorophyll fluorescence (hcf) phenotype as a screen. Seeds from small bulk populations (7 plants each) of chemically mutagenized (ethyl methane sulfonate) plants were collected and the progeny screened for the presence of hcf phenotypes. Of 570 bulk populations screened, 251 contained hcf seedlings in the M2 generation. This suggests that there are a high number of nuclear loci that are required for functional photosynthetic electron transport. Twenty-one mutant lines, each originating from a separate M1 bulk segregating for the hcf phenotype as a single nuclear recessive gene were isolated. Eight lines that appeared to be blocked early in the photosynthetic electron transport chain were selected for further study. They were characterized with respect to fluorescence induction kinetics, chloroplast pigment composition, photosystem I and H electron transport activity rates, chloroplast proteins, chlorophyll-protein complexes, and RNAs. Chloroplast pigments were found in different proportions in the mutants compared to the wild-type siblings, however, all the major pigments were present. Thus pigment biosynthesis does not appear to be the cause of the hcf phenotype, but loss of one or more of the thylakoid membrane complexes involved in electron transport is responsible. Four of the mutants (hcf2, hct3, hcf5 and hcf6) are reduced in all components of the electron transport chain (photosystems I and II, and the cytochrome complex). The block in electron transport in these mutants can be correlated with abnormal processing and/or stability of chioroplast RNA transcripts or steady state levels of chloroplast RNAS encoding protein products components of photosystem II and cytochrome complex. In mutant hcf2, the chloroplastpetA transcript (encoding the cytochrome fapoprotein) is at significantly higher steady state levels in the mutant. The reason for this is unknown at the present, as the cytochrome fpolypeptide is in the thylakoid membrane at reduced levels. Mutant hcf3 appears to have a lower level of chloroplast mRNAs, but the differences were not quantified. Mutants hcf5 and hcf6 both have altered levels in some, but not all, of the transcripts derived from polycistronic chioroplast operons. The results on these mutants suggests that a nuclear gene product involved in the stability or processing of the chioroplast transcripts is missing in these mutants. The hcf phenotype in the remainder of the mutants (hcfl, hcf4, hc17 and hct) is due to a block in electron transport at the photosystem I complex. Photosystem II and the cytochrome complex are normal, except for mutant hcf4 which appears to have a slightly lower level of Photosystem II electron transport activity rate. There was a correlation between photosystem I ChI protein complex (CPI), photosystem I thylakoid membrane polypeptides and photosynthetic electron transport activity rate for mutants hcfl and hcf8. Mutant hcf4 on the other hand, in spite of the lower level of photosystem I ChI-protein complex (CPI) and photosystem I polypeptides, had a normal level of photosystem electron transport activity rate. The reason for this is unknown, although it is possible that a soluble component in the electron transport chain is lost in the mutant. Chloroplast and nuclear steady state mRNA levels are at wild-type levels in all of the mutants reduced in photosystem I. This suggests that a nuclear gene product is required post-transcriptionally in assembly and/or stability of the photosystem I complex in these mutants.

Thesis/Dissertation

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2008-12-20

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http://hdl.handle.net/2429/3269

Botany

University of British Columbia

UBCV

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