Identification of a Novel Gene That Plays a Role in High Light Tolerance in the Green Micro-alga Chlamydomonas reinhardtii

Kevin Manoap-Anh-Khoa Nguyen, Ja'Von Swint, Joel Page III, Kenneth Kim, Katherine Smith, Tai Truong, Kasey Swilley

Abstract


Photo-autotrophic growth under different light intensities is regulated by a complex interplay of several physiological processes. Chlamydomonas reinhardtii is a model green micro-alga. A haploid genome, short replication time, autotrophic and heterotrophic growth ability, amenability to nuclear and chloroplast transformation, and a fully sequenced genome, make it a model system for studying oxygenic photosynthesis. Our lab generated a mutant library of Chlamydomonas by random insertional mutagenesis using the pBC1 vector. This vector contains the APHVIII gene that confers resistance to paromomycin. The mutant library was screened under heterotrophic, mixotrophic, and photo-autotrophic growth conditions under different light intensities leading to the isolation of differentially light-sensitive 20 mutants. One of the isolated high light-sensitive mutant is 10E35/lsr1a. Mutant lsr1a is chlorophyll-deficient, hyper-sensitive to high light in photo-autotrophic and in mixotrophic growth conditions and photo-bleaches on exposure to high light. lsr1a has a slow growth rate in photo-autotrophic condition compared to that of the wild type strain. lsr1a fails to grow photo-autotrophically under dim light in the presence of Rose Bengal, a singlet oxygen generator. Adapter Ligation-Mediated-PCR was performed on lsr1a genomic DNA. DNA sequencing of the PCR products revealed two insertion sites of pBC1 in lsr1a. One pBC1 insertion site is in the fourth exon of a novel functionally uncharacterized gene, namely Cre11.g467757 (LSR1). The second insertion site of pBC1 is in Cre02.g095095. Cre02.g095095 codes for a secretory cell wall protein pherophorin-C12 (PHC12) of unknown function. A strong indication that Cre11.g467757 is responsible for the lsr1a growth phenotype is the observation that its growth phenotype is similar to that of another uncharacterized Chlamydomonas Library Project (CLiP) mutant (lsr1b), which has a mutation in the fourth intron of the gene Cre11.g467757. We will be presenting our physiological and molecular research on lsr1 mutants.


Keywords


Photo-protection; Chlamydomonas reinhardtii; Non-photochemical quenching

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