Date of Award
Doctor of Philosophy (PhD)
Chlamydomonas acidophila is a unicellular green alga of the order Chlamydomonadales. Our research efforts were allied along two lines: (1) Characterization of the C. acidophila mitochondrial genome (mtDNA) and (2) Elucidation of any molecular events responsible for C. acidophila's heavy metal tolerance. The mitochondrial genomes of the protists have been underrepresented in the sequence databases. Among the protists, the alga genera Chlamydomonas shows a reduced mtDNA content with a highly rearranged gene structure. It was decided to sequence C. acidophila's mtDNA to further elucidate the evolutionary paths among the Chlamydomonads and add to the protist sequence database. A 15 kb fragment of C. acidophila's mtDNA was cloned and sequenced. The genes identified included apocytochrome b; partial sequences of subunits 2 and 5 and a complete subunit 1 of the NADH dehydrogenase complex; subunit 1 of the cytochrome oxidase complex; discontinuous and scrambled large and small subunit ribosomal rRNA; and four tRNAs whose anticodons specify tryptophan, glutamine, and 2 methionines (one of which appears to be a pseudogene). The mtDNA of C. acidophila, therefore, probably encodes a reduced gene coding capacity common among the Chlamydomonadales. In fact the basic gene order is colinear with that of C. eugametos. However, C. acidophila appears to have two distinctive features: (1) The reduced size of intergenic spacers, and (2) Non-synonymous insertion of a number of group I introns within the partial sequence. These differences suggest a recent divergence between C. acidophila and C. eugametos, and place them very close phylogenetically. It was also noticed that C. acidophila exhibits a higher tolerance for cadmium than do other Chlamydomonas species. Cadmium is a potent environmental toxin and carcinogen that is accumulating in the environment through anthropogenic and natural means. Knowledge of the characteristics of metal tolerant species has yielded valuable insights into the nature of cadmium tolerance, and may one day aid in the safe disposal of this metal. In an attempt to understand the role of mtDNA during cadmium exposure, a 5 kb Hind III fragment of mtDNA was cloned onto a pGem vector (pJB2). That fragment was hybridized to Northern blots of cadmium challenged C. acidophila cells, and a transcript of ∼300 bp in size was shown to increase during cadmium challenge. Restriction studies and DNA sequencing has revealed that the transcript was produced from a 1500 bp region and appears to be rRNA.
Hoffman, Scott A., "The DNA Sequence From A Cloned 15 Kilobase Fragment Of The Chlamydomonas Acidophila Mitochondrial Genome And RNA Transcript Production In Response To Cadmium" (1999). Theses and Dissertations. 759.