title: In Vivo Competition between Plastocyanin and a Copper-Dependent Regulator of the Chlamydomonas reinhardtii Cytochrome c(6) Gene.
abstract: The Chlamydomonas reinhardtii gene encoding cytochrome c(6) (Cyt c(6)) is transcriptionally repressed by cupric ions. In quantitating the level of expression of this gene as a function of cupric ions available per cell, we find that transformed Chlamydomonas reinhardtii cells that accumulate high levels of plastocyanin (a type I copper protein) have a higher sensory threshold for copper-dependent repression of the Cyt c(6) gene than do untransformed, otherwise isogenic, cells that are plastocyanin-deficient. Also, in wild-type cells, the extent to which the gene is expressed at any given ratio of copper/cell is exactly correlated with the predicted deficiency (at this level of copper) in the organism's capacity to synthesize holoplastocyanin. These results support a simple model in which the sensory threshold for transcriptional repression of the Cyt c(6) gene is determined by direct competition for intracellular copper ions between a copper-binding regulator of this gene and plastocyanin. Thus, the organism is able to maintain a constant amount of Cyt c(6) plus plastocyanin per Photosystem I. With the use of in vitro-generated Cyt c(6)-encoding transcripts as a standard for the quantitation of cellular Cyt c(6) mRNA levels, we estimate that whereas copper-deficient wild-type cells maintain approximately 1 x 10(2) to 4 x 10(2) Cyt c(6)-specific transcripts per cell, copper-supplemented cells contain, on average, less than one Cyt c(6)-encoding mRNA. Thus, repression of the Cyt c(6) gene by copper ions is essentially 100%, making it unlikely that Cyt c(6) has any essential metabolic function in copper-supplemented cells. We find also that the steady-state levels of several transcripts, including those for Cyt c(6), are influenced by cell density, so that cells harvested at low density contain several-fold as many copies of a particular message as cells harvested near stationary phase.
authors.affiliation: Department of Chemistry and Biochemistry, University of California at Los Angeles, Los Angeles, California 90024.
authors.forename: K L
authors.initials: KL
authors.lastname: Hill
authors.forename: S
authors.initials: S
authors.lastname: Merchant