Biosynthesis of carbonic anhydrase in Chlamydomonas reinhardtii during adaptation to low CO(2).
The unicellular green alga Chlamydomonas reinhardtii synthesizes carbonic anhydrase in response to low levels of CO(2) (i.e., air levels of CO(2)). This enzyme, localized predominantly in the periplasmic space of the alga (or associated with the cell wall), is an important component of the machinery required for the active accumulation of inorganic carbon by C. reinhardtii and the saturation of ribulose-1,5-bisphosphate carboxylase at low extracellular carbon concentrations. We have begun to examine the synthesis and compartmentalization of carbonic anhydrase in C. reinhardtii. The monomeric species associated with carbonic anhydrase activity is synthesized as a precursor on 80S cytoplasmic ribosomes. This precursor can be detected immunologically in the profiles of translation products when a reticulocyte lysate, cell-free system is primed with poly(A)-RNA from either air-grown C. reinhardtii or cells shifted from growth on 5% CO(2) to air for 12 hr. It is not synthesized when the in vitro system is primed with poly(A)-RNA from CO(2)-grown algae. Since translatable RNA for the polypeptide responsible for carbonic anhydrase activity was only present in cells that experienced low levels of CO(2), the adaptation process either involves the regulation of transcription of the carbonic anhydrase gene (and perhaps other genes involved in adaptation) or the post-transcriptional processing of the messenger RNA. Furthermore, the appearance of the mature polypeptide associated with carbonic anhydrase activity in the periplasmic space of C. reinhardtii is inhibited by tunicamycin, an antibiotic that prevents core glycosylation of polypeptides on the endoplasmic reticulum. Together, these results suggest that the biosynthesis of this extracellular algal enzyme involves the translation of mRNA for the carbonic anhydrase monomer on ribosomes bound to the endoplasmic reticulum, the cleavage of a signal sequence during transport of the nascent polypeptide into the lumen of the endoplasmic reticulum, and subsequent glycosylation events prior to export across the plasmalemma.