Biosynthesis of carbonic anhydrase in Chlamydomonas reinhardtii during adaptation to low CO 2

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Abstract
The unicellular green alga C. reinhardtii synthesizes anhydrase in response to low levels of CO2 (i.e., air levels of CO2). 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 C by C. reinhardtii and the saturation of ribulose-1,5-bisphosphate carboxylase at low extracellular C concentrations. The synthesis and compartmentalization of carbonic anhydrase in C. reinhardtii was examined. The monomeric species associated with carbonic anhydrase activity is synthesized as a precursor on 80S cytoplasmic ribosomes. This precursor can be deteced 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% CO2 to air for 12 h. It is not synthesized when the in vitro system is primed with poly(A)-RNA from CO2-grown algae. Since translatable RNA for the polypetide response for carbonic anhydrase activity was only present in cells that experienced low levels of CO2, 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. 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. The biosynthesis of this extracellular algal enzyme apparently 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 polypetide into the lumen of the endoplasmic reticulum and subsequent glycosylation events prior to export across the plasmalemma.