Rapid recovery of chloroplast mutations affecting ribulosebisphosphate carboxylase/oxygenase in Chlamydomonas reinhardtii

Abstract

Based on the unique ability of chloroplast genes to recombine in Chlamydomonas reinhardtii, a collection of acetate-requiring mutants was screened for recombination with a mutation affecting ribulose-1,5-bisphosphate carboxylase/oxygenase [Rbu-1,5-P(2) carboxylase/oxygenase; 3-phospho-D-glycerate carboxy-lyase (dimerizing), EC 4.1.1.39]. This chloroplast mutation, rcl-u-1-10-6C, causes the absence of Rbu-1,5-P(2) carboxylase/oxygenase activities and alters the isoelectric point of the larger subunit. Several mutants that displayed little or no recombination with 10-6C were recovered, and two lacked carboxylase activity. These new chloroplast mutants lack both large and small Rbu-1,5-P(2) carboxylase/oxygenase subunits. The approach demonstrated here permits the routine recovery of chloroplast mutations affecting this enzyme. Multiple mutations in the Rbu-1,5-P(2) carboxylase/oxygenase large-subunit gene can be used to investigate the function and regulation of this enzyme and the regulation of chloroplast genes in general.