ABSTRACTCarotenoids constitute an important component of the defense system against photooxidative stress in bacteria. InAzospirillum brasilenseSp7, a nonphotosynthetic rhizobacterium, carotenoid synthesis is controlled by a pair of extracytoplasmic function sigma factors (RpoEs) and their cognate zinc-binding anti-sigma factors (ChrRs). Its genome harbors two copies of the gene encoding geranylgeranyl pyrophosphate synthase (CrtE), the first critical step in the carotenoid biosynthetic pathway in bacteria. Inactivation of each of twocrtEparalogs found inA. brasilensecaused reduction in carotenoid content, suggesting their involvement in carotenoid synthesis. However, the effect ofcrtE1deletion was more pronounced than that ofcrtE2deletion. Out of the five paralogs ofrpoHinA. brasilense, overexpression ofrpoH1andrpoH2enhanced carotenoid synthesis. Promoters ofcrtE2andrpoH2were found to be dependent on RpoH2 and RpoE1, respectively. Using a two-plasmid system inEscherichia coli, we have shown that thecrtE2gene ofA. brasilenseSp7 is regulated by two cascades of sigma factors: one consisting of RpoE1and RpoH2 and the other consisting of RpoE2 and RpoH1. In addition, expression ofcrtE1was upregulated indirectly by RpoE1 and RpoE2. This study shows, for the first time in any carotenoid-producing bacterium, that the regulation of carotenoid biosynthetic pathway involves a network of multiple cascades of alternative sigma factors.IMPORTANCECarotenoids play a very important role in coping with photooxidative stress in prokaryotes and eukaryotes. Although extracytoplasmic function (ECF) sigma factors are known to directly regulate the expression of carotenoid biosynthetic genes in bacteria, regulation of carotenoid biosynthesis by one or multiple cascades of sigma factors had not been reported. This study provides the first evidence of the involvement of multiple cascades of sigma factors in the regulation of carotenoid synthesis in any bacterium by showing the regulation of a gene encoding geranylgeranyl pyrophosphate synthase (crtE2) by RpoE1→RpoH2→CrtE2 and RpoE2→RpoH1→CrtE2 cascades inA. brasilense. It also provides an insight into existence of an additional cascade or cascades regulating expression of another paralog ofcrtE.
Engineering the carotenoid biosynthetic pathway in Rhodothermus marinus for lycopene production
