The Regulatory Mechanism Of Sigma Factors During Heterocyst Differentiation In Anabaena Sp. PCC 7120

Anabaena sp. strain PCC 7120 is a kind of diazotrophic and filamentous cyanobacterium. Upon nitrogen-depletion, part of cells will differentiate into heterocysts, specialized cells capable of nitrogen fixation. The distribution of heterocysts in filaments has a semi-regular pattern, with about 10-20 vegetative cells being intercalated between every two heterocyst.As an integral subunit of prokaryotic RNA polymerase, sigma factor promotes the specific recognition and binding of the enzyme to the target DNA strand. Twelve sigma factor are identified in Anabaena sp. strain PCC 7120, and they are conventionally divided into three groups:group 1 consists of SigA, group 2 consists of SigB, SigC, SigD and SigE, and group 3 consists of SigF, SigG, SigI and SigJ. Previous studies suggest that SigC and SigG may be involved in the regulation of heterocyst differentiation as both sigC and sigG had increased expression in nitrogen starvation conditions.To investigate whether SigC and SigG are participated in heterocyst development regulation, we constructed the sigC and sigG deletion mutants, named as MsigC and MsigG, respectively. After nitrogen deprivation, the heterocyst frequency of MsigC or MsigG appeared lower than of the wild type. After 24 h and 48 h of nitrogen stavation, the average vegetative cell numbers between neighboring heterocysts were 20.6 and 18.4 in MsigC,22.6 and 19.9 in MsigG, and 16 and 12 in the wild type. The decreased heterocyst differentiation frequencies in MsigC or MsigG suggest that SigC and SigG play a role in heterocyst development.We further carried out complementation experiments by expressing sigC and sigG in their corresponding mutant, resulting in CsigC and CsigG, respectively. After 24 h and 48 h of nitrogen stavation, the average vegetative cell numbers between neighboring heterocysts were reduced to 18.1 and 16.1 in CsigC, and to 18.9 and 17.2 in CsigG. The expression of SigC and SigG increased the heterocyst frequency of MsigC and MsigG, further indicates that SigC and SigG are involved in heterocyst development regulation.To better understanding the underlying mechanism by which SigC and SigG regulate heterocyst differentiation, we examined 11 well-known heterocyst-formation related genes in MsigC and MsigG using transcriptional GFP fusions and Real-Time PCR. The results showed that the expression of both ntcA and hetR were down regulated in either MsigC or MsigG, while the expression of hglD was completely suppressed in MsigG These results suggest that sigC and sigG may affect heterocyst frequency by regulating the expression of heterocyst-formation related genes.