Effects Of S-(3,4-dichlorobenzyl) Isothiourea On Cell Division Related Events In The Cyanobacterium Anabaena Sp. Strain PCC 7120

Anabaena PCC 7120 is a nitrogen-fixing filamentous cyanobacterium. When the combined-nitrogen source is limited in the environment, heterocysts are produced to fix nitrogen from the air. Heterocysts show a semiregular pattern in the filaments, and provide a micro-oxic environment for the functionality of the oxygen-sensitive nitrogenase complex, which catalyzes the reaction of nitrogen fixation.A small molecule compound, S-(3,4-dichlorobenzyl) isothiourea (A22), can inhibit cell growth, induce cell morphology changes, division defects, and irregular chromosome segregation in Escherichia coli and Caulobacter crescentus. A22 has been reported to specifically inhibit the function of MreB, an actin-like protein in rod-shaped bacteria. In this study, the effects of A22 in Anabaena PCC 7120 had been studied. A22 could severely inhibit the cell growth, and even lead to cell disruption. By measuring the growth curves, the MIC of A22 for Anabaena PCC 7120 was determined to be around 2.0μg/ml. We found that in Anabaena PCC 7120, A22 could inhibit cell growth, cause abnormal cellular morphology, and bring about asymmetric cell division and irregular chromosome segregation. However, A22 had little effect on heterocyst formation. These results demonstrated that the phenotype after the treatment of A22 was not the same as that of the mreB-depletion mutant. An A22-resistant mutant named C23 was isolated by growing cells on A22-containing plates. C23 showed stronger resisitance to A22 than wild type. It had normal appearance of cell shape, division, and DNA content distribution when treated by A22. But this mutant kept a wild-type allele of mreB, suggesting that A22 may have different target(s) in the cyanobacterium Anabaena PCC 7120.In the two-component system, two histidine kinase encoding genes, all2899 and all2897, and a response regulator encoding gene, all2898, form a gene cluster on the chromosome. The disruption mutants of all2897 and all2899 can form heterocysts when the combined-nitrogen source is limited in the media, but the interrupted-inactivation mutant of all2898 can not differentiate into heterocysts in response to combined nitrogen deprivation. We confirmed that the components in the gene cluster are co-transcribed as an operon by using RT-PCR. They are up-regulated when the combined-nitrogen source is limited in the medium, indicating that they may mediate the cellular nitrogen metabolism. The overexpression of all2898 had no effect on heterocyst development, and complement strains didn’t restore heterocyst development in the mutant of all2898.