Disruption analysis of the gene for a cold-regulated RNA-binding protein, rbpA1, in Anabaena: Cold-induced initiation of the heterocyst differentiation pathway

Abstract

A cold-regulated operon, rbpA1-rpsU, encodes an RNA-binding protein and a ribosomal protein in Anabaena variabilis M3. The level of expression of this gene cluster was about ten times higher at temperatures below 30°C than at 38°C. To study the role of the RbpA1 protein in vivo, we constructed insertional disruptants of rbpA1. These strains were totally devoid of RbpA1 protein but contained a normal level of the ribosomal protein S21, a product of the rpsU gene. The disruptants were morphologically normal at 38°C, but at 22°C they produced unusual cells at a low frequency. These cells were probably at an initial stage of proheterocyst formation. Various molecular events that are related to heterocyst initiation, namely, excision of the 11-kbp DNA element in nifD and the accumulation of transcripts of xisA and hetR, also occurred in the disruptants at 22°C in the presence of nitrate ions, but these events did not occur in the presence of ammonium ions or at 38°C. The results suggest that RbpA1 is required for enhanced repression of heterocyst initiation at low temperatures in the presence of nitrate. Possible mechanisms of the action of RbpA1 are discussed.