H-037. Regulation of the Alkylresorcinol Biosynthetic Gene arsA in Azotobacter vinelandii

Y. Romero-Ramirez1, D. Segura-Gonzalez1, S. Moreno1, M. Catañeda-Lucio2, G. Espin1;
1Univ. Nacional Autonoma de Mexico, Cuernavaca, Morelos, MEXICO, 2Benemerita Univ. Autonoma de Puebla, Puebla, MEXICO.

A. vinelandii is a bacterium which undergoes a process of differentiation leading to the formation of metabolically dormant cysts. The cyst consists of a contracted cell surrounded by a capsule made up of a thin outer layer (exine) and a thicker inner layer (intine). When this bacterium encysts, 5-alkylresorcinols (AR) lipids unique to encysting cells are synthesized, accumulate in the exine layer, and replace the phospholipids in the membrane. The synthesis of these lipids in A. vinelandii has been shown to occur by the activity of a type III polyketide synthase (ArsB), and the arsABCD operon, involved in AR synthesis, has been described. However, little is know about the regulation of AR synthesis. In this study, we examined the regulation of the expression of the ars genes in A. vinelandii using an arsA-gusA transcriptional fusion reporter and real-time RT-PCR. We found that arsA expression was induced (14 fold) in ageing cultures of vegetative cells, when a small percentage of cyst are formed (less than 0.001 %), whereas a 200 fold induction was observed in cultures induced to encyst, in accordance with the increase observed in alkylresorcinol synthesis. We identified, by Tn5 mutagenesis, a LysR-type transcriptional regulator named ArsR, involved in the activation of arsA transcription. We also found that the global regulators GacA, a two component response regulator, the sigma factors AlgU and RpoS, and the CsrA/CsrB system, implicated in the control of the synthesis of other cysts components (i.e., alginate and poly-ß-hydroxybutyrate), are also involved in the control of alkylresorcinol synthesis. We determined that these regulators exert their effects on arsA through the control of arsR expression.