Multiple sense and antisense promoters contribute to the regulated expression of the isc-suf operon for iron-sulfur cluster assembly in Rhodobacter

Abstract

A multitude of biological functions relies on iron-sulfur clusters. The formation of photosynthetic complexes goes along with an additional demand for iron-sulfur clusters for bacteriochlorophyll synthesis and photosynthetic electron transport. However, photooxidative stress leads to the destruction of iron-sulfur clusters, and the released iron promotes the formation of further reactive oxygen species. A balanced regulation of iron-sulfur cluster synthesis is required to guarantee the supply of this cofactor, on the one hand, but also to limit stress, on the other hand. The phototrophic alpha-proteobacterium Rhodobacter sphaeroides harbors a large operon for iron-sulfur cluster assembly comprising the iscRS and suf genes. IscR (iron-sulfur cluster regulator) is an iron-dependent regulator of isc-suf genes and other genes with a role in iron metabolism. We applied reporter gene fusions to identify promoters of the isc-suf operon and studied their activity alone or in combination under different conditions. Gel-retardation assays showed the binding of regulatory proteins to individual promoters. Our results demonstrated that several promoters in a sense and antisense direction influenced isc-suf expression and the binding of the IscR, Irr, and OxyR regulatory proteins to individual promoters. These findings demonstrated a complex regulatory network of several promoters and regulatory proteins that helped to adjust iron-sulfur cluster assembly to changing conditions in Rhodobacter sphaeroides.