A Transcription Factor Cascade Involving Fep1 and the CCAAT-Binding Factor Php4 Regulates Gene Expression in Response to Iron Deficiency in the Fission Yeast Schizosaccharomyces pombe

Abstract

ABSTRACT We have identified genes encoding candidate proteins involved in iron storage ( pcl1 + ), the tricarboxylic acid cycle ( sdh4 + ), and iron-sulfur cluster assembly ( isa1 + ) that are negatively regulated in response to iron deprivation. Promoter deletion and site-directed mutagenesis permitted identification of a new cis -regulatory element in the promoter region of the pcl1 + gene. This cis -acting regulatory sequence containing the pentanucleotide sequence CCAAT is responsible for transcriptional repression of pcl1 + under low iron supply conditions. In Schizosaccharomyces pombe , the CCAAT-binding factor is a heteromeric DNA-binding complex that contains three subunits, designated Php2, Php3, and Php5. Inactivation of the php2 + locus negatively affects the transcriptional competency of pcl1 + . A fourth subunit, designated Php4, is not essential for the transcriptional activation of target genes under basal and iron-replete conditions. We demonstrate that, in response to iron-limiting conditions, Php4 is required for down-regulation of pcl1 + , sdh4 + , and isa1 + mRNA levels. In vivo RNase protection studies reveal that the expression of php4 + is negatively regulated by iron and that this regulated expression requires a functional fep1 + gene. The results of these studies reveal that Fep1 represses php4 + expression in response to iron. In contrast, when iron is scarce, Fep1 becomes inactive and php4 + is expressed to act as a regulatory subunit of the CCAAT-binding factor that is required to block pcl1 + , sdh4 + , and isa1 + gene transcription.