FIG. 1.

Regulation of proline utilization in A. nidulans. (A) Overview of proline cluster regulation. The prnD-B intergenic region is shown. prnD encodes the proline oxidase, and prnB encodes the specific proline transporter (12, 14). The pathway-specific transcription factor PrnA is essential for proline induction of both genes. In the absence of preferential carbon (glucose) and nitrogen (ammonium) sources and the presence of proline, PrnA and the GATA factor AreA bind to their cognate sites in the intergenic region, resulting in the expression of prnD and prnB. Repression requires both glucose activation of the negative regulator CreA and ammonium inactivation of AreA. Full repression occurs only in the simultaneous presence of glucose and ammonium. Repression acts directly on prnB expression; prnD repression is indirect and results from inducer exclusion. (B) Effect of adaB and gcnE deletion on prnB and prnD transcription. Strains (Table 1) were pregrown in liquid minimal medium under noninducing conditions (5 mM urea-0.1% fructose) with the appropriate supplements, harvested, divided into aliquots, and further incubated for 2 h under the conditions indicated. Noninducing (NI), 5 mM urea and 0.1% fructose; inducing (I), induced by 20 mM proline; inducing-repressing (IR), 20 mM proline and repression by 1% glucose and 20 mM ammonium-l(+)-tartrate. Expression levels (bottom) were quantified using phosphorimaging and ImageQuant software analysis. Normalized signals were obtained by comparison of specific signals with actin gene (acnA) expression signals. The induced levels in the adaB⁺ and gcnE⁺ strains are given in every case the arbitrary value of 100; filled columns represent prnB, and open columns represent prnD expression.