Summary: The gene from encodes an inducible alkaline extracellular protease. Its complex regulation involves pH, carbon, nitrogen and peptones. Two previously identified upstream activating sequence (UAS) regions were analysed in a reporter system, outside the context. Fragments from the UAS regions were inserted upstream of a minimal promoter directing the expression of a reporter gene. The activity of the hybrid promoters was assessed following integration into the genome. This study confirmed the presence of two UASs composed of several interacting elements. Within the distal UAS (UAS1), a TUF/RAP1 binding site exhibited a UAS activity, which was enhanced by the presence of two adjacent repeats, overlapping sites similar to the upstream repressing sequence from Within the proximal UAS (UAS2), the UAS activity required the interaction of both an ABF1-like binding site and a decameric repeat, containing PacC site consensus sequences. This decameric repeat was able to mediate repression due to carbon and/or nitrogen sources as well as pH-dependent activation. A study in the context of -regulatory mutations in the gene showed that the PacC-like sites, potential binding sites for YIRim101p, were implicated in the derepression of UAS2-driven expression at neutral-alkaline pH. The response of the PacC-like decamers to external pH was dependent on the status of the pH-regulated activator YIRim101p, which is homologous to the PacC regulator. The carbon/nitrogen regulation imposed on the decamers was shown to be independent of YIRim101p and to override its effects.

Keyword(s): promoter , regulation , UAS , XPR2 and Yarrowia

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