Regulation of the xylanase‐encoding xlnA gene of Aspergilius tubigensis

Abstract

A gene encoding an endo-1,4-β-xylanase from Aspergillus tubigensis was cloned by oligonucleotide screening using oligonucleotides derived from amino acid sequence data obtained from the purified protein. The isolated gene was functional as it could be expressed in the very closely related fungus Aspergillus niger. The xylanase encoded by this gene is synthesized as a protein of 211 amino acids. After cleavage of the presumed prepropeptide this results in a mature protein of 184 amino acids with a molecular weight of 19 kDa and an isoelectric point of 3.6. The regulatory region of the xlnA gene was studied with respect to the response to xylan induction and carbon catabolite repression. By deletion analysis of the 5′ upstream region of the gene a 158bp region involved in the xylan specific induction was identified. To study this regulatory element a reporter system for transcriptional activating sequences was developed that is based on the A. niger glucose oxidase-encoding gene. From the results with this reporter system it is concluded that this 158bp fragment not only contains the information required for induction of transcription but that it also plays a role in carbon catabolite repression of the xlnA gene. The region directly upstream of this fragment contains four potential CREA target sites; deletion of this region leads to an increase in the level of transcription. These results suggest that carbon catabolite repression of the xinA gene is controlled at two levels, directly by repression of xlnA gene transcription and indirectly by repression of the expression of a transcriptional activator. This type of mechanism would be similar to the double lock mechanism for the regulation of gene expression of alcA in Aspergillus nidulans. The reporter system was also used to study the regulation of expression via the functions located on this fragment in A. niger and in A. niduians. Essentially the same pattern of regulation was found in both of these hosts. Therefore, regulation of xylanase gene expression is basically conserved in ail three aspergilli.