The orlA gene from Aspergillus nidulans encodes a trehalose‐6‐phosphate phosphatase necessary for normal growth and chitin synthesis at elevated temperatures

Abstract

A cosmid carrying the orlA gene from Aspergillus nidulans was identified by complementation of an orlA1 mutant strain with DNA from the pKBY2 cosmid library. An orlA1 complementing fragment from the cosmid was sequenced. orlA encodes a predicted polypeptide of 227 amino acids (26360 Da) that is homologous to a 211-amino-acid domain from the polypeptide encoded by the Saccharomyces cerevisiae TPS2 gene and to almost the entire Escherichia coli otsB-encoded polypeptide. TPS2 and otsB each specify a trehalose-6-phosphate phosphatase, an enzyme that is necessary for trehalose synthesis. orlA disruptants accumulate trehalose-6-phosphate and have reduced trehalose-6-phosphatate phosphatase levels, indicating that the gene encodes a trehalose-6-phosphatate phosphatase. Disruptants have a nearly-wild-type morphology at 32 degrees C. When germinated at 42 degrees C, the conidia and hyphae from disruptants are chitin deficient, swell excessively, and lyse. The lysis is almost completely remedied by osmotic stabilizers and is partially remedied by N-acetylglucosamine (GlcNAc). The activity of glutamine:fructose-6-phosphate amido-transferase (GFAT), the first enzyme unique to aminosugar synthesis, is reduced and is labile in orlA disruption strains. The findings are consistent with the hypothesis that trehalose-6-phosphate reduces the temperature stability of GFAT and other enzymes of chitin metabolism at elevated temperatures. The results extend to filamentous organisms the observation that mutations in fungal trehalose synthesis are highly pleiotropic and affect aspects of carbohydrate metabolism that are not directly related to trehalose synthesis.

The Filamentous Fungus Aspergillus niger Contains Two “Differentially Regulated” Trehalose-6-phosphate Synthase-encoding Genes, tpsA and tpsB

“…A comparison of the amino acid sequences of the tpsA and tpsB gene products does not provide a clue as to the reason for this. Interestingly, while this paper was prepared for publication, Borgia et al (43) reported on the presence of two trehalose-6-phosphate phosphatases in A. nidulans but did not investigate whether the two gene products may have different functions. It will be interesting to see if this is a general feature of multicellular fungi or a feature unique to aspergilli.…”

Characterization and Regulation of the Trehalose Synthesis Pathway and Its Importance in the Pathogenicity of Cryptococcus neoformans

“…In C. albicans, the tps2 mutant flocculates during stationary growth, while in A. nidulans, the mutant produces nonviable germination of spores through a deficiency in chitin production. Furthermore, both of these phenotypes can be suppressed by the addition of osmoprotectants to the culture medium (7,53,59). We did not observe a flocculation phenotype in a C. neoformans-like tps2 strain of C. albicans, but we did find different phenotypes between tps1 and tps2 mutants at low and high temperatures.…”

“…T6P is an intermediate in trehalose biosynthesis that has been shown to possess regulatory potential (5,26 (7,21,22,47,53,59) and give further insights into T6P. In C. albicans, the tps2 mutant flocculates during stationary growth, while in A. nidulans, the mutant produces nonviable germination of spores through a deficiency in chitin production.…”

The SskA and SrrA Response Regulators Are Implicated in Oxidative Stress Responses of Hyphae and Asexual Spores in the Phosphorelay Signaling Network ofAspergillus nidulans

“…7). 27,[43][44][45] It was found that conidia from ÁsskA contained a significantly lower amount of transcript for each of these stress-tolerant or germination-associated genes. These results together support the view that SskA plays more sophisticated or wider roles in stress-tolerant spore development (Figs.…”