Two chitin synthase genes from Ustiago maydis Beat

Departamentos de Genetica y Biolog(a Molecular1 and Ingenieria Genetica (Unidad Irapuato)2, Centro de lnvestigacidn y de Estudios Avanzados del IPN, Irapuato, Gto, Mexico PCR was used to amplify fragments corresponding to CHS genes from Ustilago maydis, utilizing as primers oligonucleotides devised according to the conserved regions of fungal CHS genes. The PCR product was employed as a probe to screen a genomic library of the fungus. Two different CHS genes (Umchsl and Umhs2) were thus identified in the positive clones recovered. Their sequence revealed high similarity with the CHS genes previously cloned from other fungi, especially in their central region. Alignment with the deduced protein sequences of all CHS genes reported up to date showed the existence of seven conserved domains. Transcripts from both genes were detected in the yeast and mycelial forms. In general, the transcripts from the Umchsl gene appeared to be present at a higher level than the transcripts from the Umchs2 gene; the transcripts from both genes appeared to be more abundant in the mycelial form. Gene replacement of either gene and analysis of the resulting phenotype demonstrated that they are non-essential. Nevertheless, growth, chitin synthase activity levels, and chitin content of mycelial cells induced by cultivation in acidic media were all reduced in chsl and chs2 mutants. However, mating, virulence and dimorphic behaviour were unaffected. Overall, the results indicate that the CHSf and CHS2 genes encode products with redundant functions in U. maydis.