Biomaterials, Biodegradables and Biomimetics Research Group

Conference Abstract -ISI Web of Science Indexed

Genetic analysis of the double resistance phenotype to β(1,3)D-glucan synthase from the Schizosaccharomyces pombe mutant pbr1-8


The cell wall can be considered a specific target in the development of new antifungal agents. The biosynthesis of (1,3)β-D-glucan, one of the structural elements of the fungal cell wall, represents a valuable model for morfogenetic studies. We have identified four bgs (beta glucan synthesis) genes that probably are the catalytic subunits of four different (1,3)β-D-glucan synthase activities in S. pombe. bgs4+ was cloned from a cosmid clone located in the same region where we described two genetically linked mutant genes, both related with the
(1,3)β-D-glucan synthesis: cwg1-1, a termosensitive mutant affected in the (1,3)β-D-glucan synthase catalytic subunit and pbr1-8, implicated in the resistance to antifungal drugs that specifically inhibit the (1,3)β-D-glucan synthase activity.
We found that cwg1+ and bgs4+ are the same gene. On the other hand, bgs4does not suppress the recessive phenotype of a pbr1–8 mutant. By the contrary, a bgs4D/pbr1–8 diploid, containing a plasmid expressing bgs4+ under the control of the thiamine-repressible nmt1+ promoter 81X, is resistant to the antifungals when the promoter is repressed and sensitive when it is induced. Besides, bgs4+ cloned from a pbr1–8 mutant, induces a resistant phenotype in a bgs4D but not in a bgs4+ strain. Therefore, bgs4+ is at least partially responsible for the resistant phenotype of pbr1–8 mutant. These results suggest that it is possible the existence of a second gene, very tightly linked to bgs4+, that would be responsible for an additional resistance phenotype. We performed a bgs4+ deletion analysis from a pbr1–8 strain containing the thiamine-repressible plasmid p81X-bgs4+, to maintain the deletion strain viable. Deletion of the complete bgs4pbr1-8 ORF suppressed the mutant phenotype. However, deletions of (1) 1 Kb promoter region plus ATG codon, (2) 200 bases of 5 end ORF, or (3) 1 Kb of 3 end ORF showed, in the three cases, that the truncated Bgs4p were not functional but the resulting bgs4pbr1-8 strains displayed the resistance phenotype. We are currently performing new bgs4+ partial deletions and complementation tests. These and new results will be discussed.

European Journal of Biochemistry - The FEBS Journal
pbr1-8, Schizosaccharomyces pombe
Restricted Access
Peer Reviewed
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