CLONING AND DISRUPTION OF THE ORNITHINE DECARBOXYLASE GENE OF USTILAGO-MAYDIS - EVIDENCE FOR A ROLE OF POLYAMINES IN ITS DIMORPHIC TRANSITION

The gene encoding ornithine decarboxylase (ODC) from Ustilago maydis w as cloned. A conserved PCR product amplified from U. maydis DNA was sy nthesized and used to screen a genomic library of the fungus. Alignmen t of its deduced protein sequence with those of other cloned ODCs show ed a high degree of homology. Gene replacement was obtained by removal of a central part of the gene and insertion of the hygromycin resista nce cassette. The null mutant thus obtained displayed no ODC activity and behaved as a polyamine auxotroph. This result is evidence that a s ingle ODC gene exists in the fungus, and that U. maydis utilizes the O DC pathway as the only mechanism for polyamine biosynthesis. When grow n in polyamine-containing media, the null mutant accumulated a polyami ne pool which further sustained its normal rate of growth in polyamine -free media for approximately 12-16 h. When putrescine concentrations lower than 0-5 mM were employed, the mutant grew at a normal rate but was unable to engage In the dimorphic transition. Under conditions fav ourable for mycelial growth, the mutant grew with a yeast-like morphol ogy in liquid media, and formed smooth colonies consisting of yeast ce lls on solid media. Reversion to normal dimorphic phenotype required h igh concentrations of putrescine or spermidine. These results are evid ence that concentrations of polyamines higher than those necessary to sustain vegetative growth are required for the dimorphic transition in U. maydis.