POLYAMINES, DNA METHYLATION, AND FUNGAL DIFFERENTIATION

Mucorales constitute a group of fungi that, because of their growth ch aracteristics, have been used extensively in the study of cell differe ntiation, cell morphogenesis, and stimuli perception. We have studied the role of polyamine metabolism in the development of different Mucor ales, with emphasis on Mucor and Phycomyces species. It has been obser ved that previous to each differentiative step, the cellular levels of the most regulated enzyme of the pathway, ornithine decarboxylase (OD C), and polyamines suffer a noticeable increase. Addition of diaminobu tanone (DAB), a competitive inhibitor of ODC, blocks all the correspon ding differentiative phenomena. In its presence, germinating spores fa il to produce germ tubes and keep growing isodiametrically; mycelia do not sporulate but continue their vegetative growth, and yeast cells a re unable to engage in a dimorphic transition without alterations in t heir growth rate. This differential effect of the ODC inhibitor in gro wth and development is apparently due to the location of ODC in at lea st two different cell compartments, one of which is impermeable to the drug. Inhibition of development is counteracted by putrescine and mor e noticeably by 5-azacytidine (5AC), a strong inhibitor of DNA methyla tion. Methylation levels of DNA are high in spores, and they become re duced after germination. Demethylation is inhibited by hydroxyurea, wh ich blocks DNA replication, and by DAB. The effect of the latter is re versed by 5AC. These results suggest a relationship between polyamines and DNA methylation. Analysis of metallothioneine gene (CUP) behavior and expression during spore germination has confirmed this hypothesis . The gene is methylated and not transcribed in the spore, and becomes demethylated and transcriptionally active after germination. Both pro cesses, demethylation and transcription, are inhibited by DAB. The hyp othesis has been forwarded that in Mucorales (and perhaps in other fun gi), polyamine levels regulate gene expression during development, aff ecting DNA methylation by an as yet unknown mechanism.