Fate of transforming DNA in pathogenic fungi.
Genetic engineering is an important tool in helping us to define the molecular basis of pathogenicity and is also useful in helping us to identify new therapeutic targets in pathogenic fungi. Molecular genetic manipulation of micro-organisms requires the development of plasmid-mediated transformation systems that include: (i) infusion of exogenous DNA into recipient cells, (ii) expression of genes present on the incoming DNA, and (iii) stable maintenance and replication of the inserted DNA leading to expression of the desired phenotypic trait. Transformation systems have been developed for only a handful of fungi that are pathogenic to humans including several species of Candida, Cryptococcus neoformans, Histoplasma capsulatum, Blastomyces derrmatitidis, Aspergillus fumigatus, Wangiella dermatitidis (Exophiala dermatitidis) and Coccidioides immitis. Except for Candida species and A. fumigatus, where passage of exogenous DNA into recipient cells has been achieved readily using methods developed for transformation of Saccharomyces cerevisiae and Aspergillus nidulans, respectively, development of transformation systems in other pathogenic fungi has been delayed considerably and has only been possible recently with the introduction of electroporation and biolistic methods. Conventional spheroplasting methods or cell wall permeabilization methods using lithium acetate have not been successful for transformation of C. neoformans and work with only low efficiency in H. capsulatum. The fate of incoming DNA varies greatly in these pathogenic species regardless of their phylogenetic relationships. Understanding the fate of incoming DNA is critical for the construction of transforming vectors and the molecular manipulation of the organisms. In this symposium, recent advances in molecular genetic systems including transformation systems, the fate of incoming DNA and strategies for targeted integration are discussed in relation to four pathogenic fungi.
Kwon-Chung
K J
KJ
National Institutes of Health, Bethesda, Maryland 20892, USA.
Goldman
W E
WE
Klein
B
B
Szaniszlo
P J
PJ
D004271:DNA, Fungal / Q000235:genetics*; D005658:Fungi / Q000235:genetics*; D006801:Humans; D009181:Mycoses / Q000382:microbiology*; D014170:Transformation, Genetic