Cloning and analysis of mold-specific genes in the dimorphic fungus Histoplasma capsulatum

A critical feature in the pathogenesis of the respiratory pathogen Histopla sma capsulatum is the conversion from the mold form (found in soil) to the yeast form in the lungs of the host. Little is known about the molecular bi ology of Histoplasma dimorphism. In particular, the possible roles of genes which are transcriptionally silent in yeast (i.e. mold-specific) have not been studied. We have produced a cDNA library highly enriched for mold-upre gulated clones by fragmenting cDNA and removing yeast-specific and common s equences with a highly efficient enzyme degrading subtraction method. Scree ning of randomly selected clones identified cDNA fragments representing 16 different mold-upregulated genes. Because multiple cDNA fragments can be tr eated as alleles in a genetic screen, we were able to apply probability ana lysis to estimate the total number of mold-upregulated genes. We estimate t hat there are 27 upregulated genes, cDNA fragments of 16 have been isolated . Here we report the first isolation and analysis of cDNA from two mold-spe cific genes, MS8 (GenBank AF292398) and MS88 (GenBank AF357882). The MS8 tr anscript was very strongly expressed in mold but not detected on Northern b lots with yeast RNA. The putative MS8 protein was predicted to be 21.3 kDa (203 aa), very rich in glutamine and glycine and had a calculated pI of 6.7 6. The MS88 transcript was weakly expressed in mold and not detected in yea st. The putative MS88 protein was predicted to be 22.5 kDa (219 aa) with a pI of 4.46. GenBank similarity searches revealed that the putative MS8 prot ein was similar to a glutamine-rich protein, of unknown function, from the fungus Colletotrichum gloeosporioides (GenBank U94186). No significant matc hes were found for the putative MS88 protein. (C) 2001 Elsevier Science B.V . All rights reserved.