YEAST EXO-BETA-GLUCANASES CAN BE USED AS EFFICIENT AND READILY DETECTABLE REPORTER GENES IN SACCHAROMYCES-CEREVISIAE

Yeast exo-1,3-beta-glucanases are secretable proteins whose function i s basically trophic and may also be involved in cell wall glucan hydro lytic processes. Since fluorescein di(beta-D-glucopyranoside) is a flu orogenic substrate detectable and quantifiable by flow cytometry, it w as used for testing the ability of the EXG1 gene product of Saccharomy ces cerevisiae and its homologous gene in Candida albicans to function as reporter genes. These open reading frames were coupled to differen t promoters in multicopy plasmids, and exoglucanase activity quantifie d at flow cytometry. Exoglucanases were found to be useful tools for t he study of promoter regions in S. cerevisiae. This technique has the advantage over other reporter gene systems-such as beta-galactosidase fusions-that it does not require permeabilization of yeast cells and t herefore it allows the recovery of viable cells-by sorting-after flow cytometry analysis.