Graduate Thesis Or Dissertation
 

Defining roles for sterols in Saccharomyces cerevisiae : characterization of bulk membrane and high specificity sparking functions

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  • The ability of cholestanol to support growth of yeast sterol auxotrophs has been examined. Growth on this stanol was precluded unless minute quantities of ergosterol were available, a phenomenon designated sparking. The low levels of ergosterol were insufficient to support growth alone. The data indicate that cholestanol is fulfilling a bulk membrane requirement(s) while ergosterol satisfies a high specificity sparking function(s). A variety of sterols and stanols were analyzed for their ability to satisfy these two sterol functions in yeast sterol auxotrophs. While many sterols and stanols satisfied bulk membrane requirements, only those possessing a C-5,6 unsaturation or capable of being desaturated at C-5 in vivo fulfilled the sparking function. Addition of a bromo or iodo moiety to the sterol side chain had no effect on the ability of cholesterol to satisfy either sterol function. However, the presence of a keto or hydroxy group on the sterol side chain completely abolished the ability of cholesterol to satisfy either sterol requirement. Unsaturation of the A-ring or a-saturation of a C-5,6 unsaturation rendered both sterol and stanol unsuitable for either function. The C-28 methyl group of ergosterol, while found not to be required for growth, allowed for greater ease of in vivo desaturation at C-5. As a result some sterols and stanols lacking the C-28 methyl were incapable of satisfying the sparking requirement while identical compounds possessing the C-28 methyl were able to fulfill the sparking function(s). These data are extended to hypothesize a role for the C-28 methyl group as well as other functional moieties of ergosterol in yeast. Physiological studies are also undertaken in an attempt to define the sparking phenomenon better.
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