| Summary: | <i>Cryptococcus neoformans</i> (<i>Cn</i>) is a pathogenic yeast that is the leading cause of fungal meningitis in immunocompromised patients. Various <i>Cn</i> virulence factors, such as the enzyme laccase and its product melanin, phospholipase, and capsular polysaccharide have been identified. During a screen of knockout mutants, the gene resistance to aminocholesterol 1 (<i>RTA1</i>) was identified, the function of which is currently unknown in <i>Cn</i>. Rta1 homologs in <i>S. cerevisiae</i> belong to a lipid-translocating exporter family of fungal proteins with transmembrane regions and confer resistance to the antimicrobial agent 7-aminocholesterol when overexpressed. To determine the role of <i>RTA1</i> in <i>Cn</i>, the knock-out (<i>rta1Δ</i>) and reconstituted (<i>rta1Δ+RTA1</i>) strains were created and phenotypically tested. <i>RTA1</i> was involved in resistance to 7-aminocholesterol, and also in exocyst complex component 3 (Sec6)-mediated secretion of urease, laccase, and the major capsule component, glucuronoxylomannan (GXM), which coincided with significantly smaller capsules in the <i>rta1Δ</i> and <i>rta1Δ+RTA1</i> strains compared to the wild-type H99 strain. Furthermore, <i>RTA1</i> expression was reduced in a secretory 14 mutant (<i>sec14Δ</i>) and increased in an RNAi Sec6 mutant. Transmission electron microscopy demonstrated vesicle accumulation inside the <i>rta1Δ</i> strain, predominantly near the cell membrane. Given that Rta1 is likely to be a transmembrane protein located at the plasma membrane, these data suggest that Rta1 may be involved in both secretion of various fungal virulence factors and resistance to 7-aminocholesterol in <i>Cn</i>.
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