Microtubules and actin cytoskeleton in Cryptococcus neoformans compared with ascomycetous budding and fission yeasts

Actin cytoskeleton and microtubules were studied in a human fungal pathogen , the basidiomycetous yeast Cryptococcus neoformans (haploid phase of Filob asidiella neoformans), during its asexual reproduction by budding using flu orescence and electron microscopy, Staining with rhodamine-conjugated phall oidin revealed an F-actin cytoskeleton consisting of cortical patches, cabl es and cytokinetic ring. F-actin patches accumulated at the regions of cell wall growth, i.e, in sterigma, bud and septum, In mother cells evenly dist ributed F-actin patches were joined to F-actin cables, which were directed to the growing sterigma and bud. Some F-actin cables were associated with t he cell nucleus. The F-actin cytokinetic ring was located in the bud neck, where the septum originated. Anti-tubulin TAT1 antibody revealed a microtub ular cytoskeleton consisting of cytoplasmic and spindle microtubules, In in terphase cells cytoplasmic microtubules pointed to the growing sterigma and bud. As the nucleus was translocated to the bud for mitosis, the cytoplasm ic microtubules disassembled and were replaced by a short intranuclear spin dle. Astral microtubules then emanated from the spindle poles. Elongation o f the mitotic spindle from bud to mother cell preceded nuclear division, fo llowed by cytokinesis (septum formation in the bud neck). Electron microsco py of ultrathin sections of chemically fixed and freeze-substituted cells r evealed filamentous bundles directed to the cell cortex. The bundles corres ponded in width to the actin microfilament cables. At the bud neck numerous ribosomes accumulated before septum synthesis. We conclude: (i) the topolo gy of F-actin patches, cables and rings in C, neoformans resembles ascomyce tous budding yeast Saccharomyces, while the arrangement of interphase and m itotic microtubules resembles ascomycetous fission yeast Schizosaccharomyce s, The organization of the cytoskeleton of the mitotic nucleus, however, is characteristic of basidiomycetous yeasts. (ii) A specific feature of C, ne oformans was the formation of a cylindrical sterigma, characterized by inva sion of F-actin cables and microtubules, followed by accumulation of F-acti n patches around its terminal region resulting in development of an isodiam etrical bud.