The beauty of the yeast: Live cell microscopy at the limits of optical resolution

Abstract

The yeast Saccharomyces cerevisiae is a very powerful system for cell biological research. Recent advances in electronic light microscopy together with the application of green fluorescent protein and other in vivo staining techniques have allowed novel and exciting insights into structural organization and dynamics of cells as small as yeast. Methods for staining yeast for microscopic inspection and for introducing tags for localization studies of proteins in living or fixed cells are summarized. Electronic light microscopy, video/deconvolution methods, and confocal laser scanning microscopy as novel tools for structural analyses, and their practical applications in yeast, are discussed. Microsc. Res. Tech. 51:511–529, 2000. © 2000 Wiley‐Liss, Inc.

The Yeast Plasma Membrane Protein Alr1 Controls Mg2+ Homeostasis and Is Subject to Mg2+-dependent Control of Its Synthesis and Degradation

“…Cells were prepared for micros-copy as described (23). For FM4 -64 staining of the yeast vacuole, cells were pulse-labeled for 1 h in synthetic SD medium containing 30 g/l FM4 -64 (T-3166, Molecular Probes, Inc.), and FM4 -64 fluorescence was visualized after a chase of 1 h.…”

Tsc13p Is Required for Fatty Acid Elongation and Localizes to a Novel Structure at the Nuclear-Vacuolar Interface in Saccharomyces cerevisiae

“…The ceramide moiety is comprised of a fatty acid joined in amide linkage to a long-chain base (LCB). In Saccharomyces cerevisiae, the fatty acid is a hydroxylated C 26 very long chain fatty acid (VLCFA), the LCB is usually phytosphingosine, and the polar head group is an inositolphosphoryl moiety that can be further decorated by mannosylation and a second inositolphosphorylation ( Fig. 1; for a review, see reference 13).…”

“…Candidates for yeast cells defective in the elongating system were identified in two independent screens. These screens took advantage of the observation that fas2 mutants will grow in medium supplemented with myristate (C 14 ) as long as this fatty acid can be converted to the long-chain (C 16 and C 18 ) and very long chain (typically C 26 ) fatty acids by the elongating systems. An additional mutation in the fas2 background rendering cells unable to grow on FIG.…”

“…Palmitoyl-CoA is synthesized from acetyl-CoA and malonyl-CoA by soluble FAS. Palmitoyl-CoA is elongated to a C 26 VLCFA by a membrane-associated fatty acid elongating system (A, left branch). Each cycle of elongation requires four successive reactions and lengthens the growing fatty acid by two carbon units; condensation of malonyl-CoA myristate but able to grow on palmitate (C 16 ) identified the ELO1 gene (14,59).…”

Characterization, localization and functional analysis of Gpr1p, a protein affecting sensitivity to acetic acid in the yeast Yarrowia lipolytica

“…Cells were prepared for fluorescence microscopy as described by Kohlwein (2000). A sample (0?5 ml) of cell culture was placed on a coverslip and covered with a thin sheet of agarose, to immobilize cells under conditions of nutrient and oxygen supply.…”