Scars, bud

Fig. 1. Scanning electron micrograph showiag ceUs of S. cerevisiae. Bud scars are visible at the ends of ceUs. Scale bar is 5 p.m.

Apart from the presence of chitin, no gross structural differences were detected between the bud scars of S. cerevisiae and the rest of the cell wall."... 

Many articles have appeared concerning the content and location of chitin, which is virtually universal in the cell walls of fungi. Both chi-tin (17%) and chitosan, which lacks N-acetyl groups, are present in the cell wall of Choanephora curcurbitarum.194 Chitin may be detected in bud scars, cell wall, and cytoplasm, near the plasmalemma of Sac-charomyces cerevisiae and Candida utilis, as detected by wheat-germ agglutinin.82... 

Flow cytometry [141, 142] is a technique that allows the measurement of multiple parameters on individual cells. Cells are introduced in a fluid stream to the measuring point in the apparatus. Here, the cell stream intersects a beam of light (usually from a laser). Light scattered from the beam and/or cell-associated fluorescence are collected for each cell that is analysed. Unlike the majority of spectroscopic or bulk biochemical methods it thus allows quantification of the heterogeneity of the cell sample being studied. This approach offers tremendous advantages for the study of cells in industrial processes, since it not only enables the visualisation of the distribution of a property within the population, but also can be used to determine the relationship between properties. As an example, flow cytometry has been used to determine the size, DNA content, and number of bud scars of individual cells in batch and continuous cultures of yeast [143,144]. This approach can thus provide information on the effect of the cell cycle on observed differences between cells that cannot be readily obtained by any other technique. 

Other vital stains take advantage of different cellular properties which can be correlated with cellular physiology Propidium Iodide, Ethidium Bromide, Ethidium Monoazide, Calcofluor White have been widely used to indicate the presence of dead eukaryotes or prokaryotes cells. 2-(p-iodophenyl-)3)(p-nitro-phenyl)-5-phenyl tetrazolium chloride (INT) belongs to a class of stains which can be used to determine if a cell or hyphal compartments [180] can maintain an internal reducing environment (Fig. 20a). There are, however, still a large debate about the reliability of those techniques, depending upon the cells under consideration [181]. Calcofluor (Aex = 380 nm, Aem 420 nm) is a specific cell wall stain which enables to counts buds scars on Saccharomyces cerevisiae [29] to estimate the age of a cell. 

A historical review of the development of our knowledge of the yeast cell wall was given by Phaff (I). Most information based on chemical studies has been derived, by far, from studies with cell walls from baker s yeast, Saccharomyces cerevisiae, and closely related species. The principal components of Saccharomyces walls are several types of glucan and a mannan-protein complex which may contain variable proportions of phosphate. A low content of chitin (ca. 1% ) may be present depending on the number of times a cell has produced buds. The reason for this is that chitin is present only in the bud scars (ca. 3 pm2 in area) produced on the surface of a mother cell (2), each at a different place on the cell surface. 

Some yeast cells divide by the production of a bud on the cell surface. The bud is much smaller than the cell itself and grows till it reaches adult dimensions. The release of each bud leaves a "bud scar" on the parent cell. It is not possible for this... 

Since the coumarin compounds show phosphodiesterase inhibitory activity, the observed change in the morphology of Candida albicans could be partly due to this property. Each Candida yeast cell can normally have a budding scar or the daughter cell still attached to it [319]. The possible cause of multibudding phenomenon could be PD inhibition. A similar observation was done in treatment with caffeine, which at the subinhibitory concentrations caused an increase in unusual modes of proliferation with signs of multiple budding in Candida albicans [320]. 

Key words Chemotropism, Mating pheromone. Bud scar, Gradient, Zygote... 

UV excitation (excitation bandpass filter 340-380 nm, 400 nm dichroic, and emission longpass filter 425 nm) to visualize the bud scars (see Note 16). 

Fig. 3. Examples of zygotes with both bud scars and birth scars visible. Bud scars and birth scars are in different focal planes.

The percentages of cells with the bud scar proximal, medial, or distal to the site of fusion are then calculated for the wild-type and mutant strains. In general, in contrast to wild-type cells, chemotropism mutants mate by default and thus display growth sites during mating that are adjacent to the bud scar. As a result, while wild-type cells should have relatively equal percentages of proximal, medial, and distal bud scars relative to the site of fusion, chemotropism mutants should have a majority of cells with bud scars proximal to the site of fusion (Fig. 4). It should be noted that such analysis is not possible with mutants that perturb axial bud site selection. 

If the mutant cells are chemotropism defective, then the mating efficiency should be substantially lower (e.jj., 20-50-fold) than in wild-type cells. This plating step is thus included to provide further confirmation that cells showing nonrandom growth relative to the bud scar are indeed chemotropism defective. 

It is important to distinguish the birth scar, which is also visible upon calcofluor white staining, from the bud scar. The birth scar is typically larger (>1 pm) and less strongly stained than the bud scar, which is usually visible as a clear ring of fluorescence (Fig. 3). 

Figure 6.4 A. The Putative Structure of the Yeast Cell Wall. Disulphide bridges hold the mannan glycoprotein and glycan together. Chitin is confined to bud scars. B. The putative structure of the wall of a hyphal fungus. Septa differ slightly from hyphal walls in many species. The plasma membrane is indicated by M.

The position of chitin in the wall of 5. cere vwiae has long been a matter of uncertainty and argument, but recent evidence puts it quite firmly in the region of bud scars where it appears to form a ring, perhaps for reinforcement. 

Recent evidence indicates that in addition to mannoproteins, some P-glucan is e iosed on the surface of C. albicans yeasts, at sites that presumptively represent bud scars. During cell division, B-glucans and chitin become exposed at the site of separation of the dai ter from the mother... 

Fig. 16.2 Diagram of an electron micrograph of a section through a resting cell of bakers yeast Saccharomyces certvisiae). ER, endoplasmic reticulum M, mitochondrion N, nucleus Nm, nuclear membrane Nn, nucleolus Pi, invagination PI, plasmalemma V, vacuole Vp, polymetaphosphate granule W, cell wall Ws, bud scar L, lipid granule (sphaerosome).

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