Vacuole segregation and mycelial development of Candida albicans

Candida albicans is the most common fungus that is associated with life-threatening infection. It is one of many fungi that can grow either as an ovoid yeast that grows by forming expanding buds or as a mould that elaborates branching tubular cells to form a mycelium. Because infections are associated with transitions between these two forms the process of yeast-to-hypha transition has been heavily studied in recent years. However, most of these investigations have focussed on the signals that stimulate the transitions rather than the growth of the primary germ tube and its associated branches. This is important because mycelial growth may be vital for tissue invasion, and because it has become clear that the process of hyphal C. albicans is unusual and enables novel hypotheses to be tested about the physiological requirements for cell division of eukaryotic cells in general. Consequently we will investigate how mycelial cells grow and divide both from the point of view of its role in fungal disease and the insights it can provide into the cell biology of the cell cycle. We have observed that during the growth of germ tubes, a large vacuole forms behind the growing tip that fills up most of the cell. Although the overall dimensions of this cell are similar to other cells, it has less cytoplasm. We propose that it does not have sufficient cytoplasm to progress through the cell division cycle. At a specific point in the cell cycle called START, cells must achieve a certain minimal cell size. We therefore hypothesise that the large vacuole left behind after cell division prevents these cells from passing through START in the division cycle. We have also shown that the vacuole is not equally divided between the two daughter cells formed after cell division, and that the younger daughter cell acquires most of the cytoplasm and the mother cell retains most of the vacuole. Vacuole division and inheritance is a carefully regulated process about which we have learned much from the related yeast-like fungus Saccharomyces cerevisiae. To test out hypothesis that vacuole inheritance and distribution determines whether growth or growth arrest of cells occurs within the mycelium, we will make defined mutations in genes that regulate vacuole partitioning at cell division. We can predict from studies in S. cerevisiae exactly what mutations we will need to make to be able to alter the normal vacuole pattern in cells. These mutants will enable us not only to find out how vacuoles control cell division, but also to address questions about how the cell cycle in regulated in higher eukaryotic cells. With a range of mutants we will be able to dissect the regulatory pathways through which hyphae and branches of mycelia grow. We will also be able to assess whether normal branching is important for the ability of this fungus to establish diseased lesions in animal tissues. Importantly, many other important fungi, including many plant pathogens, control the type of growth they undergo by forming a greater or lesser amount of vacuole. We will therefore be able to advance the field of fungal physiology, by undertaking the first in- depth study of the genetic links between vacuolation and fungal growth.