Research: Investigations

Small Steps Toward Large-Scale Analysis of the Yeast Genome

Research in the Kumar lab focuses on the application of genomics to the study of cell and molecular biology in the budding yeast Saccharomyces cerevisiae. Defined broadly, genomics is the global or systematic study of the genes and proteins in an organism, encompassing large-scale studies of gene expression, protein activity, protein localization, and protein interactions. In contrast to classical molecular biology, genomic data provide broad functional surveys of hundreds to thousands of genes, offering insights into higher-order patterns of gene function unobtainable from individual studies of a single gene. The complementary nature of genomic and classical data sets is clear; by analogy, consider genomics to be a mechanism by which we don’t lose sight of the forest for the trees.

Yeast colony indicating the transition from vegetative growth to pseudohyphal growth

Although my lab utilizes genomic approaches to study a variety of cell responses in the budding yeast, significant effort is focused upon understanding yeast filamentous-form growth. In response to certain environmental stresses (e.g., nitrogen deprivation), some strains of yeast undergo a dramatic change in growth form: The yeast cells elongate and remain connected after budding, forming chains (or filaments) of cells called pseudohyphae. Pseudohyphal growth in yeast has garnered intense interest over recent years as a model of fungal pathogenesis. Many fungal pathogens, including the causative agents of candidiasis and meningitis, undergo an essentially identical transition to a filamentous growth form. This transition has been found to be absolutely essential for virulence in these pathogens, such that strains impaired in their ability to undergo filamentous growth are no longer infectious. Furthermore, yeast pseudohyphal growth (PHG) has proven to be an excellent model of tumor cell metastasis; the cytoskeletal rearrangements underlying yeast PHG are very similar to those observed in tumor cells during metastasis.

My lab uses four approaches to study PHG in yeast: gene expression profiling, large-scale protein localization studies, mass spectrometry, and bioinformatics. To investigate gene expression during yeast PHG, DNA microarrays are used. The DNA microarray is a miniaturized array of oligonucleotide probes for the detection of corresponding RNA transcripts over the genome as a whole. By microarray analysis, we have identified hundreds of genes apparently functioning in PHG, including several previously overlooked signaling pathways. Microarray studies are complemented with largescale studies of protein localization by which we can track individual proteins within the yeast cell. For these studies, we have fused a fluorescent protein to hundreds of yeast proteins for subsequent analysis by fluorescence microscopy. By this approach, we have identified many novel regulatory mechanisms controlling protein function and localization during yeast PHG. We have also utilized the Michigan Proteome Consortium for expertise in mass spectrometry. Using new approaches in mass spectrometry, we have identified a subset of yeast proteins differentially abundant during PHG; these proteins very likely facilitate yeast cell-cell adhesion during filamentous growth. Finally, most of our data sets are made publicly available through our online database Organelle DB. Organelle DB provides a catalog of over 25,000 proteins from nearly 60 organelles, subcellular structures, and protein complexes in 154 organisms spanning the eukaryotic kingdom. As such, Organelle DB is the first online resource devoted to the identification and presentation of eukaryotic proteins localized to organelles and subcellular structures. Organelle DB may be freely accessed at http://organelledb.lsi.umich.edu.

In total, these studies are providing a fuller knowledge base defining yeast filamentous-form growth, with relevance to our understanding of fungal pathogenesis as well as cancer biology.

— Anuj Kumar, PhD, October 2005