Yeast genomic expression patterns in response to low-shear modeled microgravity

Abstract

The low-shear microgravity environment, modeled by rotating suspension culture bioreactors called high aspect ratio vessels (HARVs), allows investigation in ground-based studies of the effects of microgravity on eukaryotic cells and provides insights into the impact of space flight on cellular physiology. We have previously demonstrated that low-shear modeled microgravity (LSMMG) causes significant phenotypic changes of a select group of Saccharomyces cerevisiae genes associated with the establishment of cell polarity, bipolar budding, and cell separation. However, the mechanisms cells utilize to sense and respond to microgravity and the fundamental gene expression changes that occur are largely unknown. In this study, we examined the global transcriptional response of yeast cells grown under LSMMG conditions using DNA microarray analysis in order to determine if exposure to LSMMG results in changes in gene expression.

Results: LSMMG differentially changed the expression of a significant number of genes (1372) when yeast cells were cultured for either five generations or twenty-five generations in HARVs, as compared to cells grown under identical conditions in normal gravity. We identified genes in cell wall integrity signaling pathways containing MAP kinase cascades that may provide clues to novel physiological responses of eukaryotic cells to the external stress of a low-shear modeled microgravity environment. A comparison of the microgravity response to other environmental stress response (ESR) genes showed that 26% of the genes that respond significantly to LSMMG are involved in a general environmental stress response, while 74% of the genes may represent a unique transcriptional response to microgravity. In addition, we found changes in genes involved in budding, cell polarity establishment, and cell separation that validate our hypothesis that phenotypic changes observed in cells grown in microgravity are reflected in genome-wide changes. This study documents a considerable response to yeast cell growth in low-shear modeled microgravity that is evident, at least in part, by changes in gene expression. Notably, we identified genes that are involved in cell signaling pathways that allow cells to detect environmental changes, to respond within the cell, and to change accordingly, as well as genes of unknown function that may have a unique transcriptional response to microgravity. We also uncovered significant changes in the expression of many genes involved in cell polarization and bud formation that correlate well with the phenotypic effects observed in yeast cells when grown under similar conditions. These results are noteworthy as they have implications for human space flight.

Figure 1

Gene expression response to low-shear modeled microgravity. Two-dimensional hierarchical cluster heat map analysis of the microarray transcriptional profile of gene response to low-shear modeled microgravity (LSMMG) in S. cerevisiae wild-type diploid cells (BY4743) grown in HARVs (experimental group) for 5 generations (3E5, 4E5, 5E5) or 25 generations (3E25, 4E25, 5E25) versus the control group grown in HARVs for 5 generations (1C5, 2C5, 3C5, 4C5) or 25 generations (1C25, 2C25). The analysis was performed on 1372 statistically significant genes (from the filtered list of 3850 genes) with a fold change of at least 2 in one of three comparisons as described in Methods. Levels of expression are indicated on a color scale where red represents the up-regulated genes and green, the down-regulated genes. Similarities or differences in the experimental versus control gene expression patterns are represented in the horizontal dendrogram. The vertical dendrogram shows similarities in expression patterns between cultures exposed to LSMMG versus control cultures.

Figure 2

A comparison of the low-shear modeled microgravity response (SMG) with a previously identified list of 868 S. cerevisiae Environmental Stress Response (ESR) genes with a fold change >2 identified by Gasch et al. (2002). 363 genes from our dataset change > 2-fold under all conditions.

Figure 3

IPS functional classification of the list of 1006 significant genes that respond to LSMMG that are not general environmental stress response genes as identified by Gasch et al., 2000. Black bars represent genes that are up-regulated, gray bars represent the down-regulated genes.

Figure 4

A. A comparison of the low-shear modeled microgravity response (SMG) with 311 MIPS functional category (43.01.03.05) genes involved in budding and cell polarity functions. B. The 31genes from the list of 311 that are up-regulated greater than 2-fold compared to controls. C. The 30 genes from the list of 311 that are down-regulated greater than 2-fold compared to controls. Expression levels are indicated on the color scale bars.