The Genetic Architecture of Biofilm Formation in a Clinical Isolate of Saccharomyces cerevisiae

Abstract

Biofilms are microbial communities that form on surfaces. They are the primary form of microbial growth in nature and can have detrimental impacts on human health. Some strains of the budding yeast Saccharomyces cerevisiae form colony biofilms, and there is substantial variation in colony architecture between biofilm-forming strains. To identify the genetic basis of biofilm variation, we developed a novel version of quantitative trait locus mapping, which leverages cryptic variation in a clinical isolate of S. cerevisiae. We mapped 13 loci linked to heterogeneity in biofilm architecture and identified the gene most closely associated with each locus. Of these candidate genes, six are members of the cyclic AMP-protein kinase A pathway, an evolutionarily conserved cell signaling network. Principal among these is CYR1, which encodes the enzyme that catalyzes production of cAMP. Through a combination of gene expression measurements, cell signaling assays, and gene overexpression, we determined the functional effects of allelic variation at CYR1. We found that increased pathway activity resulting from protein coding and expression variation of CYR1 enhances the formation of colony biofilms. Four other candidate genes encode kinases and transcription factors that are targets of this pathway. The protein products of several of these genes together regulate expression of the sixth candidate, FLO11, which encodes a cell adhesion protein. Our results indicate that epistatic interactions between alleles with both positive and negative effects on cyclic AMP-protein kinase A signaling underlie much of the architectural variation we observe in colony biofilms. They are also among the first to demonstrate genetic variation acting at multiple levels of an integrated signaling and regulatory network. Based on these results, we propose a mechanistic model that relates genetic variation to gene network function and phenotypic outcomes. BIOFILMS are complex, surface-adherent communities of microbes. They are ubiquitous in nature and in the human environment (Donlan and Costerton 2002;López et al. 2010). Not only can they be life threatening when they form in the human body (Hall-Stoodley et al. 2004), but biofilms also create problems when they form on human-made structures, ranging from merely annoying (shower curtains) to hazardous (nuclear power plants) (Flemming 2002). Because of the frequency with which we interact with biofilms, characterizing the cellular mechanisms that regulate them and understanding the genetic basis of their variation are of great interest.The ability to form biofilms is a multifactorial trait, resulting from the interactions of several lower-level properties, including cell-cell and cell-substrate adhesion; production of extracellular matrix; and spatial heterogeneity in cell morphology, growth, and division. Recent studies have demonstrated that the budding yeast Saccharomyces forms microbial communities that have these biofilm characteristics (Kuthan et al. 2003;Váchová et al. 2011). These ch...

Production of protein and metabolites by yeast grown in solid state fermentation: present status and perspectives

“…The composition and regulation analysis of gene expression for the constitution of biofilms comes mainly from studies on pathogenic yeast such as Candida sp . However, there are also studies that have focused on other yeasts with biotechnological interest such as S. cerevisiae . In addition, the formation of biofilm yeast has several metabolic implications; on the one hand, formation of these aggregates exposes differentially cells that are in the interior of the biofilm from those that are on the surface of the aggregate, as can be demonstrated by the work of Lopez et al ., where the application of an inducer, such as methanol, in atomized (aerosol) form on a mosaic of cell aggregates of Pichia pastoris embedded in polyurethane foam, resulted in a smaller amount of recombinant laccase with respect to a smaller amount of yeast suspended in liquid medium within shake flasks.…”

“…studied the production of bio‐surfactants using SSF without the need to contaminate the product with tensoactive compounds because there was no foam in the system. In addition, it is necessary to emphasize that there are no studies of biofilms for genus of yeasts such as Pichia and Kluyveromyces, however, studies with Saccharomyces and other yeasts have provided relevant information about the genes involved in cellular aggregation . On the other hand, the analysis of micrographs of cell aggregates of P. pastoris growing on polyurethane foam, show the capacity of the yeast to grow in the form of biofilms (Fig.…”

Biofilms 2012: New Discoveries and Significant Wrinkles in a Dynamic Field

“…Using next-generation sequencing of fungal isolates exhibiting substantial variation in colony architecture and a subsequent genome-wide expression analysis of biofilm and nonbiofilm segregants, candidate genes responsible for biofilm variation were identified. The results led to the development of a mechanistic model that relates genetic variation to gene network function and phenotypic outcomes (63).…”

Revisiting Mortimer’s Genome Renewal Hypothesis: Heterozygosity, Homothallism, and the Potential for Adaptation in Yeast

“…There is abundant phenotypic variation for each of the traits and the multivariate phenotypic space represented by these offspring is equally rich (Magwene, unpublished data). Illustrative of the genetic and phenotypic diversity of such strains, my laboratory has recently used such a population to map QTLs for biofilm formation in S. cerevisiae (Granek et al, 2013). …”

“…288 homozygous diploid offspring were generated by sporulation followed by autodiploidization. Traits assessed include: A) resistance to the antifungal drug fluconazole (MIC 50 as determined in microtiter plates); B) a measure of colony biofilm complexity (Granek et al, 2013); C) a measure of invasive growth on agar substrates (logarithm of the ratio of post-wash to pre-wash colony density; (Drees et al, 2005)); and D) a measure of growth at high temperature on agar plates (square root of the mean spot density for three replicates of each segregant grown at 42°C). The red arrow in each subfigure indicates the phenotype of the parental strain YJM311.…”