AG alpha 1 is the structural gene for the Saccharomyces cerevisiae alpha-agglutinin, a cell surface glycoprotein involved in cell-cell interactions during mating.

Abstract

We have cloned the a-agglutinin structural gene, AGal, by the isolation of a-specific agglutination-defective mutants, followed by isolation of a complementing plasmid. Independently isolated a-specific agglutinationdefective mutations were in a single complementation group, consistent with biochemical results indicating that the a-agglutinin is composed of a single polypeptide. Mapping results suggested that the complementation group identified by these mutants is allelic to the agal mutation identified previously. Expression ofAGal RNA was a specific and inducible by a-factor. Sequences similar to the consensus sequences for positive control by MATal and pheromone induction were found upstream of the AGaJ initiation codon. The AGal gene could encode a 650-amino-acid protein with a putative signal sequence, 12 possible N-glycosylation sites, and a high proportion of serine and threonine residues, all of which are features expected for the a-agglutinin sequence. cells of other yeast species (7,54). Therefore, the glycoprotein-glycoprotein interactions are quite specific even though the analogous agglutinins of different yeast species share biochemical characteristics.Biochemical analysis has indicated that the a-agglutinin of S. cerevisiae is a glycoprotein of approximate 160 kilodaltons (kDa) (45). Removal of N-linked carbohydrate generates a major species of 72 kDa and several minor species. Structural analysis and determination of agglutinin binding determinants have been hampered by low yields and carbohydrate-generated heterogeneity of the agglutinins. A genetic approach allowed the isolation of a putative a-agglutinin structural mutant, agal (43). The agal mutation results in an a-specific agglutination defect, resulting from the lack of expression of active a-agglutinin, and a decrease in a mating efficiency. Additional mutants have been isolated that result in agglutination defects in both a and a cells and also affect pheromone production and response (17); the pleiotropic effects of these mutations indicate that they do not identify agglutinin structural genes.We have initiated a genetic and molecular approach to the study of agglutinin structure and function, which will complement the biochemical approach that has been used previ-

Genome-wide screen for genes involved in eDNA release during biofilm formation by Staphylococcus aureus

“…S4). This result is akin to the phenotype seen in traditional agglutination assays, in which agglutinated erythrocytes or yeast cells form diffuse pellets, whereas nonagglutinated cells form compact pellets (21,22). Imaging revealed that the wild-type cells were in large clumps, whereas the mutant cells and DNase I-treated cells were in much smaller clumps (Fig.…”

Genetically Engineered Transvestites Reveal Novel Mating Genes in Budding Yeast

“…We also drove the expression of BAR1 with an engineered version of the haploid-specific promoter P FUS1 (Ingolia and Murray 2007), which is expressed in both MATa and MATa cells (Trueheart et al 1987), and the expression of the a-factor transporter STE6 with the MFa1 promoter. Since we tested mating efficiency on solid media, we did not manipulate the expression of the mating agglutinins, which function mainly when cells mate in liquid (Lipke et al 1989;Roy et al 1991).…”

“…One of these is the MATa-specific a-factor protease, Bar1 (Sprague and Herskowitz 1981;MacKay et al 1988), which helps MATa cells detect an a-factor gradient and polarize toward MATa partners (Jackson and Hartwell 1990; Barkai et al 1998). Yeast cells also express mating-type-specific agglutinins, which help cells attach to mating partners (Cappellaro et al 1991) in liquid but individually have little effect on mating efficiency on solid media (Lipke et al 1989;Roy et al 1991;de Nobel et al 1995). Evidence for the final, characterized MATa-specific gene was produced by Bender and Sprague (1989) who noted that cells expressing MATa-specific proteins and Ste3 were unable to mount a pheromone response.…”

Delineation of Functional Regions within the Subunits of theSaccharomyces cerevisiae Cell Adhesion Molecule a-Agglutinin

“…The ␣-agglutinin gene AG␣1 (2,3) encodes a 650-residue precursor with an N-terminal signal sequence that directs ␣-agglutinin to the secretory pathway and a C-terminal hydrophobic sequence that results in attachment to a glycosyl phosphatidylinositol (GPI) 1 anchor (4,5). Attachment of many cell wall proteins, including the agglutinins, involves transfer from a plasma membrane-linked GPI anchor to cell wall glycan by a transglycosylation reaction (5,6).…”