Pheromone and Receptor

The cloning strategy of the a locus is based on the observation that when both al and a2 alleles are introduced by transformation in the same cell, U. maydis behaves as a "double mater," i.e., it can mate with strains of either a mating type (Figures 8.5 and 8.6). The

Figure 8.5 Diagram of test for mating reaction in Ustilago maydis on charcoal-containing medium. Cells were cospotted as indicated on the top and on the left. Genes introduced by transformation are in parenthesis. (a) Double mater phenotype of a2b2 transformed with al. (b) albl and a1b2 were transformed with receptor gene pra2. Compatible cells produce white-filament (fuz+ phenotype). Based on Bolker, M., Urban, M. and Kahmann, R. (1992). Cell 68: 442-450, © 1992. With permission from Elsevier.

Figure 8.5 Diagram of test for mating reaction in Ustilago maydis on charcoal-containing medium. Cells were cospotted as indicated on the top and on the left. Genes introduced by transformation are in parenthesis. (a) Double mater phenotype of a2b2 transformed with al. (b) albl and a1b2 were transformed with receptor gene pra2. Compatible cells produce white-filament (fuz+ phenotype). Based on Bolker, M., Urban, M. and Kahmann, R. (1992). Cell 68: 442-450, © 1992. With permission from Elsevier.

: Mfa 1 <| : Mfa 2 Y: pra 1 V: pra 2

Figure 8.6 Schematic representation of interactions between pheromones and receptor coded by the a locus of Ustilago maydis. (a) Mating interaction between compatible cells differing at both a and b loci. (b) Double mater phenotype of a2 strain transformed with the pheromone gene from the opposite a mating type. Mating occurs between haploid strains of both a mating types. (c) Strains of the same a1 mating types can fuse if both are transformed with the receptor gene (pra2) from the opposite mating type. mfa and pra introduced by transformation are denoted in parentheses. Reprinted from Cell 68, Bolker, M., Urban, M. and Kahmann, R. (1992), pp. 442-450, © 1992. With permission from Elsevier.

nucleotide sequence of a loci revealed that they encode small size polypeptides with a carboxy-terminus sequence of Cys-A-A-X, where A is an aliphatic amino acid and X any amino acid with a farnesyl group attached to cysteine—a motif that is characteristic of pheromones (Figure 8.7). Moreover, the nucleotide sequencing data disclosed two genes called pral andpra2 that encode proteins having seven hydrophobic, potential membrane-spanning mfa1 MLSIFAQTTQTSASEPQQSPTAPQGRDNGSPIGYSSCVVA mfa2 MLSIFETVAAAAPVTAETQQASNNENRGOPGYYCLIA

Figure 8.7 Ustilago maydis. Polypeptide sequences encoded by mfal and mfa2. Amino acids are shown by one-letter symbols. The Cys-A-A-X motif at the C-terminal end is underlined. After Bolker et al. (1992).

regions, a characteristic feature of the receptors found in the membrane. From this it is inferred that a locus encodes a pheromone polypeptide and a receptor; it is the primary determinant of cell-cell recognition. Mating of al and a2 compatible cells occurs if the mfal receptor recognizes the a2 pheromone and the mfa2 receptor recognizes the al pheromone. Haploid strains of the same al mating type fuse if both are transformed with the receptor gene (pra2), provided they contained the different b allele.

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