Mutational analysis of the [Het-s] prion analog of Podospora anserina: A short N-terminal peptide allows prion propagation

The het-s locus is one of nine known het (heterokaryon incompatibility) loc i of the fungus Podospora anserina. This locus exists as two wild-type alle les, het-s and het-S, which encode 289 amino acid proteins differing at 13 amino acid positions. The het-s and het-S alleles are incompatible as their coexpression in the same cytoplasm causes a characteristic cell death reac tion. We have proposed that the HET-s protein is a prion analog. Strains of the het-s genotype exist in two phenotypic states, the neutral [Het-s*] an d the active [Het-s] phenotype. The [Het-s] phenotype is infectious and is transmitted to [Het-s*] strains through cytoplasmic contact. het-s and het- Swere associated in a single haploid nucleus to generate a self-incompatibl e strain that displays a restricted and abnormal growth. In the present art icle we report the molecular characterization of a collection of mutants th at restore the ability of this self-incompatible strain to grow. We also de scribe the functional analysis of a series of deletion constructs and site- directed mutants. Together, these analyses define positions critical for re activity and allele specificity. We show that a 112-amino-acid-long N-termi nal peptide of HET-s retains [Het-s] activity. Moreover, expression of a mu tant het-s allele truncated at position 26 is sufficient to allow propagati on of the [Het-s] prion analog.