An apparent increase in meiotic silencing strength in crosses involving inbred Neurospora crassa strains.

Meiotic silencing by unpaired DNA is a presumed RNAi-mediated elimination of the transcripts of any gene that is not properly paired with a homolog in meiosis. Wild-isolated strains of Neurospora crassa can be classified into three types based on the apparent strength of meiotic silencing of the bml (ß-tubulin) and mei-3 genes in crosses of the wild strains with the tester strains ::Bml(r) and ::mei-3. Crosses with "OR" and "Sad" type wild-isolates, respectively, did or did not silence both the genes, whereas crosses with the "Esm" type silenced bml, but not mei-3(+). Presumably, bml is more sensitive to silencing than mei-3, and silencing is strongest in crosses with the OR type, weakest (or non-evident) in crosses with the Sad type, and of intermediate strength in crosses with Esm type. Now, while constructing recombinant inbred lines from the Sad type wild strains Bichpuri-1 a and Spurger-3 A, we found that although crosses of Bichpuri-1 a and Spurger-3 A, and of most of their f1 progeny, with the ::Bml(r) and ::mei-3 testers were consistently and reproducibly Sad type, those of later generation strains of each line could show Sad, Esm, or a variably expressive Sad/Esm type. Since any genotype found in a later generation is, in principle, obtainable in the f1, the observed transition from an apparently stable Sad phenotype to an apparently unstable Sad/Esm phenotype appears not to be due to genotype differences between the generations. Therefore, the Sad versus Esm difference may have a genotype-independent basis.

Meiotic silencing by unpaired DNA is expressed more strongly in the early than the late perithecia of crosses involving most wild-isolated Neurospora crassa strains and in self-crosses of N. tetrasperma.

Meiotic silencing by unpaired DNA is a presumed RNAi-mediated elimination of the transcripts of any gene that is not properly paired with a homolog in meiosis. Eighty wild-isolated strains of Neurospora crassa were classified into three types based on the apparent strength of meiotic silencing of the bml (ß-tubulin) and mei-3 genes in crosses with the ::Bml(r) and ::mei-3 tester strains. "OR" and "Sad" type wild-isolates, respectively, did or did not silence both the genes, whereas the "Esm" type (68 strains) silenced bml but not mei-3(+), suggesting an intermediate strength of silencing. Many wild strains, especially of the Esm type, gave fertile crosses with strains bearing the 148 kbp chromosome segment duplication Dp(EB4) but their crosses with the 405 kbp Dp(IBj5) were barren. Larger Dps might include more genes whose sensitivity to silencing is comparable with that of bml, consequently their crosses with the OR and Esm type strains would be about equally barren, whereas small Dps, with presumably few or no bml-like genes suffer little gene silencing in crosses with Esm and Sad strains and thus remain fertile. Using lollipop-shaped asci as indicators of silencing in crosses of the ::act tester with Esm type strains, and white-spored asci in Neurospora tetrasperma self-crosses heterozygous for an ectopic insertion of the asm-1 gene, we found that meiotic silencing was expressed more strongly in the early but not in the late perithecia of these crosses.

A factor in a wild isolated Neurospora crassa strain enables a chromosome segment duplication to suppress repeat-induced point mutation.

Repeat-induced point mutation (RIP) is a sexual stage-specific mutational process of Neurospora crassa and other fungi that alters duplicated DNA sequences. Previous studies from our laboratory showed that chromosome segment duplications (Dps) longer than (approx.) 300 kbp can dominantly suppress RIP, presumably by titration of the RIP machinery, and that although Dps less than 200 kbp did not individually suppress RIP, they could do so in homozygous and multiply heterozygous crosses, provided the sum of the duplicated DNA exceeds (approx.) 300 kbp. Here we demonstrate suppression of RIP in a subset of progeny carrying the normally sub-threshold 154 kbp Dp(R2394) from a cross of T(R2394) to the wild isolated Carrefour Mme. Gras strain (CMG). Thus, the CMG strain contains a factor that together with Dp(R2394) produces a synthetic RIP suppressor phenotype. It is possible that the factor is a cryptic Dp that together with Dp(R2394) can exceed the size threshold for titration of the RIP machinery and thereby causes RIP suppression.

Carrefour Mme. Gras: a wild-isolated Neurospora crassa strain that suppresses meiotic silencing by unpaired DNA and uncovers a novel ascospore stability defect.

Ordinarily, RIP-induced erg-3 mutant Neurospora crassa ascospores and their erg(+) siblings do not differ in stability during long-term storage. Consequently, the frequency of RIP-induced erg-3 mutants remains about constant regardless of the time that has elapsed between ascospore harvest and germination. We found, however, that RIP-induced erg-3 mutants were apparently selectively lost with time from among the ascospores stored from a cross with the wild-isolated Carrefour Mme. Gras strain from Haiti. The Haitian strain was also found to exert a dominant suppression of meiotic silencing by unpaired DNA. Similar loss of RIP-induced erg-3 mutant ascospores was seen among the stored ascospores from a subset of crosses heterozygous for the semi-dominant Sad-1 or Sad-2 suppressors of meiotic silencing. Our results suggest that crosses suppressed in meiotic silencing can compromise the stability during storage of ascospores that inherit RIP-induced mutations.