Complex Mechanisms Regulate Developmental Expression of thematA(HMG) Mating Type Gene in HomothallicAspergillus nidulans

Abstract

Sexual reproduction is a fundamental developmental process that allows for genetic diversity through the control of zygote formation, recombination, and gametogenesis. The correct regulation of these events is paramount. Sexual reproduction in filamentous fungi, including mating strategy (self-fertilization/homothallism or outcrossing/heterothallism), is determined by the expression of mating type genes at mat loci. Aspergillus nidulans matA encodes a critical regulator that is a fungal ortholog of the hSRY/SOX9 HMG box proteins. In contrast to well-studied outcrossing systems, the molecular basis of homothallism and role of mating type genes during a self-fertile sexual cycle remain largely unknown. In this study the genetic model organism, A. nidulans, has been used to investigate the regulation and molecular functions of the matA mating type gene in a homothallic system. Our data demonstrate that complex regulatory mechanisms underlie functional matA expression during self-fertilization and sexual reproduction in A. nidulans. matA expression is suppressed in vegetative hyphae and is progressively derepressed during the sexual cycle. Elevated levels of matA transcript are required for differentiation of fruiting bodies, karyogamy, meiosis, and efficient formation of meiotic progeny. matA expression is driven from both initiator (Inr) and novel promoter elements that are tightly developmentally regulated by position-dependent and position-independent mechanisms. Deletion of an upstream silencing element, matA SE, results in derepressed expression from wildtype (wt) promoter elements and activation of an additional promoter. These studies provide novel insights into the molecular basis of homothallism in fungi and genetic regulation of sexual reproduction in eukaryotes.S EXUAL reproduction is a central part of the life cycle in most eukaryotic organisms and has a profound impact on the evolution and biology of species. Successful exchange of genetic material requires establishment of sexual identity (male and female function). Sexual identity in animals is governed by genes located on sex chromosomes, whereas in fungi mating type genes residing at mat loci (sexual-identity loci) control cell-type identity and sexual development. The correct regulation and spatiotemporal expression of sex determining genes is crucial during reproduction. Mechanisms underlying gene regulation in eukaryotic sexual development are not well understood (Harley et al. 2003;Fraser and Heitman 2005;Kashimada and Koopman 2010). Mating type genes have been described in numerous heterothallic (cross-fertile) and homothallic (self-fertile) filamentous fungi, where they function as master regulators of sexual reproduction (Kronstad and Staben 1997;Debuchy et al. 2010). The mat loci typically encode transcription factors, either Mat-HMG (high-mobility group box) or Mat-a (alpha box), proteins that coordinate expression of sex-specific genes. Several lines of study indicate that mating type genes in fungi share structural and functiona...

Novel Sexual-Cycle-Specific Gene Silencing in Aspergillus nidulans

“…Elevated hyphal expression might be the result of expression coming from two matA genes and/or regulatory derepression associated with ectopic matA copy. This later possibility is more likely and is consistent with our previous data showing similar levels of elevated hyphal matA expression from a single ectopic copy of matA (Czaja et al 2011). By contrast, expression from both endogenous and ectopic matA genes was dramatically suppressed during the latter stages of development ( Figure 4B).…”

“…Deletion of 830 bp of the upstream regulatory region of matA resulted in a 140-fold increase of matA RNA abundance in both hyphal and sexually differentiated tissue relative to the wild type. However, silencing is not triggered in this strain, and fertility is like the wild type (Czaja et al 2011).Therefore, neither MatA protein nor the level of matA RNA per se is involved in gene silencing. It appears that the transgene-derived RNA itself triggers the gene-silencing phenomenon.…”

“…Homologs of the budding yeast pheromone receptor genes gprA (a-factor receptor) and gprB (a-factor receptor) in homothallic A. nidulans have been previously reported (Seo et al 2004). Our previous studies have determined that the MatA transcription factor modulates expression of the gprA gene during sexual development having activator and/or a combination of activator/inhibitor functions (Czaja et al 2011). …”

“…First, we used the UI465 matA(0) strain that carries a deletion of the entire matA transcription unit (from +1 to +1851 nt) at the resident locus. UI465 is sterile and does not differentiate fruiting bodies or ascospores (Czaja et al 2011). UI465 was complemented with an ectopically integrated matA transgene (from 21001 to +3056 nt) ( Figure 5A).…”

“…A second approach used a strain carrying a 299-nt partial deletion of the matA-coding region (+148 to +446) at the resident locus. Strain UI464 (matAD) was also sterile, having a phenotype identical to that of the matA (0) deletion strain (Czaja et al 2011). UI464 was complemented by a matA transgene (from 21001 to +3056 nt) integrated ectopically.…”