Penicillium chrysogenum is a filamentous fungus of major medical and historical importance, being the original and present-day industrial source of the antibiotic penicillin. The species has been considered asexual for more than 100 y, and despite concerted efforts, it has not been possible to induce sexual reproduction, which has prevented sexual crosses being used for strain improvement. However, using knowledge of mating-type (MAT) gene organization, we now describe conditions under which a sexual cycle can be induced leading to production of meiotic ascospores. Evidence of recombination was obtained using both molecular and phenotypic markers. The identified heterothallic sexual cycle was used for strain development purposes, generating offspring with novel combinations of traits relevant to penicillin production. Furthermore, the MAT1-1-1 mating-type gene, known primarily for a role in governing sexual identity, was also found to control transcription of a wide range of genes with biotechnological relevance including those regulating penicillin production, hyphal morphology, and conidial formation. These discoveries of a sexual cycle and MAT gene function are likely to be of broad relevance for manipulation of other asexual fungi of economic importance.sexual recombination | secondary metabolism | ascomycete

Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the ␣ domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of crossspecies microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes.Sex, one mechanism of the genetic diversity of species, is ubiquitous across kingdoms. To avoid self-crossing, genetic barriers have evolved that prevent selfing, and these often culminate in sexual dimorphism. In filamentous ascomycetes, sexual dimorphism is almost nonexistent, and in many cases individuals are hermaphrodites. Here, sex is determined genetically by a sex-specific region in the genome known as the mating-type locus (MAT) (10).Fungi exhibit two different sexual life styles, homothallism (self-fertility) and heterothallism (self-sterility) (46). This phenomenon was first discovered by Blakeslee in the group of zygomycetes, a lineage that diverged early within the fungal kingdom (5). Only recently was the sequence of the matingtype locus of Phycomyces blakesleeanus (Zygomycota) discovered (36). It was shown that each mating-type locus contains one single gene coding for a protein with a high-mobility group (HMG) (31) (Fig. 1). Both genes show low-level amino acid similarity and confer the ability to mate as either a MAT (Ϫ) or a MAT (ϩ) strain.Similarly, heterothallic ascomycetes contain a single matingtype locus with two alternate alleles. The DNA sequences at the mating-type locus in individuals of different mating types show almost no homology. To emphasize the dissimilarity between and the different origins of the genes of different matingtype loci, they have been termed idiomorphs instead of alleles (49).In ascomycetes, mating is best characterized at the molecular level in the budding yeast Saccharomyces cerevisiae. The MAT idiomorphs, MATa and MAT␣, encode regulatory proteins which, in combination with other transcription factors, are responsible for a distinct pattern of expression in the three yeast cell types: haploid MATa and MAT␣ cells and d...

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DanksagungMein besonderer Dank gilt Frau Prof. Dr. Stefanie Pöggeler für ihre exzellente Betreuung dieser Doktorarbeit, für die aktuelle und interessante Themenstellung und ihr aufrichtiges Interesse am Fortgang dieser Arbeit. Nicht zuletzt möchte ich mich für ihre zahlreichen wissenschaftlichen Anregungen und der Bereitstellung von Laborplatz und Arbeitsmitteln bedanken. Ein großer Dank gebührt Herrn Prof. Dr. Gerhard Braus für die Übernahme des Korreferats dieser Arbeit. Am Lehrstuhl für Allgemeine und Molekulare Botanik in Bochum möchte ich mich bei Frau Swenja Elßel für ihre Hilfe und Anleitung bei der RNA-Präparation für die Microarray Experimente bedanken. Frau Dr. Minou Nowrousian war eine unschätzbare Hilfe bei der Auswertung eben jener Experimente. Herrn Jay C. Dunlap und Frau Jennifer J. Loros (Dartmouth Medical School, NH, USA) möchte ich für die Bereitstellung der Microarray Chips danken und Frau Carol Ringelberg für die Durchführung der Screens. Ein Dankeschön gilt allen wissenschaftlichen, technischen und ehemaligen Mitarbeitern der Abteilung Genetik eukaryotischer Mikroorganismen für das gute Arbeitsklima. Ganz besonders möchte ich mich hiermit bei Herrn Oliver Voigt und Herrn Ronny Lehneck für die Durchsicht dieser Arbeit und ihre hilfreichen Kommentaren bedanken. Frau Sabine Riedel gebührt mein Dank für ihre Hilfe bei den Yeast-Two-Hybrid Analysen. Mein ganzer Dank gilt meiner Familie für die außergewöhnliche Unterstützung auch in schwierigen Zeiten. Ganz besonderer Dank gebührt meinen Eltern, meiner Großmutter und Frau Madeleine Hübenthal ohne die diese Arbeit niemals möglich gewesen wäre. Leider reicht der Platz hier nicht aus um allen Menschen zu danken, die mit Rat, Tat und Unterstützung zu dieser Arbeit beigetragen haben. Allen nicht Namentlich erwähnten sage ich hiermit: Danke.