Autophagy-Associated Protein SmATG12 Is Required for Fruiting-Body Formation in the Filamentous Ascomycete Sordaria macrospora.
In filamentous fungi, autophagy functions as a catabolic mechanism to overcome starvation and to control diverse developmental processes under normal nutritional conditions. Autophagy involves the formation of double-membrane vesicles, termed autophagosomes that engulf cellular components and bring about their degradation via fusion with vacuoles. Two ubiquitin-like (UBL) conjugation systems are essential for the expansion of the autophagosomal membrane: the UBL protein ATG8 is conjugated to the lipid phosphatidylethanolamine and the UBL protein ATG12 is coupled to ATG5. We recently showed that in the homothallic ascomycete Sordaria macrospora autophagy-related genes encoding components of the conjugation systems are required for fruiting-body development and/or are essential for viability. In the present work, we cloned and characterized the S. macrospora (Sm)atg12 gene. Two-hybrid analysis revealed that SmATG12 can interact with SmATG7 and SmATG3. To examine its role in S. macrospora, we replaced the open reading frame of Smatg12 with a hygromycin resistance cassette and generated a homokaryotic ΔSmatg12 knockout strain, which displayed slower vegetative growth under nutrient starvation conditions and was unable to form fruiting bodies. In the hyphae of S. macrospora EGFP-labeled SmATG12 was detected in the cytoplasm and as punctate structures presumed to be phagophores or phagophore assembly sites. Delivery of EGFP-labelled SmATG8 to the vacuole was entirely dependent on SmATG12.
Institute of Microbiology and Genetics, Department of Genetics of Eukaryotic Microorganisms, Georg-August University, Göttingen, Germany.
Antonia
A
Werner
Institute of Microbiology and Genetics, Department of Genetics of Eukaryotic Microorganisms, Georg-August University, Göttingen, Germany.
Britta
B
Herzog
Institute of Microbiology and Genetics, Department of Genetics of Eukaryotic Microorganisms, Georg-August University, Göttingen, Germany.
Stefan
S
Frey
Institute of Microbiology and Genetics, Department of Genetics of Eukaryotic Microorganisms, Georg-August University, Göttingen, Germany.|Göttingen Center for Molecular Biosciences (GZMB), Georg-August University, Göttingen, Germany.
Stefanie
S
Pöggeler
D000595:Amino Acid Sequence; D001343:Autophagy / Q000235:genetics*; D000071192:Autophagy-Related Protein 12 / Q000235:genetics* / Q000378:metabolism; D000071193:Autophagy-Related Protein 7 / Q000235:genetics / Q000378:metabolism; D000071190:Autophagy-Related Protein 8 Family / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D048690:Fruiting Bodies, Fungal / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D025301:Hyphae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D016366:Open Reading Frames; D010588:Phagosomes / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D020037:Sordariales / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D020798:Two-Hybrid System Techniques; D014617:Vacuoles / Q000378:metabolism / Q000648:ultrastructure
D000071192:Autophagy-Related Protein 12; D000071190:Autophagy-Related Protein 8 Family; D005656:Fungal Proteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; D000071193:Autophagy-Related Protein 7