Unexpected Link between Metal Ion Deficiency and Autophagy in Aspergillus fumigatus

Abstract

Autophagy is the major cellular pathway for bulk degradation of cytosolic material and is required to maintain viability under starvation conditions. To determine the contribution of autophagy to starvation stress responses in the filamentous fungus Aspergillus fumigatus, we disrupted the A. fumigatus atg1 gene, encoding a serine/threonine kinase required for autophagy. The ⌬Afatg1 mutant showed abnormal conidiophore development and reduced conidiation, but the defect could be bypassed by increasing the nitrogen content of the medium. When transferred to starvation medium, wild-type hyphae were able to undergo a limited amount of growth, resulting in radial expansion of the colony. In contrast, the ⌬Afatg1 mutant was unable to grow under these conditions. However, supplementation of the medium with metal ions rescued the ability of the ⌬Afatg1 mutant to grow in the absence of a carbon or nitrogen source. Depleting the medium of cations by using EDTA was sufficient to induce autophagy in wild-type A. fumigatus, even in the presence of abundant carbon and nitrogen, and the ⌬Afatg1 mutant was severely growth impaired under these conditions. These findings establish a role for autophagy in the recycling of internal nitrogen sources to support conidiophore development and suggest that autophagy also contributes to the recycling of essential metal ions to sustain hyphal growth when exogenous nutrients are scarce.Nutrient limitation is one of the most significant stresses encountered by microorganisms in nature. Autophagy is a catabolic membrane trafficking system that counters such nutrient stress by initiating a process of limited intracellular digestion to support the organism during periods of reduced nutrient availability (26,32,37,60). The process begins with the formation of isolation membranes within the cytoplasm, whose origin is not entirely clear. These membranes progressively expand, nonselectively encapsulating cytosolic material into a doublemembrane vesicle called the autophagosome. The autophagosome fuses its outer membrane with a vacuole, releasing the autophagic body and its contents into the vacuolar lumen for degradation by resident hydrolases. Autophagy is upregulated in response to starvation stress, resulting in the generation of a pool of recycled molecules that provide the building blocks for continued synthesis of essential components until nutrient conditions improve (26,37,39,60). However, some autophagy remains constitutively active at low levels, even under nutrientreplete conditions, where it serves as a mechanism to remove old or damaged proteins and organelles (17,25,28,46,54). This provides an important form of quality control that combats the toxic accumulation of abnormal cytoplasmic components.The degradative functions of autophagy contribute to several important aspects of cell physiology, including autophagydependent programmed cell death (2, 14), cellular remodeling during development and differentiation (21,36,40,49,57), maintenance of endoplasmic reticulum homeostasis (4, 30, 6...

Autophagy-deficient Schizosaccharomyces pombe mutants undergo partial sporulation during nitrogen starvation

“…In Sch. pombe or the filamentous fungus Aspergillus fumigatus, addition of ammonium chloride as a nitrogen source is able to initiate sporulation during nitrogen starvation (Kohda et al, 2007;Richie et al, 2007). This observation suggests that a nitrogen source is necessary for sporulation, but it does not reveal the pathway of amino acid recycling that follows autophagy.…”

Guidelines for the use and interpretation of assays for monitoring autophagy

“…193,194,[798][799][800][801] In addition, macroautophagy seems to be involved in many normal developmental processes such as sexual and asexual reproduction, where there is a need for reallocation of nutrients from one part of the mycelium to another to supply the developing spores and spore-bearing structures. 194,798,799,801,802 Similarly, autophagy also affects conidial germination under nitrogen-limiting conditions. 194 In Podospora anserina, autophagy has been studied in relation to incompatibility reactions between mating strains where it seems to play a prosurvival role.…”

“…Most commonly, ATG1 and/or ATG8 has been deleted. 193,798,799,801,802,804 To confirm that the deletion(s) affects autophagy, the formation of autophagic bodies in the wild type and the mutant can be compared. In filamentous fungi the presence of autophagic bodies can be detected using MDC staining, 193,798 TEM 193,799 or fluorescence microscopy to monitor Atg8 tagged with a fluorescent protein.…”

Direct effects of non-antifungal agents used in cancer chemotherapy and organ transplantation on the development and virulence ofCandidaandAspergillusspecies

“…159,160 In a study of A. fumigatus, autophagy was required for aerial hyphae formation, conidial germination, hyphal foraging and growth under nutrient-deficient conditions; recent data have also suggested that autophagy contributes to metal ion homeostasis but the mechanism by which this occurs requires further investigation. 155,161 As for C. albicans, autophagy does not appear to be essential for virulence in mice with aspergillosis, 155,161 the reason for which remains to be defined. One possible explanation is that for A. fumigatus, the fungus is able to survive on dead/decaying matter and is relatively resistant to nutrient stress.…”