Control of morphology and virulence by ADP-ribosylation factors (Arf) in Mucor circinelloides

Abstract

Mucor circinelloides is a dimorphic fungus used to study cell differentiation that has emerged as a model to characterize mucormycosis. In this work, we identified four ADP-ribosylation factor (Arf)-encoding genes (arf1-arf4) and study their role in the morphogenesis and virulence. Arfs are key regulators of the vesicular trafficking process and are associated with both growth and virulence in fungi. Arf1 and Arf2 share 96% identity and Arf3 and Arf4 share 89% identity, which suggests that the genes arose through gene-duplication events in M. circinelloides. Transcription analysis revealed that certain arf genes are affected by dimorphism of M. circinelloides, such as the arf2 transcript, which was accumulated during yeast development. Therefore, we created knockout mutants of four arf genes to evaluate their function in dimorphism and virulence. We found that both arf1 and arf2 are required for sporulation, but these genes also perform distinct functions; arf2 participates in yeast development, whereas arf1 is involved in aerobic growth. Conversely, arf3 and arf4 play only minor roles during aerobic growth. Moreover, we observed that all single arf-mutant strains are more virulent than the wild-type strain in mouse and nematode models, with the arf3 mutant being most virulent. Lastly, arf1/arf2 and arf3/arf4 double mutations produced heterokaryon strains that did not reach the homokaryotic state, indicating that these genes participate in essential and redundant functions. Overall, this work reveals that Arfs proteins regulate important cellular processes in M. circinelloides such as morphogenesis and virulence, laying the foundation to characterize the molecular networks underlying this regulation.

Comparative genomics applied to Mucor species with different lifestyles

“…In this context, Vongsangnak et al proposed a metabolic network of the oleaginous strain Mucor circicinelloides WJ11 [10,11], Corrochano et al shed new light on Mucor sensory perception [12], and multiple genes potentially involved in virulence were investigated and discovered in Mucor spp. [13][14][15][16][17][18][19][20][21]. Following the annotation of the first Mucor genome sequence (Mucor circinelloides CBS 277.49), researches on Mucor benefited and will continue to benefit from different sequencing projects including the Zygolife initiative (http://zygolife.org/home/) which aims to provide a better phylogenetical classification to the formerly called Zygomycetes which include the Mucor genus (see [1]).…”

Conditional Expression of the Small GTPase ArfA Impacts Secretion, Morphology, Growth, and Actin Ring Position in Aspergillus niger

“…In S. cerevisiae , for example, cognate GAPs for Arf1 include Glo3, Age1/2, and Gcs1 (Poon et al, 1996 , 1999 ; Zhang et al, 1998 ; Suda et al, 2018 ). In addition to the well characterized role of Arfs in budding yeast, these GTPases have also been well studied in mammals (Gillingham and Munro, 2007 ), and there is growing interest in the role of these proteins regulating morphology and virulence during disease, for example in the human infecting dimorphic yeast Candida albicans and the zygomycete Mucor circinelloides (Labbaoui et al, 2017 ; Patiño-Medina et al, 2017 ). Despite comprehensive molecular and genetic characterization of Arfs, their cognate GEFs and GAPs, and the regulatory role of these proteins on multiple processes in yeast and mammalian systems, little is known about their role in Aspergilli, and filamentous fungi in general.…”

Genes, Pathways, and Mechanisms Involved in the Virulence of Mucorales

“…Reverse genetics approaches have identified several virulence factors in Mucorales. The deletion in Mucorales of genes characterized previously in other organisms led to the identification of virulence-reduced phenotypes in mutants for genes involved in iron uptake [17,78], ADP-ribosylation factor (Arf) [79] and Calcineurin [29]. Nowadays, the availability of mucoralean genomes and the knowledge of RNAi mechanisms have allowed strategies of structural and functional genomics to identify a growing number of new virulence factors in these fungi.…”