Evidence for the conservation of circadian rhythms in the fungal order Diaporthales.

Slupski, M.*, Naidoo, S., Van der Merwe, N. A.

Department of Biochemistry, Genetics and Microbiology, Forestry and Agricultural Biotechnology Institute, University of Pretoria, Pretoria, South Africa

Circadian rhythms constitute an organism’s ability to anticipate and respond to regularly changing environmental stimuli. These endogenous rhythms are controlled by a network of circadian rhythm proteins. In fungi, this network is well conserved and involves a central transcriptional/translational negative feedback loop. One of the most important core genes in this loop is frq, which works mainly as the negative arm of the feedback loop. As a result, its gene expression varies cyclically throughout a 24-hour day and acts as an endogenous timekeeper in response to external rhythms such as light/dark cycles, temperature or nutrient conditions. Recent research showed that circadian rhythms influence the virulence of fungi, which highlights a need for research into the circadian clocks of pathogenic fungi. The fungal pathogens in the genus Chrysoporthe are good candidates for such studies, due to their importance in the infection of economically important Eucalyptus trees in Southern Africa, and their use as a model species to study tree-pathogen interactions. We aimed to characterise the evolution of circadian rhythm genes within Chrysoporthe species and in the order Diaporthales to which they belong. Additionally, we aimed to identify the presence of circadian rhythmicity within the model species Chrysoporthe austroafricana. The evolution of these genes was investigated via phylogenetic and protein domain analyses. Functional domains were highly conserved within the order Diaporthales, providing good evidence for the functional conservation of circadian rhythms within this order of fungi. Following these results, circadian rhythmicity in C. austroafricana was assessed by analysing gene expression of frq via qPCR over a period of 48 hours under constant conditions. From this work, we have established a foundation for future research into the impact of circadian rhythms on pathogenic fungi, their hosts, and on how experimental design can be improved to better take changing environmental conditions into account.

Keywords: circadian rhythm; Diaporthales; phylogenetics; plant pathogenic fungi