Date on Senior Honors Thesis
Senior Honors Thesis
College of Arts and Sciences
GFP; mCherry; mitochondrial inheritance; mitochondria; pathogenic fungi; confocal microscopy
Sporisorium reilianum is a dimorphic fungus that inhabits and infects a host corn plant (Zea mays). In order for the fungus to reproduce sexually, compatible haploid mating types must form a dikaryon that goes on to cause infection in the host. This infection causes leaf chlorosis and gall formation, while ultimately allowing for the dispersal of fungal teliospores in the later stages of infection. To grow, the fungus requires energy production in the form of ATP from its mitochondria. As a countermeasure to infection, host plants release harsh reactive oxygen species that may damage DNA, lead to apoptosis, and reduce the ability of organisms to produce ATP via the electron transport chain in the mitochondrion. The fungus may survive this assault by utilizing an alternative oxidase, which gives the fungi an alternative pathway in the electron transport chain. Thus, alternative oxidase (Aox) is a necessary part of the fungal mitochondria and will be highly expressed at some points during its life cycle. This leads to the unique opportunity to make use of the presence of Aox in the mitochondria and tag the protein fluorescently to track it, and also mitochondrial dynamics, throughout the fungal lifecycle. Fluorescent tagging will allow for a better understanding of fungal mitochondria and what happens to the mitochondria of fungi when sexual reproduction occurs between different mating types. This study serves to investigate the utility of fluorescently tagging Aox for the purpose of better understanding these mating-type interactions. Two mating partners 5-1 and 5-2 of S. reilianum were tagged and observed using confocal microscopy. The strains were then crossed and fusion was observed between the 5-1 mCherry #2 and 5-2 eGFP #6 strains.
Schroeder, Luke, "Using fluorescent microscopy to follow mitochondrial Inheritance through tagged alternative oxidase in sporisorium reilianum." (2022). College of Arts & Sciences Senior Honors Theses. Paper 276.
Retrieved from https://ir.library.louisville.edu/honors/276
A protein known as Aox (alternative oxidase) that is located in the inner mitochondrial membrane of Sporisorium reilianum cells was tagged with a fluorescent protein using molecular genetic techniques. Strains of the fungus with this modified protein were then observed using confocal microscopy which allowed for tracking of the fluorescent mitochondria during sexual reproduction.