Poster number 103 submitted by Sean Duffy
Upf1 is required for translational response to oxidative stress in Cryptococcus neoformans
Sean R. Duffy (Department of Microbiology and Immunology, SUNY Buffalo), John C. Panepinto (Department of Microbiology and Immunology, SUNY Buffalo)
Nonsense-mediated decay (NMD) is an evolutionarily conserved pathway that is important for mRNA quality control and is initiated by the RNA helicase Upf1. NMD targets mRNAs which possess premature termination codons (PTCs) resulting from DNA damage, RNA damage, transcriptional errors, or PTCs programmed into the mRNA sequence. The fungal pathogen Cryptococcus neoformans must adapt to the host-lung environment, which includes temperature and oxidative stress. In response to stress, C. neoformans represses translation by decreasing translation initiation, clearing homeostatic mRNAs, and promoting the translation of mRNAs necessary for stress adaptation. Previous studies have investigated the role of Upf1 in the retention of introns and the control of translation through start codon context in C. neoformans. However, the role of Upf1 during stress adaptation remains unknown. Utilizing a upf1Δ mutant we found that translation is more readily repressed by oxidative stress in the absence of NMD. In polysome profiles, the upf1Δ mutant showed a greater decrease in high molecular weight complexes than the WT strain after exposure to hydrogen peroxide. Western blotting also revealed that upf1Δ displays increased eIF2α phosphorylation, a known hallmark for global translation inhibition, at lower peroxide concentrations. Northern blot assessment of the ribosomal protein transcript, RPL2, also demonstrated slower decay of homeostatic transcripts in the upf1Δ mutant following oxidative stress. Upf1 along with the other core NMD factors Upf2 and Upf3 are conserved in both humans and yeast. However, the additional cofactors involved in targeting NMD-regulated transcripts for decay vary. BLAST analyses revealed two genes, CNAG_05233 and CNAG_07351, in C. neoformans which encode for proteins that have sequence similarity and domain conservation to the human NMD cofactors, hSmg5/7, and hSmg6, respectively, than to the yeast NMD cofactors, Ebs1p and Nmd4p, respectively. Future work will aim to explore the NMD complex in C. neoformans and understand the mechanism by which Upf1 contributes to translational regulation and translatome reprogramming in response to oxidative stress.
Keywords: Translation, NMD, Stress