Biology subjects

Magalhaes, A. C.

Publications and source records attributed to Magalhaes, A. C..

2 recordsLinked to original sources

bioRxiv2023

Fusarium graminearum Ste2 and Ste3 Receptors Undergo Peroxidase Induced Heterodimerization

F. graminearum FgSte2 and FgSte3 are G-protein coupled receptors (GPCRs) recently shown to play roles in mediating fungal hyphal chemotropism and plant pathogenesis in response to activity arising from host-released peroxidases. Here, we follow up on the previous observation that chemotropism is dependent on both FgSte2 and FgSte3 being present at the same time; testing the possibility that this effect might be due to formation of an FgSte2-FgSte3 heterodimer. Initially the recombinant cell-surface expression of the F. graminearum GPCRs was validated in S. cerevisiae by confocal immunofluorescence microscopy. Bioluminescence resonance energy transfer analyses were subsequently conducted, where the addition of horse radish peroxidase (HRP) was found to increase the transfer of energy from the inducibly-expressed FgSte3-Nano luciferase (FgSte3-NLuc) donor, to the constitutively-expressed FgSte2-yellow fluorescent protein (FgSte2-YFP) acceptor, compared to controls. A partial response was also detected when an HRP-derived ligand-containing extract was enriched from F. graminearum spores and applied to the S. cerevisiae BRET system directly. The selectivity of the interaction was demonstrated by comparison to treatment with pheromones as well as an unrelated bovine GPCR, rhodopsin, fused to YFP as acceptor, that yielded no response when co-expressed with FgSte3-NLuc. Finally, the peroxidase-stimulated heterodimerization was validated by affinity pulldown. Taken together these findings demonstrate the formation of HRP and HRP-derived ligand stimulated heterodimers between FgSte2 and FgSte3. Outcomes are discussed from the context of the roles of ligands and reactive oxygen species in GPCR dimerization.

biochemistry↗

bioRxiv2022

Human cytomegalovirus vMIA inhibits MAVS oligomerization at peroxisomes in an MFF-dependent manner

Upon intracellular recognition of viral RNA, RIG-I-like proteins interact with MAVS at peroxisomes and mitochondria, inducing its oligomerization and the downstream production of direct antiviral effectors. The human cytomegalovirus (HCMV) is able to specifically evade this antiviral response, via its antiapoptotic protein vMIA. Besides suppressing the programmed cell death of infected cells, vMIA inhibits the antiviral signalling at mitochondria by inducing the organelles fragmentation, consequently hindering the interaction between MAVS and the endoplasmic reticulum protein STING. Here, we demonstrate that vMIA interferes with the peroxisomal antiviral signalling via a distinct mechanism that is independent of the organelles morphology and does not affect STING. vMIA interacts with MAVS at peroxisomes and inhibits its oligomerization, restraining downstream signalling, in an MFF-dependent manner. This study also demonstrates that vMIA is totally dependent on the organelles fission machinery to induce peroxisomal fragmentation, while this dependency is not observed at mitochondria. Furthermore, although we demonstrate that vMIA is also able to inhibit MAVS oligomerization at mitochondria, our results indicate that this process, such as the whole vMIA-mediated inhibition of the mitochondrial antiviral response, is independent of MFF. These observed differences in the mechanisms of action of vMIA towards both organelles, likely reflect their intrinsic differences and roles throughout the viral infection. This study uncovers specific molecular mechanisms that may be further explored as targets for antiviral therapy and highlights the relevance of peroxisomes as platforms for antiviral signalling against HCMV.

cell biology↗