Biology subjects

Egan, M. J.

Publications and source records attributed to Egan, M. J..

3 recordsLinked to original sources

bioRxiv2025

Synchronous spatio-temporal control of autophagy and organelle trafficking is necessary for appressorium-mediated plant infection by Magnaporthe oryzae

The blast fungus Magnaporthe oryzae infects plants using a specialised infection structure called an appressorium that generates physical force to break the rice leaf cuticle. Appressorium development follows a cell cycle-controlled morphogenetic program, requiring autophagy-associated cell death of the fungal spore from which the infection cell develops. How proliferative growth of the fungus is regulated at the same time as programmed cell death, however, is unknown. In this study, we provide evidence that each cell of the conidium undergoes a separate developmental program, which is necessary for plant infection. Using quantitative live-cell imaging, we monitored trafficking of ten organelle types during appressorium morphogenesis in a wild-type M. oryzae strain and isogenic {Delta}atg8 autophagic mutant. High-resolution microscopy using a photoactivatable green fluorescent protein revealed that organelle trafficking occurs from a single conidium cell into the appressorium, while the remaining two cells undergo autophagy. Organelle inheritance operates independently of cell cycle checkpoints but is always associated with spore germination. We furthermore defined the temporal sequence of organelle movement and de novo organelle biogenesis in the incipient appressorium using photoconvertible fluorescent localisation microscopy. Taken together, our study reveals how synchronous spatiotemporal control of autophagy and organelle trafficking is necessary for rice blast infection.

molecular biology↗

bioRxiv2023

Aspergillus SUMOylation mutants have normal dynein function but exhibit chromatin bridges

AbstractFunctions of protein SUMOylation remain incompletely understood in different cell types. The budding yeast SUMOylation machinery interacts with LIS1, a protein critical for dynein activation, but dynein-pathway components were not identified as SUMO-targets in the filamentous fungus Aspergillus nidulans. Via A. nidulans forward genetics, here we identified ubaBQ247*, a loss-of-function mutation in a SUMO-activation enzyme UbaB. Colonies of the ubaBQ247*, {Delta}ubaB and {Delta}sumO mutants looked similar and less healthy than the wild-type colony. In these mutants, about 10% of nuclei are connected by abnormal chromatin bridges, indicating the importance of SUMOylation in the completion of chromosome segregation. Nuclei connected by chromatin bridges are mostly in interphase, suggesting that these bridges do not prevent cell-cycle progression. UbaB-GFP localizes to interphase nuclei just like the previously studied SumO-GFP, but the nuclear signals disappear during mitosis when the nuclear pores are partially open, and the signals reappear after mitosis. The nuclear localization is consistent with many SUMO-targets being nuclear proteins, for example, topoisomerase II whose SUMOylation defect gives rise to chromatin bridges in mammalian cells. Unlike in mammalian cells, however, loss of SUMOylation in A. nidulans does not apparently affect the metaphase-to-anaphase transition, further highlighting differences in the requirements of SUMOylation in different cell types. Finally, loss of UbaB or SumO does not affect dynein-and LIS1-mediated early-endosome transport, indicating that SUMOylation is unnecessary for dynein or LIS1 function in A. nidulans.

cell biology↗

bioRxiv2019

Spatial and temporal alterations in protein structure by EGF regulate cryptic cysteine oxidation

Stimulation of receptor tyrosine kinases (RTK) such as EGF locally increase reactive oxygen species (ROS) levels at the plasma membrane that oxidize cysteines in proteins to enhance downstream signaling. Spatial confinement of ROS is an important regulatory mechanism to redox signaling, but it remains unknown why stimulation of different receptor tyrosine kinases (RTKs) at the plasma membrane target distinct sets of downstream proteins. To uncover additional mechanisms specifying which cysteines are redox regulated by EGF stimulation, we performed time-resolved quantification of the oxidation of 4,200 cysteine sites subsequent to EGF stimulation in A431 cells. EGF induces three distinct spatiotemporal patterns of cysteine oxidation in functionally organized protein networks, consistent with the spatial confinement model. Unexpectedly, protein crystal structure analysis and molecular dynamic simulation indicate widespread redox regulation of cryptic cysteines that are only solvent exposed upon changes in protein conformation. Phosphorylation and increased flux of nucleotide substrates serve as two distinct modes by which EGF specifies which cryptic cysteines become solvent exposed and redox regulated. Since proteins structurally regulated by different RTKs or cellular perturbations are largely unique, solvent exposure and redox regulation of cryptic cysteines is an important mechanism contextually delineating redox signaling networks.\n\nSignificance StatementCellular redox processes are interconnected, but are not in equilibrium. Thus, understanding the redox biology of cells requires a systems-level, rather than reductionist, approach. Factors specifying which cysteines are redox regulated by a stimulus remain poorly characterized but are critical to understanding the fundamental properties of redox signaling networks. Here, we show that EGF stimulation induces oxidation of specific cysteines in 3 distinct spatiotemporal patterns. Redox regulated proteins include many proteins in the EGF pathway as well as many cysteines with known functional importance. Many redox regulated cysteines are cryptic and solvent exposed by changes in protein structure that were induced by EGF treatment. The novel finding that cryptic cysteines are redox regulated has important implications for how redox signaling networks are specified and regulated to minimize crosstalk. In addition, this time-resolved dataset of the redox kinetics of 4,200 cysteine sites is an important resource for others and is an important technological achievement towards systems-level understanding of cellular redox biology.

biochemistry↗