Investigating signaling network evolution phenomena in the fungus Magnaporthe oryzae

Research in evolutionary dynamics has demonstrated that – apart from bacteria – also eukaryotic organisms like fungi can undergo rapid evolutionary changes within a "very short" timeframe. The molecular mechanisms driving these rapid adaptations remain poorly understood, largely due to their dependence on rare and unpredictable changes. Adaptive evolution in microorganisms typically involves advantageous genetic, transcriptomic, or proteomic changes that persist through natural selection. Understanding these mechanisms is crucial for addressing global challenges such as the emergence of new pathogens, the spread of invasive species, and the development of multi-drug resistance.

Within this work the rice blast pathogen Magnaporthe oryzae is used to highlight the complexity of evolution in regulatory networks of physiological and biochemical processes. The High Osmolarity Glycerol (HOG) pathway regulates cellular adaptation to environmental osmolarity. Loss-of-function (lof) mutants of the HOG pathway are osmosensitive and fail to produce the main critical osmotic stress response solute arabitol as it is in the wildtype strain. Interestingly, when these lof mutants are exposed to constant osmotic pressure, stable suppressor strains emerge that produce high amounts of the osmolyte glycerol instead of arabitol.

This study aimed to identify the genes responsible for adaptation to long-term stress and the shift from arabitol to glycerol production. Candidate genes were identified and their roles in primary metabolite production. Within this work an effective bioinformatic pipeline for label-free quantification of proteome and phosphoproteome data was developed. Using M. oryzae as a model, a data-independent acquisition (DIA) approach was developed, significantly improving the quality and completeness of the data. This method reduced the LC-MS/MS analysis time and increased the identification of phosphosites, establishing a refined methodology and a comprehensive basis for studying signaling processes in filamentous fungi. Within this method, a new set of candidate genes involved in the adaptation process was identified. These findings contribute to a deeper understanding of the complex evolutionary mechanisms in M. oryzae and underscore the need for continued research to unravel the molecular basis of rapid evolutionary adaptations in microorganisms.