Dynamics of a Kinetochore Protein Dam1 during pathogenic development in the Rice-Blast fungus

Hiral Shah Rajesh Patkar Johannes Manjrekar
Bharat Chattoo Genome Research Centre, Department of Microbiology & Biotechnology Center, The Maharaja Sayajirao University of Baroda, Vadodara, India

Filamentous pathogenic fungi exhibit multiple morphological forms, associated with changes in cell polarity. Development in the rice-blast fungus Magnaporthe oryzae during vegetative hyphal growth proceeds by apical extension and lateral branching, while pathogenic development requires polarised elongation of the germ tube followed by an isotropic expansion of its tip into the infection structure called appressorium. These developmental transitions are tightly regulated by correct mitotic progression, nuclear segregation and migration. The kinetochore is a multi-protein complex that brings about high-fidelity nuclear segregation during anaphase. The fungus-specific outer kinetochore protein Dam1 plays a crucial role in maintaining the spindle structure, and thus in proper nuclear segregation and viability in yeasts. In addition to kinetochore localisation, Dam1 is also seen associated with microtubules in fission yeast. In vitro studies with S. cerevisiae proteins show Dam1 complex as oligomeric rings and patches around microtubules. However, the requirement of Dam1 function for viability and its localisation during cell cycle progression vary among different yeast species. Here, we found that the GFP-tagged Dam1 puncta(e) localise to the nucleus at the onset of mitosis, and intensify during nuclear segregation and migration in the filamentous fungus M. oryzae. Further, loss of Dam1 function (dam1Δ) led to prolonged mitosis with sluggish nuclear movement and difficulty in segregation, resulting in delayed and aberrant appressorium formation. Interestingly, GFP-Dam1 also localised to the tip of the growing hyphae or germ tubes during interphase, suggesting its role in cell polarity. The dam1Δ mutant showed reduced hyphal growth and aberrant hyphal morphology. Thus, our findings indicate that Magnaporthe Dam1, in addition to its role in nuclear division, likely has a broader function in the dynamics of the microtubular network and/or cell polarity even during interphase.