The role of the lissencephaly protein Pac1 during nuclear migration in budding yeast

During mitosis in Saccharomyces cerevisiae, the mitotic spindle moves into the mother–bud neck via dynein-dependent sliding of cytoplasmic microtubules along the cortex of the bud. Here we show that Pac1, the yeast homologue of the human lissencephaly protein LIS1, plays a key role in this process. First, genetic interactions placed Pac1 in the dynein/dynactin pathway. Second, cells lacking Pac1 failed to display microtubule sliding in the bud, resulting in defective mitotic spindle movement and nuclear segregation. Third, Pac1 localized to the plus ends (distal tips) of cytoplasmic microtubules in the bud. This localization did not depend on the dynein heavy chain Dyn1. Moreover, the Pac1 fluorescence intensity at the microtubule end was enhanced in cells lacking dynactin or the cortical attachment molecule Num1. Fourth, dynein heavy chain Dyn1 also localized to the tips of cytoplasmic microtubules in wild-type cells. Dynein localization required Pac1 and, like Pac1, was enhanced in cells lacking the dynactin component Arp1 or the cortical attachment molecule Num1. Our results suggest that Pac1 targets dynein to microtubule tips, which is necessary for sliding of microtubules along the bud cortex. Dynein must remain inactive until microtubule ends interact with the bud cortex, at which time dynein and Pac1 appear to be offloaded from the microtubule to the cortex

Extraction and purification of DNA from wood at various stages of decay for metabarcoding of wood-associated fungi

Assessment of endophytic and saprotrophic microbial communities from wood-extracted DNA presents challenges due to the presence of surface microbes that contaminate samples and plant compounds that act as inhibiting agents. Here, we describe a method for decontaminating, sampling, and processing wood at various stages of decay for high-throughput extraction and purification of DNA

Initial wood trait variation overwhelms endophyte community effects for explaining decay trajectories

Microbial organisms, environmental conditions and their interactions govern many ecosystem processes. Recent studies have highlighted the importance of priority effects, that is, the identity of potential decomposers present early in community assembly, in determining resulting decay rates especially for wood. In diverse forests, available woody substrates differ chemically and structurally with implications for their role as both habitats and resources for microbes. Both wood traits and microbial communities at the start of the decay process affect subsequent decay rates, but the relative magnitude of effects is not known. In this work, we sought to ask a simple question: what are the relative effects of microbial communities and wood traits? We characterized fungal and oomycete endophytes with amplicon sequencing from stems of 22 woody species growing in woodlands near Richmond, NSW, Australia, and measured 11 traits to capture variation in the physical and chemical wood substrates. To evaluate the consequence of endophyte diversity and wood traits on the trajectory of decay, stem samples were sequentially harvested over 5 years to quantify the decay rate, its consistency and how it varies through time. We did not find evidence to support particular initial endophyte compositions leading to faster decay. Instead, initial wood attributes were much more powerful in explaining decay trajectories with smaller, less dense stems with high water, low N and low lignin concentrations decomposing consistently faster. These data show that initial wood traits have long-lasting consequences on decay unlike natural variation in endophyte communities, supporting the idea that community member functions are highly redundant. Wood substrate-driven environmental filtering, rather than endophyte-driven priority effects, had a stronger effect on decay when real-world levels of diversity in wood traits were considered. Read the free Plain Language Summary for this article on the Journal blog