Effect of Rapamycin on Aspergillus nidulans Cell Wall Material Properties

Filamentous fungi play vital roles in human health, agriculture and bioprocessing. In all of these situations fungi are often exposed to nutrient limitation, which can impact their behavior. Autophagy is a eukaryotic cellular process, induced under nutrient deficient conditions, which recycles internal components for cell survival. Autophagy can also be gratuitously induced in rich growth medium, using the drug rapamycin. To address our hypotheses, we have studied fungi and cell wall material properties of the model fungus, Aspergillus nidulans, under autophagic conditions. For example, it has been observed that during fungal fermentation nutrient limitation leads to a greater degree of hyphal fragmentation. We hypothesize that under nutrient deprived conditions autophagy is involved in recycling cell wall components, thereby weakening the wall and resulting in this increased degree of fragmentation. In all experiments, fungi were subjected to two media conditions - with and without rapamycin. To understand changes in the cell wall material properties we studied changes in cell wall tensile strength using a fragmentation assay, changes in cell wall thickness using transmission electron microscopy, and changes in cell wall elastic properties using atomic force microscopy (AFM). The fragmentation assay showed that cells grown in presence of rapamycin for 16 hours were stronger than the cells grown in the absence of rapamycin. More data is required to determine whether the observed changes in cell wall strength are due to changes in cell wall thickness. Typical techniques used to study the changes in cell wall material properties are tedious, time intensive and only measure the population average. As an alternative, we propose the use of Atomic Force Microscope (AFM) to observe changes at the single mycelium level. Previously, our lab developed a method to study fungi grown in suspension culture, using AFM and we used this method to study cells grown on cover slips enabling us to study individual mycelia.