Assays for genetic dissection of septin filament assembly in yeast, from de novo folding through polymerization

In Saccharomyces cerevisiae, septin mutations have severe effects on colony-forming ability, particularly at high temperatures, allowing the full variety of genetic tools available in this model organism to be applied to the study of septin biology. Although many details of septin function remain unknown, one can exploit a small number of easily scored phenotypes-proliferation capacity, cell morphology, septin localization, and septin ring integrity-as sensitive readouts of properly assembled septin filaments. Accordingly, this chapter focuses on genetic approaches targeted toward understanding the molecular mechanisms of de novo septin folding, heterooligomerization, and polymerization into filaments. The same general methods can be used to interrogate septin function, although interpretation of results can be more complicated. As genetic-based methodologies are technically simple but particularly dependent on interpretation, here I focus on the logic underlying the most common interpretations of results using septin mutants.