The role of microtubules and myosin II in the formation of Hirano bodies

Abstract

Hirano bodies are intracellular, paracrystalline, F-actin rich, structures that are most commonly found in the autopsied brains of humans suffering from neurodegenerative diseases. However, their biochemistry and physiology are not well understood despite their association with aging and several diseases. Recently, an in vitro and in vivo model to induce formation of Hirano bodies in living cells by expression of various mutants of a 34 kDa actin bundling protein mutant (Maselli et, al., 2002, 2003; Davis et, al., 2008) was developed. This allows further questions concerning Hirano bodies to be investigated. To examine what is necessary for the formation of Hirano bodies, we investigated the role of microtubules and an actin-dependent motor protein, myosin II. An inducible promoter for Hirano body formation fused to green fluorescent protein (pVEII-E60K-GFP) was introduced in Dictyostelium discoideum amoeba in the presence of pharmaceuticals. Cells were treated with nocodazole, a microtubule depolymerizing drug, or blebbistatin, a myosin II ATPase inhibitor. Cells were analyzed via fluorescence microscopy. After 24 hours of induction, no statistical difference in Hirano body size was seen between control cells and those treated with nocodazole, but blebbistatin-treated cells contained significantly smaller Hirano bodies, bust still retained some large Hirano bodies. This suggests that myosin II plays a role in the formation of Hirano bodies but is not essential. Further experiments are needed to elucidate what components of the cytoskeleton are absolutely essential for the formation of Hirano bodies.