Molecular mechanisms underlying morphological effects of protein kinase C under normal conditions and cellular stress

Detta är en avhandling från Lund University, Faculty of Medicine

Sammanfattning: The protein kinase C (PKC) family of serine/threonine kinases consists of 10-15 members. The PKC isoforms have central roles in many essential cellular processes, including proliferation, apoptosis, differentiation, cytoskeletal changes, and migration. The PKC family can be subgrouped into classical, novel and atypical isoforms depending on structure and sensitivity to the activators diacylglycerol, phorbol esters and Ca2+. We have previously reported of a neurite-inducing effect specific for PKCε, a novel PKC isoform. The effect has been shown to be mediated through the regulatory domain and independently of the catalytic activity of PKCε. Here we identify residues located in the base of the C1b domain important for the neurite-inducing effect of PKCε. We have in an approach to elucidate the mechanisms that mediate PKCε-induced neurite outgrowth performed a screening for proteins that interact with the most potent neurite-inducing PKCε construct. We have identified several novel PKCε-interacting proteins, including the intermediate filament peripherin and several mRNA-binding proteins. Peripherin and three RNA-binding proteins G3BP2, PABPC1, and IGF2BP3 were further investigated regarding their putative function in PKCε-mediated neurite outgrowth. However, no involvement could be detected for any of the investigated interacting proteins. Interactions between peripherin, G3BP2, PABPC1, or IGF2BP3 and PKCε have been confirmed for endogenous proteins. The peripherin-PKCε interaction is mediated by the C1b domain. We have found that PKCε induces peripherin aggregation when the expression levels of peripherin are elevated and that an activation of PKC induces apoptosis of cells overexpressing peripherin. These novel findings can be of importance in understanding the mechanism behind the neurodegenerative disease amyotrophic lateral sclerosis (ALS), where aggregates containing peripherin are essentially always seen in afflicted tissues. The RNA-binding proteins were found to localize to specific RNA-granules, formed when cells are exposed to stress. PKCalpha, rather than PKCε, was found to be the main PKC isoform found in these stress granules. We have furthermore discovered a role for PKCalpha in regulating stress granule formation.