The microtubule-affinity regulating kinase (MARK) proteins are a family of Ser/Thr kinases involved in controlling cytoskeletal dynamics and maintaining cell polarity. A primary function of MARK2 is to phosphorylate microtubule-associated proteins such as MAP2, MAP4 and tau within their microtubule-binding repeat (R) domains. In neurons, tau preserves the structural integrity of the cytoskeleton by binding and stabilizing microtubules. The phosphorylated state of tau plays an important role in regulating its interaction with tubulin; tau that is phosphorylated dissociates from tubulin, leaving microtubules in highly dynamic states. Under pathological conditions, tau proteins can become abnormally phosphorylated, leading to loss of microtubule stability and neuronal cell death. Indeed, hyper-phosphorylated tau isoforms have been implicated in the pathogenesis of neurodegenerative disorders such as Alzheimer’s disease and frontotemporal dementia. In this study, our objective was to develop a peptide-based kinase inhibitor of MARK2 function. This peptide (designated tR1) was designed as a direct sequence mimic of the tau R1 domain to selectively inhibit the MARK2-mediated phosphorylation of tau proteins <i>in vitro</i> and in cultured primary neurons. <i>In vitro</i> experiments showed that tR1 inhibits the MARK2-mediated phosphorylation of Ser262 within the tau R1 microtubule-binding domain. It was further demonstrated that tR1 is internalized by rat primary cortical neurons and can be delivered to the cytoplasm when co-treated with small-molecule endosome disruptors such as bafilomycin A1 or chloroquine. More critically, the tR1 peptide was found to inhibit the phosphorylation of endogenous tau at Ser262 in rat primary cortical neurons displaying hyper-active MARK2. Finally, the effects of tR1 were found to be selective for inhibiting tau phosphorylation at MARK2-dependent sites, as tR1 did not interfere with the activity of other kinases, such as GSK-3β, that phosphorylate tau at Thr231. Importantly, these results have established tR1 as a novel peptide-based kinase inhibitor of MARK2 function that has significant therapeutic potential and the promising capacity to be developed as a tool to dissect the complex nature of MARK:tau biology.

Reducing tau phosphorylation using synthetic peptides: Developing peptide-based inhibitors of microtubule affinity regulating kinase 2 (MARK2)

256th National Meeting (2018)

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