Glycogen Synthase Kinase (GSK) 3β Directly Phosphorylates Serine 212 in the Regulatory Loop and Inhibits Microtubule Affinity-regulating Kinase (MARK) 2

Abstract

MARK/Par-1, a kinase family with diverse functions particularly in inducing cell polarity, can phosphorylate microtubule-associated proteins in their repeat domain and cause their detachment from microtubules, and thereby microtubule destabilization. Because of its role in abnormal phosphorylation of the Tau protein in Alzheimer disease, we searched for regulatory kinases. MARK family kinases can be activated by phosphorylation of a conserved threonine (Thr-208 in MARK2), and inactivated by phosphorylation of a serine (Ser-212), both in the activation loop of the catalytic domain. Activation is achieved by the kinases MARKK/TAO1 or LKB1, although the inactivating kinase was unknown. We show here that GSK3␤ serves the role of the inhibitory kinase. Because GSK3␤ can also phosphorylate Tau at sites outside the repeat domain, the activation of GSK3␤, and concomitant inactivation of MARK can shift the pattern of pathological phosphorylation of Tau protein in Alzheimer disease.

LRRK2G2019S Mutation Induces Dendrite Degeneration through Mislocalization and Phosphorylation of Tau by Recruiting Autoactivated GSK3β

“…The involvement of GSK3␤ in PD development is substantiated by a recent study that PD risk is associated with genetic polymorphisms of GSK3␤ (Kwok et al, 2005). Knowing that activities of tau kinases are also regulated by phosphorylation events (Kim and Kimmel, 2006;Timm et al, 2008), LRRK2 disease mutations may affect tau phosphorylation through upstream kinases such as GSK3␤, thereby contributing to PD pathogenesis.…”

Microtubule Affinity Regulating Kinase Activity in Living Neurons Was Examined by a Genetically Encoded Fluorescence Resonance Energy Transfer/Fluorescence Lifetime Imaging-based Biosensor

“…9). The activity of MARK had been shown in the tips of neurites in differentiated and fixed PC12 cells by staining with antibodies specific for active MARK (Thr(P)-208) and for its phosphorylated primary substrate Tau (Ser(P)-262) (19). However, the reporter now allowed us to observe MARK activity in living cells in a timeresolved manner and to follow the response to a variety of cues, such as activators or treatments with kinase inhibitors.…”

“…In cells the catalytic activity of MARK is regulated by intramolecular domain interactions (e.g. ubiquitin-associated domain and kinase associated domains), phosphorylation by other kinases (activating: MARKK, LKB1 (17,18); inhibiting: GSK3␤ (19)), interactions with other cytoskeleton-directed kinases (PAK5 (p21-activated kinase 5), TESK (testis-specific kinase) (20,21)), or by interactions with scaffold proteins such as 14-3-3 (22).…”

Aβ Oligomers Cause Localized Ca2+Elevation, Missorting of Endogenous Tau into Dendrites, Tau Phosphorylation, and Destruction of Microtubules and Spines

“…Cells were fixed in 3.7% formaldehyde for 20 min, permeabilized for 6 min in 80% ice-cold methanol and incubated with the following antibodies (for details, see above): K9JA (DAKO Tau-A0024, 1:500); DA9 (mAb against Tau, gift from P. Davies, 1:200); mAb AP20 against MAP2 (Sigma; 1:100); polyclonal anti-neurofilament 200 (Sigma, 1:100) polyclonal amyloid-␤ (Cell Signaling Technology; 1:200); mAb 12E8 (1:500); mAb AT8 (1:100); polyclonal MARK phospho-T208 [prepared as described by Timm et al (2008); 1:100]; phosphop70S6Kinase (pT389, clone 108D2, Cell Signaling Technology; 1:50); polyclonal BRSK1 (Acris, 1:100); phospho-MAPK (Cell Signaling Technology; 1:50); polyclonal phospho-JNK1 ϩ 2 (pT183ϩpY185, Abcam; 1:50); phospho-GSK3␤ (Cell Signaling Technology, 1:50); phospho-CDK5 (pS159, Eurogentec; 1:100); actin (Sigma, clone AC-40, 1:500). Fluorescently labeled secondary antibodies (Cy2, Cy3, Cy5) were from Dianova.…”