TTBK2 kinase substrate specificity and the impact of spinocerebellar-ataxia-causing mutations on expression, activity, localization and development

Abstract

Mutations that truncate the C-terminal non-catalytic moiety of TTBK2 (tau tubulin kinase 2) cause the inherited, autosomal dominant, SCA11 (spinocerebellar ataxia type 11) movement disorder. In the present study we first assess the substrate specificity of TTBK2 and demonstrate that it has an unusual preference for a phosphotyrosine residue at the +2 position relative to the phosphorylation site. We elaborate a peptide substrate (TTBKtide, RRKDLHDDEEDEAMSIYpA) that can be employed to quantify TTBK2 kinase activity. Through modelling and mutagenesis we identify a putative phosphate-priming groove within the TTBK2 kinase domain. We demonstrate that SCA11 truncating mutations promote TTBK2 protein expression, suppress kinase activity and lead to enhanced nuclear localization. We generate an SCA11-mutation-carrying knockin mouse and show that this leads to inhibition of endogenous TTBK2 protein kinase activity. Finally, we find that, in homozygosity, the SCA11 mutation causes embryonic lethality at embryonic day 10. These findings provide the first insights into some of the intrinsic properties of TTBK2 and reveal how SCA11-causing mutations affect protein expression, catalytic activity, localization and development. We hope that these findings will be helpful for future investigation of the regulation and function of TTBK2 and its role in SCA11.

Cep164 triggers ciliogenesis by recruiting Tau tubulin kinase 2 to the mother centriole

“…6 E and F). In light of a previous report on the regulation of TTBK2 activity (41), one could also argue that the C-terminal domain of TTBK2 may be required for regulation of kinase activity. Although we cannot rigorously exclude this possibility, we note that our rescue experiments with TTBK2-Cep164 chimeras demonstrate that constructs lacking the C-terminal part of TTBK2 exhibited sufficient kinase activity to restore ciliogenesis in Cep164-depleted cells (Fig.…”

Upregulation of Na+,Cl--Coupled Betaine/ γ-Amino-Butyric Acid Transporter BGT1 by Tau Tubulin Kinase 2

“…TTBK2 [1-450] has been identified in SCA11 patients [34] and leads to decreased kinase activity [7]. Further constructs used were the kinase inactive mutants TTBK2-KD and TTBK2 [1-450] -KD in which an aspartic acid at position 163 was replaced by alanine [D163A].…”

“…Accordingly, loss of function mutations of TTBK2 lead to autosomal dominant spinocerebellar ataxia type 11 (SCA11) [7]. Moreover, TTBK2 may contribute to the resistance of kidney tumor cells and melanoma cells to therapy [6].…”

“…Moreover, TTBK2 may contribute to the resistance of kidney tumor cells and melanoma cells to therapy [6]. Gene targeted mice lacking functional TTBK2 die at embryonic day 10 [7]. …”

Tau Tubulin Kinase TTBK2 Sensitivity of Glutamate Receptor GluK2

“…The tau tubulin kinase 2 (TTBK2) is a serine/threonine kinase [1] underlying the pathophysiology of the rare [2], inherited, autosomal dominant spinocerebellar ataxia type 11 (SCA11) [3,4], a neurodegenerative movement disorder affecting primarily cerebellar neurons [5]. TTBK2 belongs to the CK1 (casein kinase 1) superfamily and is closely related to the kinase TTBK1 which is suspected to play an important role in the pathophysiology of Morbus Alzheimer [6].…”