Spinocerebellar ataxia type 11-associated alleles of Ttbk2 dominantly interfere with ciliogenesis and cilium stability

Abstract

Spinocerebellar ataxia type 11 (SCA11) is a rare, dominantly inherited human ataxia characterized by atrophy of Purkinje neurons in the cerebellum. SCA11 is caused by mutations in the gene encoding the Serine/Threonine kinase Tau tubulin kinase 2 (TTBK2) that result in premature truncations of the protein. We previously showed that TTBK2 is a key regulator of the assembly of primary cilia in vivo. However, the mechanisms by which the SCA11-associated mutations disrupt TTBK2 function, and whether they interfere with ciliogenesis were unknown. In this work, we present evidence that SCA11-associated mutations are dominant negative alleles and that the resulting truncated protein (TTBK2SCA11) interferes with the function of full length TTBK2 in mediating ciliogenesis. A Ttbk2 allelic series revealed that upon partial reduction of full length TTBK2 function, TTBK2SCA11 can interfere with the activity of the residual wild-type protein to decrease cilia number and interrupt cilia-dependent Sonic hedgehog (SHH) signaling. Our studies have also revealed new functions for TTBK2 after cilia initiation in the control of cilia length, trafficking of a subset of SHH pathway components, including Smoothened (SMO), and cilia stability. These studies provide a molecular foundation to understand the cellular and molecular pathogenesis of human SCA11, and help account for the link between ciliary dysfunction and neurodegenerative diseases.

Phosphorylation of multiple proteins involved in ciliogenesis by Tau Tubulin kinase 2

“…These motifs resemble noncanonical CK1 motifs SLS and to some extent also a motif with acidic stretch of residues located C-terminally from the target S/T reported before (29,(66)(67)(68). Importantly, one has to bear in mind that, in contrast to motifs identified in this study and the studies where noncanonical motifs for CK1…”

“…TTBK2 is recruited to DA by CEP164 (26,27), their mutual interaction is finetuned by INNPP5E and PI3Kγ (28). In addition, TTBK2 seems uniquely positioned among the DAassociated proteins -its depletion still allows cilia initiation to proceed to its very final stages unperturbed (vesicles dock to DA) but ciliogenesis fails at the stage when axoneme should start to extend, implying that TTBK2 may function as a switch in cilia assembly pathway, turning cilia initiation programme to cilia elongation and maintenance (25)(26)(27)29).…”

“…TTBK2 mutations leading to truncated proteins are causative for development of neurodegenerative disorder spinocerebellar ataxia type 11 (SCA11) (33)(34)(35). Interestingly, these mutant variants of TTBK2, typically truncated around 450 AA, are unable to interact with CEP164 and fail to localize to DA (25,26), instead seem to act in dominant negative manner to disturb PC formation (25,29). As no other kinase can compensate for defects caused by TTBK2 mutations in SCA11, it again argues that TTBK2 has non-redundant functions.…”

Targeted Depletion of Primary Cilia in Dopaminoceptive Neurons in a Preclinical Mouse Model of Huntington’s Disease

“…Notably, the role of PC in neuronal homeostasis, and in the control of the cellular stress response signaling cascades is emerging in other neurodegenerative disorders. PC pathology has been described for example in Alzheimer's disease (AD), in Parkinson's disease (PD) and in spinocerebellar ataxias, another polyglutamine disease, although distinct mechanisms are probably involved (Steger et al, 2017;Bowie et al, 2018;Dhekne et al, 2018;Vorobyeva and Saunders, 2018). Interestingly, PC were elongated in the hippocampus of the APP/PS1 mouse models of AD compared with wild-type mice, and serotonin 5-HT6 receptors playing a critical role in AD development regulate the morphology and function of neuronal PC (Hu et al, 2017).…”

“…Nevertheless, defective ciliogenesis in striatal cholinergic neurons might impair a protective mechanism involving non-cell autonomous Sonic hedgehog between cholinergic and dopaminergic neurons (Gonzalez-Reyes et al, 2012). The Spinocerebellar ataxia type 11-associated mutation of the serine/threonine kinase Tau tubulin kinase 2 dominantly interferes with ciliogenesis and cilium stability (Bowie et al, 2018). Shortened primary cilium length and dysregulated Sonic hedgehog signaling were also reported in Niemann-Pick type C1 (NPC1) disease, a neurodegenerative lysosomal storage disorder caused by mutations in the NPC1 gene (Canterini et al, 2017).…”