Tau missorting and spastin-induced microtubule disruption in neurodegeneration: Alzheimer Disease and Hereditary Spastic Paraplegia

Abstract

In Alzheimer Disease (AD), the mechanistic connection of the two major pathological hallmarks, namely deposition of Amyloid-beta (Aβ) in the form of extracellular plaques, and the pathological changes of the intracellular protein Tau (such as phosphorylation, missorting, aggregation), is not well understood. Genetic evidence from AD and Down Syndrome (Trisomy 21), and animal models thereof, suggests that aberrant production of Aβ is upstream of Tau aggregation, but also points to Tau as a critical effector in the pathological process. Yet, the cascade of events leading from increased levels of Aβ to Tau-dependent toxicity remains a matter of debate.Using primary neurons exposed to oligomeric forms of Aβ, we have found that Tau becomes mislocalized (missorted) into the somatodendritic compartment. Missorting of Tau correlates with loss of microtubules and downstream consequences such as loss of mature spines, loss of synaptic activity, and mislocalization of mitochondria.In this cascade, missorting of Tau induces mislocalization of TTLL6 (Tubulin-Tyrosine-Ligase-Like 6) into the dendrites. TTLL6 induces polyglutamylation of microtubules, which acts as a trigger for spastin mediated severing of dendritic microtubules. Loss of microtubules makes cells unable to maintain transport of mitochondria, which in turn results in synaptic dysfunction and loss of mature spines. These pathological changes are absent in TauKO derived primary neurons. Thus, Tau mediated mislocalization of TTLL6 and spastin activation reveals a pathological gain of function for Tau and spastin in this cellular model system of AD.In contrast, in hereditary spastic paraplegia (HSP) caused by mutations of the gene encoding spastin (spg4 alias SPAST), spastin function in terms of microtubule severing is decreased at least for the gene product of the mutated allele, resulting in overstable microtubules in disease model systems. Whether total spastin severing activity or microtubule stability in human disease is also affected is not yet clear. No human disease has been associated so far with the long-chain polyglutamylation enzyme TTLL6, or the other TTLLs (1,5,11) possibly involved.Here we review the findings supporting a role for Tau, spastin and TTLL6 in AD and other tauopathies, HSP and neurodegeneration, and summarize possible therapeutic approaches for AD and HSP.

Fig. 1

Aβ oligomers induce missorting of Tau into dendrites. Missorting of Tau correlates with dendritic loss of microtubules and mislocalization of TTLL6. a, b Micrographs of fluorescently stained primary hippocampal neurons aged for 21 days in vitro after treatment with Aβ oligomers. (a) Tau is stained with an antibody against total Tau (K9JA, green color). Staining of the dendritic marker MAP2 is shown in red. In control conditions, Tau and MAP2 do not colocalize (a1, boxed area shows a dendrite and cell body without Tau). After exposure to Aβ, Tau becomes missorted into the somatodendritic compartment, where it then colocalizes with MAP2 (a2, boxed area shows a dendrite and cell body with Tau missorting). (b) Staining of f-actin in dendritic segments with phalloidin. In control conditions, there are numerous spines (b1, f-actin positive dendritic protrusions, arrowheads). After exposure to Aβ, mature spines are lost (b2). c Microtubules were stained using an antibody against tubulin after fixing and extracting to favor microtubules over free tubulin. The filamentous structures within the dendrite in c1 (framed by dotted lines) are microtubules. In control conditions dendrites display dense microtubules (c1), whereas microtubules are lost after Aβ treatment (c2). Proximal parts of dendritic segments are magnified in lower panels. cb: cell body; d: dendrites. d Brain sections from Tau transgenic mice where missorting of Tau is present, TTLL6 is mislocalized to dendrites even without exposure to Aβ. Boxed areas highlight examples of dendritic segments. Adapted from [78]

Fig. 2

Spastin executes microtubule loss after invasion of dendrites by Tau and TTLL6. a Experimental evidence for spastin mediated microtubule breakdown in a neuronal cell model of AD. Spastin was silenced using shRNA with a vector co-expressing mRFP in primary hippocampal neurons aged 16 days in vitro. Cells were then treated with 1 μM Aβ for 3 h. Silencing of spastin results in stable microtubules after Aβ treatment. Cells expressing shRNA show no microtubule reduction in dendrites (arrows, a1). Neighboring untransfected cells with normal spastin levels show loss of microtubules in dendrites after Aβ treatment (a2, arrowheads). b, c Diagrams of proposed mechanisms for microtubule breakdown in AD and overstable microtubules in HSP. (b) In Alzheimer disease and other tauopathies, spastin mediates microtubule disassembly in dendrites in the presence of missorted Tau. (c) In the case of HSP caused by mutations of sp4/SPAST, spastin is unable to sever microtubules, but might still be able to bind them, resulting in overstable and bundled microtubules. Modified from [32, 78]

Fig. 3

Spastin domain structure, and proposed mechanism of Tau mediated microtubule loss. a Domain structure of spastin. Spastin contains four main domains: TM, transmembrane domain; MIT, microtubule interacting and transport domain, MTBD, microtubule binding domain; and AAA, ATPase associated with various cellular activities domain. Adapted from [56, 60]. b Severing of microtubules by spastin and subsequent disassembly. MTs are fluorescently labeled (Atto488) and stabilized by taxol. Yellow arrowheads indicate areas where microtubules are severed by spastin. Figure by courtesy of G. Woehlke (Tech. Univ. Munich), adapted from [17]. For movies of spastin-induced severing of microtubules see [17]. c Model of Tau dependent spastin mediated severing of microtubules: (1) Missorted Tau induces translocation of TTLL6 to dendritic microtubules. (2) Tau detaches from microtubules (i.e. has a reduced dwell time on microtubules, possibly as a result of increased phosphorylation by MARK). (3) TTLL6 executes polyglutamylation of microtubules which triggers the activation of spastin and severing of microtubules