Abstract
Cytoplasmic aggregation of TDP-43 characterizes degenerating neurons in most cases of amyotrophic lateral sclerosis (ALS). Here, we develop an optogenetic TDP-43 variant (opTDP-43), whose multimerization status can be modulated in vivo through external light illumination. Using the translucent zebrafish neuromuscular system, we demonstrate that short-term light stimulation reversibly induces cytoplasmic opTDP-43 mislocalization, but not aggregation, in the spinal motor neuron, leading to an axon outgrowth defect associated with myofiber denervation. In contrast, opTDP-43 forms pathological aggregates in the cytoplasm after longer-term illumination and seeds non-optogenetic TDP-43 aggregation. Furthermore, we find that an ALS-linked mutation in the intrinsically disordered region (IDR) exacerbates the light-dependent opTDP-43 toxicity on locomotor behavior. Together, our results propose that IDR-mediated TDP-43 oligomerization triggers both acute and long-term pathologies of motor neurons, which may be relevant to the pathogenesis and progression of ALS.
Highlights
Cytoplasmic aggregation of TDP-43 characterizes degenerating neurons in most cases of amyotrophic lateral sclerosis (ALS)
To explore mechanisms of TDP-43 toxicity associated with its cytoplasmic aggregation in spinal motor neurons, we first aimed to induce TDP-43 aggregation by its overexpression in the caudal primary motor neurons (CaPs) of zebrafish, which innervate a ventral third of the myotome and are present uniquely in every spinal hemisegment (Fig. 1a, b)[31]
We injected mRNAs encoding wild-type Tardbp and mRFP1-TDP43z into the TDP-43 double knock-out (DKO) embryos at the one-cell stage and found that the blood circulation defect was rescued by both (Supplementary Fig. 1e, Supplementary Movie 1), indicating that mRFP1-TDP-43 is functional
Summary
Cytoplasmic aggregation of TDP-43 characterizes degenerating neurons in most cases of amyotrophic lateral sclerosis (ALS). OpTDP-43 forms pathological aggregates in the cytoplasm after longer-term illumination and seeds non-optogenetic TDP-43 aggregation. The TDP-43 IDR mutations that are linked to familial ALS cases enhance intrinsic aggregation propensity and protein stability of TDP-4310,11 and result in altered phase separation[9], which could contribute to disease propagation through acceleration of the formation and accumulation of pathological aggregates[12,13,14]. We develop an optogenetic TDP-43 variant (opTDP-43) carrying a light-dependent oligomerization module of cryptochrome-2 attached to the IDR, and analyze the mechanisms of TDP-43 toxicity in spinal motor neurons in vivo. Longer chronic light stimulation eventually leads to accumulation of cytoplasmic opTDP-43 aggregates that further seed aggregation of non-optogenetic TDP-43, which is accompanied by motor decline. The sequential pathological alterations of spinal motor neurons triggered by opTDP-43 oligomerization may provide clues about how motor neuron degeneration progresses at both molecular and cellular levels in a prodromal phase of ALS
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