Tau-tubulin kinase 1 phosphorylates TDP-43 at disease-relevant sites and exacerbates TDP-43 pathology.

TDP-43 pathology is a hallmark of Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal lobar degeneration (FTLD). Namely, both diseases feature aggregated and phosphorylated TDP-43 containing inclusions in the cytoplasm and a loss of nuclear TDP-43 in affected neurons. It has been reported that tau tubulin kinase (TTBK)1/2 phosphorylate TDP-43 and TTBK1/2 overexpression induced neuronal loss and behavioral deficits in a C. elegans model of ALS. Here we aimed to elucidate the molecular mechanisms of TTBK1 in TDP-43 pathology. TTBK1 levels were observed to be elevated in ALS patients' post-mortem motor cortex. Also, TTBK1 was found to phosphorylate TDP-43 at disease-relevant sites in vitro directly, and this phosphorylation accelerated TDP-43 formation of high molecular species. Overexpression of TTBK1 in mammalian cells induced TDP-43 phosphorylation and the construction of high molecular species, concurrent with TDP-43 mis-localization and cytoplasmic inclusions. In addition, when TTBK1 was knocked down or pharmacologically inhibited, TDP-43 phosphorylation and aggregation were significantly alleviated. Functionally, TTBK1 knockdown could rescue TDP-43 overexpression-induced neurite and neuronal loss in iPSC-derived GABAergic neurons. These findings suggest that phosphorylation plays a critical role in the pathogenesis of TDP-43 pathology and that TTBK1 inhibition may have therapeutic potential for the treatment of ALS and FTLD.

Loss of RAD-23 Protects Against Models of Motor Neuron Disease by Enhancing Mutant Protein Clearance.

Misfolded proteins accumulate and aggregate in neurodegenerative disease. The existence of these deposits reflects a derangement in the protein homeostasis machinery. Using a candidate gene screen, we report that loss of RAD-23 protects against the toxicity of proteins known to aggregate in amyotrophic lateral sclerosis. Loss of RAD-23 suppresses the locomotor deficit of Caenorhabditis elegans engineered to express mutTDP-43 or mutSOD1 and also protects against aging and proteotoxic insults. Knockdown of RAD-23 is further neuroprotective against the toxicity of SOD1 and TDP-43 expression in mammalian neurons. Biochemical investigation indicates that RAD-23 modifies mutTDP-43 and mutSOD1 abundance, solubility, and turnover in association with altering the ubiquitination status of these substrates. In human amyotrophic lateral sclerosis spinal cord, we find that RAD-23 abundance is increased and RAD-23 is mislocalized within motor neurons. We propose a novel pathophysiological function for RAD-23 in the stabilization of mutated proteins that cause neurodegeneration. SIGNIFICANCE STATEMENT: In this work, we identify RAD-23, a component of the protein homeostasis network and nucleotide excision repair pathway, as a modifier of the toxicity of two disease-causing, misfolding-prone proteins, SOD1 and TDP-43. Reducing the abundance of RAD-23 accelerates the degradation of mutant SOD1 and TDP-43 and reduces the cellular content of the toxic species. The existence of endogenous proteins that act as "anti-chaperones" uncovers new and general targets for therapeutic intervention.

Regulation of protein quality control by UBE4B and LSD1 through p53-mediated transcription.

Protein quality control is essential for clearing misfolded and aggregated proteins from the cell, and its failure is associated with many neurodegenerative disorders. Here, we identify two genes, ufd-2 and spr-5, that when inactivated, synergistically and robustly suppress neurotoxicity associated with misfolded proteins in Caenorhabditis elegans. Loss of human orthologs ubiquitination factor E4 B (UBE4B) and lysine-specific demethylase 1 (LSD1), respectively encoding a ubiquitin ligase and a lysine-specific demethylase, promotes the clearance of misfolded proteins in mammalian cells by activating both proteasomal and autophagic degradation machineries. An unbiased search in this pathway reveals a downstream effector as the transcription factor p53, a shared substrate of UBE4B and LSD1 that functions as a key regulator of protein quality control to protect against proteotoxicity. These studies identify a new protein quality control pathway via regulation of transcription factors and point to the augmentation of protein quality control as a wide-spectrum antiproteotoxicity strategy.

Oxygen sensing neurons and neuropeptides regulate survival after anoxia in developing C. elegans.

Hypoxic brain injury remains a major source of neurodevelopmental impairment for both term and preterm infants. The perinatal period is a time of rapid transition in oxygen environments and developmental resetting of oxygen sensing. The relationship between neural oxygen sensing ability and hypoxic injury has not been studied. The oxygen sensing circuitry in the model organism C. elegans is well understood. We leveraged this information to investigate the effects of impairments in oxygen sensing on survival after anoxia. There was a significant survival advantage in developing worms specifically unable to sense oxygen shifts below their preferred physiologic range via genetic ablation of BAG neurons, which appear important for conferring sensitivity to anoxia. Oxygen sensing that is mediated through guanylate cyclases (gcy-31, 33, 35) is unlikely to be involved in conferring this sensitivity. Additionally, animals unable to process or elaborate neuropeptides displayed a survival advantage after anoxia. Based on these data, we hypothesized that elaboration of neuropeptides by BAG neurons sensitized animals to anoxia, but further experiments indicate that this is unlikely to be true. Instead, it seems that neuropeptides and signaling from oxygen sensing neurons operate through independent mechanisms, each conferring sensitivity to anoxia in wild type animals.

GluA1 promotes the activity-dependent development of motor circuitry in the developing segmental spinal cord.

The neuronal dendritic tree is a key determinant of how neurons receive, compute, and transmit information. During early postnatal life, synaptic activity promotes dendrite elaboration. Spinal motor neurons utilize GluA1-containing AMPA (2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl) propanoic acid) receptors (AMPA-R) to control this process. This form of developmental dendrite growth can occur independently of N-methyl-d-aspartate receptors (NMDA-R). This review focuses on the mechanism by which the GluA1 subunit of AMPA-R transforms synaptic activity into dendrite growth, and describes the essential role of the GluA1 binding partner SAP97 (synapse-associated protein of 97 kDa molecular weight) in this process. This work defines a new mechanism of activity-dependent development, which might be harnessed to stimulate the recovery of function following insult to the central nervous system.