C9ORF72-derived dipeptide repeat proteins have emerged as the pathogenic cause of neurodegeneration in amyotrophic lateral sclerosis and frontotemporal dementia (C9-ALS/FTD). However, the mechanisms underlying their expression are not fully understood. Here, we demonstrate that ZNF598, the rate-limiting factor for ribosome-associated quality control (RQC), co-translationally titrates the expression of C9ORF72-derived poly(GR) protein. A Drosophila genetic screen identified key RQC factors as potent modifiers of poly(GR)-induced neurodegeneration. ZNF598 overexpression in human neuroblastoma cells inhibited the nuclear accumulation of poly(GR) protein and decreased its cytotoxicity, whereas ZNF598 deletion had opposing effects. Poly(GR)-encoding sequences in the reporter RNAs caused translational stalling and generated ribosome-associated translation products, sharing molecular signatures with canonical RQC substrates. Furthermore, ZNF598 and listerin 1, the RQC E3 ubiquitin-protein ligase, promoted poly(GR) degradation via the ubiquitin-proteasome pathway. An ALS-relevant ZNF598R69C mutant displayed loss-of-function effects on poly(GR) expression, as well as on general RQC. Moreover, RQC function was impaired in C9-ALS patient-derived neurons, whereas lentiviral overexpression of ZNF598 lowered their poly(GR) expression and suppressed proapoptotic caspase-3 activation. Taken together, we propose that an adaptive nature of the RQC-relevant ZNF598 activity allows the co-translational surveillance to cope with the atypical expression of pathogenic poly(GR) protein, thereby acquiring a neuroprotective function in C9-ALS/FTD.

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Kohlschütter-Tönz syndrome (KTS) manifests as neurological dysfunctions, including early-onset seizures. Mutations in the citrate transporter SLC13A5 are associated with KTS, yet their underlying mechanisms remain elusive. Here, we report that a Drosophila SLC13A5 homolog, I’m not dead yet (Indy), constitutes a neurometabolic pathway that suppresses seizure. Loss of Indy function in glutamatergic neurons caused “bang-induced” seizure-like behaviors. In fact, glutamate biosynthesis from the citric acid cycle was limiting in Indy mutants for seizure-suppressing glutamate transmission. Oral administration of the rate-limiting α-ketoglutarate in the metabolic pathway rescued low glutamate levels in Indy mutants and ameliorated their seizure-like behaviors. This metabolic control of the seizure susceptibility was mapped to a pair of glutamatergic neurons, reversible by optogenetic controls of their activity, and further relayed onto fan-shaped body neurons via the ionotropic glutamate receptors. Accordingly, our findings reveal a micro-circuit that links neural metabolism to seizure, providing important clues to KTS-associated neurodevelopmental deficits.

D-site albumin promoter binding protein (DBP) supports the rhythmic transcription of downstream genes, in part by displaying high-amplitude cycling of its own transcripts compared to other circadian clock genes. However, the underlying mechanism remains elusive. Here, we demonstrated that poly(C) motif within DBP proximal promoters, in addition to an E-box element, provoked the transcriptional activation through increased RNA polymerase 2 (Pol2) recruitment by inducing higher chromatin accessibility. We also clarified that heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a key regulator that binds to the poly(C) motif on single-stranded DNAs in vitro. Chromatin immunoprecipitation further confirmed the expression-dependent and rhythmic binding of hnRNP K which was inhibited through its cytosolic localization mediated by time-dependent ERK activation.Knockdown of hnRNP K triggered low-amplitude mRNA rhythms in DBP and other core clock genes through transcriptional or post-transcriptional regulation. Finally, transgenic depletion of a Drosophila homolog of hnRNP K in circadian pacemaker neurons lengthened 24-hour periodicity in free-running locomotor behaviors. Taken together, our results provide new insights into the function of hnRNP K as a transcriptional amplifier of DBP, which acts rhythmically through its intracellular localization by the ERK phosphorylation and as an mRNA stabilizer along with its physiological significance in circadian rhythms of Drosophila. Significance StatementIn the case of mood disorders and the aging process, the mRNA expression and amplitude level of clock genes, including DBP, were reported to be diminished. However, the reason behind this decrease of clock gene amplitude and expression level remained unclear. Through this study, we revealed the regulatory mechanism behind the expression of clock genes, especially of DBP mRNA expression. In addition, we discovered that hnRNP K regulates more core clock genes than what we have previously known, such as Clock and Periods. Finally, we demonstrated the physiological significance of hnRNP K in Drosophila through its RNAi line model. Hence, our findings show the regulatory mechanism of circadian rhythm that may provide insight on mood disorder and aging process.