Diverse CMT2 neuropathies are linked to aberrant G3BP interactions in stress granules.

Complex diseases often involve the interplay between genetic and environmental factors. Charcot-Marie-Tooth type 2 neuropathies (CMT2) are a group of genetically heterogeneous disorders, in which similar peripheral neuropathology is inexplicably caused by various mutated genes. Their possible molecular links remain elusive. Here, we found that upon environmental stress, many CMT2-causing mutant proteins adopt similar properties by entering stress granules (SGs), where they aberrantly interact with G3BP and integrate into SG pathways. For example, glycyl-tRNA synthetase (GlyRS) is translocated from the cytoplasm into SGs upon stress, where the mutant GlyRS perturbs the G3BP-centric SG network by aberrantly binding to G3BP. This disrupts SG-mediated stress responses, leading to increased stress vulnerability in motoneurons. Disrupting this aberrant interaction rescues SG abnormalities and alleviates motor deficits in CMT2D mice. These findings reveal a stress-dependent molecular link across diverse CMT2 mutants and provide a conceptual framework for understanding genetic heterogeneity in light of environmental stress.

Phase Engineering of Nanomaterials: Transition Metal Dichalcogenides.

Crystal phase, a critical structural characteristic beyond the morphology, size, dimension, facet, etc., determines the physicochemical properties of nanomaterials. As a group of layered nanomaterials with polymorphs, transition metal dichalcogenides (TMDs) have attracted intensive research attention due to their phase-dependent properties. Therefore, great efforts have been devoted to the phase engineering of TMDs to synthesize TMDs with controlled phases, especially unconventional/metastable phases, for various applications in electronics, optoelectronics, catalysis, biomedicine, energy storage and conversion, and ferroelectrics. Considering the significant progress in the synthesis and applications of TMDs, we believe that a comprehensive review on the phase engineering of TMDs is critical to promote their fundamental studies and practical applications. This Review aims to provide a comprehensive introduction and discussion on the crystal structures, synthetic strategies, and phase-dependent properties and applications of TMDs. Finally, our perspectives on the challenges and opportunities in phase engineering of TMDs will also be discussed.

Progressive Ataxia due to de novo Missense Variants in the CACNA1A Gene.

The CACNA1A gene encodes the alpha-1A subunit of P/Q type voltage-gated calcium channel Cav2.1, which is associated with a broad clinical spectrum and variable symptomatology. While few patients with progressive ataxia caused by CACNA1A missense variants have been reported, here we report three unrelated Chinese patients with progressive ataxia due to de novo missense variants in the CACNA1A gene, including a novel pathogenic variant (c.4999C > G) and a previously reported pathogenic variant (c.4037G > A). Our findings and a systematic literature review show the unique phenotype of progressive ataxia caused by missense variants and enlarge the genetic and clinical spectrum of CACNA1A. This suggests that in addition to routine screening for dynamic mutations, screening for CACNA1A variants is important for clinicians facing patients with progressive ataxia.

Basal forebrain mediates prosocial behavior via disinhibition of midbrain dopamine neurons.

Sociability is fundamental for our daily life and is compromised in major neuropsychiatric disorders. However, the neuronal circuit mechanisms underlying prosocial behavior are still elusive. Here we identify a causal role of the basal forebrain (BF) in the control of prosocial behavior via inhibitory projections that disinhibit the midbrain ventral tegmental area (VTA) dopamine (DA) neurons. Specifically, BF somatostatin-positive (SST) inhibitory neurons were robustly activated during social interaction. Optogenetic inhibition of these neurons in BF or their axon terminals in the VTA largely abolished social preference. Electrophysiological examinations further revealed that SST neurons predominantly targeted VTA GABA neurons rather than DA neurons. Consistently, optical inhibition of SST neuron axon terminals in the VTA decreased DA release in the nucleus accumbens during social interaction, confirming a disinhibitory action. These data reveal a previously unappreciated function of the BF in prosocial behavior through a disinhibitory circuitry connected to the brain's reward system.

Application Value of Serum Neurofilament Light Protein for Disease Staging in Huntington's Disease.

BACKGROUND: Neurofilament light protein (NfL) has been proven to be a sensitive biomarker for Huntington's disease (HD). However, these studies did not include HD patients at advanced stages or with larger CAG repeats (>50), leading to a knowledge gap of the characteristics of NfL. METHODS: Serum NfL (sNfL) levels were quantified using an ultrasensitive immunoassay. Participants were assessed by clinical scales and 7.0 T magnetic resonance imaging. Longitudinal samples and clinical data were obtained. RESULTS: Baseline samples were available from 110 controls, 90 premanifest HD (pre-HD) and 137 HD individuals. We found levels of sNfL significantly increased in HD compared to pre-HD and controls (both P < 0.0001). The increase rates of sNfL were differed by CAG repeat lengths. However, there was no difference in sNfL levels in manifest HD from early to late stages. In addition, sNfL levels were associated with cognitive measures in pre-HD and manifest HD group, respectively. The increased levels of sNfL were also closely related to microstructural changes in white matter. In the longitudinal analysis, baseline sNfL did not correlate with subsequent clinical function decline. Random forest analysis revealed that sNfL had good power for predicting disease onset. CONCLUSIONS: Although sNfL levels are independent of disease stages in manifest HD, it is still an optimal indicator for predicting disease onset and has potential use as a surrogate biomarker of treatment effect in clinical trials. © 2023 International Parkinson and Movement Disorder Society.

CAT Interruption as a Protective Factor in Chinese Patients with Spinocerebellar Ataxia Type 1.

Spinocerebellar ataxia type 1 (SCA1) is the third most common type of spinocerebellar ataxias in China. CAT interruptions in the pathogenic alleles of SCA1 patients had only been reported by limited documents and there was a lack of data based on the Chinese population. In this study, we detected CAT interrupted pathogenic alleles in SCA1 patients from 4 out of 79 (5.1%) Chinese families. Their total CAG repeats were larger (median 58 vs. 47, p < 0.001) but ages at onset were later (median 46 vs. 38, p = 0.020). The longest uninterrupted CAG repeats could explain 65.4% of the AAO variance, making an increase of 28.0% compared to the total CAG repeats. The interruption pattern was greatly different between Chinese cohort and Caucasian cohort, indicating the effect of race.

Sevoflurane Confers Protection Against the Malignant Phenotypes of Lung Cancer Cells via the microRNA-153-3p/HIF1α/KDM2B Axis.

Sevoflurane is shown to curtail lung cancer (LC) development. Herein, this research sought to investigate the underlying mechanism of sevoflurane in regard to its repressive effects on LC. Expression levels of microRNA (miR)-153-3p, HIF1α, and KDM2B in LC tissues and cells were determined with qRT-PCR. Following sevoflurane pretreatment and/or ectopic expression and knockdown experiments, the malignant phenotypes, and levels of miR-153-3p, HIF1α, and KDM2B in LC A549 cells were detected using Transwell, scratch, EdU, CCK-8, Western blot, and qRT-PCR assays. Relationship between HIF1α and miR-153-3p was verified with a dual-luciferase reporter assay. The interaction between HIF1α and KDM2B was verified with a ChIP assay. LC tissues and cells presented low miR-153-3p expression and high HIF1α and KDM2B expression. Sevoflurane pretreatment, miR-153-3p upregulation, HIF1α downregulation, or KDM2B downregulation impeded the malignant phenotypes of A549 cells. Sevoflurane pretreatment augmented miR-153-3p expression, while miR-153-3p negatively targeted HIF1α. HIF1α bound to the KDM2B promoter to upregulate KDM2B. HIF1α or KDM2B overexpression counteracted the inhibitory effects of sevoflurane pretreatment on A549 cell malignant behaviors. Sevoflurane decreased HIF1α expression through upregulation of miR-153-3p, thereby reducing KDM2B transcription to restrict the malignant phenotypes of LC A549 cells.

CALHM2 V136G polymorphism reduces astrocytic ATP release and is associated with depressive symptoms and Alzheimer's disease risk.

INTRODUCTION: Depression is considered a prodromal state of Alzheimer's disease (AD), yet the underlying mechanism(s) by which depression increases the risk of AD are not known. METHODS: Single-nucleotide polymorphism (SNP) analysis was used to determine the CALHM2 variants in AD patients. Cellular and molecular experiments were conducted to investigate the function of CALHM2 V136G mutation. We generated a new genetically engineered Calhm2 V136G mouse model and performed behavioral tests with these mice. RESULTS: CALHM2 V136G mutation (rs232660) is significantly associated with AD. V136G mutation resulted in loss of the CALHM2 ATP-release function in astrocytes and impaired synaptic plasticity. Mice homozygous for the Calhm2 V136G allele displayed depressive-like behaviors that were rescued by administration of exogenous ATP. Moreover, Calhm2 V136G mutation predisposed mice to cognitive decline in old age. DISCUSSION: CALHM2 dysfunction is a biologically relevant mechanism that may contribute to the observed clinical correlation between depression and AD.

Pathogenesis and therapeutic advances of cerebral autosomal- dominant arteriopathy with subcortical infarcts and leukoencephalopathy.

Cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common hereditary cerebral small vessel disease in adults. Many CADASIL cases were reported after NOTCH3 was identified as the causative gene of CADASIL. However, there is still no specific and effective therapies for CADASIL. In this review, we summarize recent research progress on disease models, symptomatic treatments and potential therapies for CADASIL, thereby providing a reference for follow-up CADASIL treatment research.

MicroRNA-31 mediated by interferon regulatory factor 7 signaling facilitates control of Mycobacterium tuberculosis infection.

Tuberculosis (TB) induced by Mycobacterium tuberculosis (M. tuberculosis) infection remains a global most deadly infectious disease. While development of more effective TB vaccines and therapeutics relies on identifications of true biomarkers designating an immune protection against M. tuberculosis infection, exact protective immune components against M. tuberculosis infection remain largely unidentified. We previously found that severe TB induced remarkable up-regulation of interferon regulatory factor 7 (IRF7) and IRF7-related gene signatures, implicating that some unknown downstream molecules in IRF7 signaling cascades may determine the M. tuberculosis infection outcomes and serve as a protective immune component against M. tuberculosis infection. Indeed, here, we observe that genetic ablation of IRF7 leads to more severe lung pathology, increased M. tuberculosis burdens, impaired differentiation of effector/memory T subsets, and extensively elevated expression of pro-inflammatory cytokines in lungs. Importantly, IRF7 is vital for sustaining expression of PD-1/PD-L1 and PD-1/PD-L1-modulated miRNA-31. Moreover, interventions of miRNA-31 expressions via administration of miRNA-31 agomir reduces lung pathology and bacilli burdens via inducing up-regulation of gene sets involved in biological processes of defense response or cellular and chemical homeostasis in lungs. Thus, this study uncovers previously unrecognized importance and mechanisms of IRF7-mediated miRNA-31 as a protective immune component against M. tuberculosis infection.

Features Differ Between Paroxysmal Kinesigenic Dyskinesia Patients with PRRT2 and TMEM151A Variants.

BACKGROUND: Mutations in proline-rich transmembrane protein 2 (PRRT2) are the major cause of paroxysmal kinesigenic dyskinesia (PKD). We recently reported transmembrane protein 151A (TMEM151A) mutations caused PKD. Herein, we aimed to conduct phenotypic comparisons of patients with PKD carrying PRRT2 variants, carrying TMEM151A variants, and carrying neither the PRRT2 nor TMEM151A variant. METHODS: Sanger sequencing of PRRT2 and TMEM151A was performed, and phenotypic characteristics were analyzed. RESULTS: In a cohort of 131 PKD probands (108 without PRRT2 variants and 23 newly recruited), five novel TMEM151A variants were identified and one (c.647C > A) occurred de novo. Together with our previous studies, PRRT2 and TMEM151A variants accounted for 34.7% (85/245) and 6.9% (17/245) of PKD probands, respectively. Compared with patients carrying PRRT2 variants, those with TMEM151A variants tended to exbibit dystonia with shorter durations, have no history of benign infantile epilepsy, and have residual attacks/aura when treated with carbamazepine/oxcarbazepine. CONCLUSIONS: Patients with TMEM151A variants have different features from patients with PRRT2 variants. © 2022 International Parkinson and Movement Disorder Society.

Serum Neurofilament Light Chain in Wilson's Disease: A Promising Indicator but Unparallel to Real-Time Treatment Response.

BACKGROUND: Wilson's disease (WD) currently lacks a promising indicator that could reflect neurological impairment and monitor treatment outcome. We aimed to investigate whether serum neurofilament light chain (sNfL) functions as a candidate for disease assessment and treatment monitoring of WD. METHODS: We assessed preclinical and manifested WD patients' sNfL levels compared to controls and analyzed the differences between patients with various clinical symptoms. We then explored the correlation between clinical scales and sNfL levels. And repeated measurements were performed in 34 patients before and after treatment. RESULTS: WD patients with neurological involvement had significantly higher sNfL levels than both hepatic patients and controls. Positive correlations were found between Unified Wilson's Disease Rating Scale scores and sNfL and between semiquantitative magnetic resonance imaging scales and sNfL levels in WD patients. However, in the treatment follow-up analysis, the trend of sNfL before and after treatment disaccorded with clinical response. CONCLUSION: These findings suggest that sNfL levels can be an ideal indicator for the severity of neurological involvement but fail to evaluate change in disease condition after treatment. © 2022 International Parkinson and Movement Disorder Society.

Bi-allelic loss of function variants in COX20 gene cause autosomal recessive sensory neuronopathy.

Sensory neuronopathies are a rare and distinct subgroup of peripheral neuropathies, characterized by degeneration of the dorsal root ganglia neurons. About 50% of sensory neuronopathies are idiopathic and genetic causes remain to be clarified. Through a combination of homozygosity mapping and whole exome sequencing, we linked an autosomal recessive sensory neuronopathy to pathogenic variants in the COX20 gene. We identified eight unrelated families from the eastern Chinese population carrying a founder variant c.41A>G (p.Lys14Arg) within COX20 in either a homozygous or compound heterozygous state. All patients displayed sensory ataxia with a decrease in non-length-dependent sensory potentials. COX20 encodes a key transmembrane protein implicated in the assembly of mitochondrial complex IV. We showed that COX20 variants lead to reduction of COX20 protein in patient's fibroblasts and transfected cell lines, consistent with a loss-of-function mechanism. Knockdown of COX20 expression in ND7/23 sensory neuron cells resulted in complex IV deficiency and perturbed assembly of complex IV, which subsequently compromised cell spare respiratory capacity and reduced cell proliferation under metabolic stress. Consistent with mitochondrial dysfunction in knockdown cells, reduced complex IV assembly, enzyme activity and oxygen consumption rate were also found in patients' fibroblasts. We speculated that the mechanism of COX20 was similar to other causative genes (e.g. SURF1, COX6A1, COA3 and SCO2) for peripheral neuropathies, all of which are functionally important in the structure and assembly of complex IV. Our study identifies a novel causative gene for the autosomal recessive sensory neuronopathy, whose vital function in complex IV and high expression in the proprioceptive sensory neuron further underlines loss of COX20 contributing to mitochondrial bioenergetic dysfunction as a mechanism in peripheral sensory neuron disease.

Transcriptome analysis reveals candidate genes involved in multiple heavy metal tolerance in hyperaccumulator Sedum alfredii.

Sedum alfredii Hance is a perennial herb native to China that can particularly be found in regions with abandoned Pb/Zn mines. It is a Cd/Zn hyperaccumulator that is highly tolerant to Pb, Cu, Ni, and Mn, showing potential for phytoremediation of soils contaminated with multiple heavy metals. A better understanding of how this species responds to different heavy metals would advance the phytoremediation efficiency. In this study, transcriptomic regulation of S. alfredii roots after Cd, Zn, Pb, and Cu exposure was analyzed to explore the candidate genes involved in multi-heavy metal tolerance. Although Zn and Cd, Pb and Cu had similar distribution patterns in S. alfredii, distinct expression patterns were exhibited among these four metal treatments, especially about half of the differentially expressed genes were upregulated under Cu treatment, suggesting that it utilizes distinctive and flexible strategies to cope with specific metal stress. Most unigenes regulated by Cu were enriched in catalytic activity, whereas the majority of unigenes regulated by Pb had unknown functions, implying that S. alfredii may have a unique strategy coping with Pb stress different from previous cognition. The unigenes that were co-regulated by multiple heavy metals exhibited functions of antioxidant substances, antioxidant enzymes, transporters, transcription factors, and cell wall components. These metal-induced responses at the transcriptional level in S. alfredii were highly consistent with those at the physiological level. Some of these genes have been confirmed to be related to heavy metal absorption and detoxification, and some were found to be related to heavy metal tolerance for the first time in this study, like Metacaspase-1 and EDR6. These results provide a theoretical basis for the use of genetic engineering technology to modify plants by enhancing multi-metal tolerance to promote phytoremediation efficiency.

Preliminary experience of oblique occlusion technique in robot-assisted infrahepatic inferior vena cava thrombectomy: step-by-step procedures and short term outcomes.

BACKGROUND: We aimed to compare the oncological outcomes between the oblique occlusion technique and the traditional technique for robot-assisted radical nephrectomy (RARN) with inferior vena cava (IVC) thrombectomy, and to explore the safety and effectiveness of the oblique occlusion technique. METHODS: Overall, 21 patients with renal cell carcinoma (RCC) and IVC tumor thrombus (TT) were admitted to our hospital from August 2019 to June 2020. All the patients underwent RARN with IVC thrombectomy, of which the IVC oblique occlusion technique was used in 11 patients and traditional occlusion technique was used in 10 patients. The oblique occlusion technique refers to oblique blocking from the upper corner of the right renal vein to the lower corner of the left renal vein using a vessel tourniquet or a vessel clamp (left RCC with IVCTT as an example). RESULTS: Compared with patients in the traditional group, those in the oblique group had lower serum creatinine at follow-up (3 month) (95 ± 21.1 vs. 131 ± 30.7 µmol/L, P = 0.03). There was no significant difference in operation time [149 (IQR 143-245) min vs. 148 (IQR 108-261) min, p = 0.86], IVC clamping time [18 (IQR 12-20) min vs. 20 (IQR 14-23) min, p = 0.41], and estimated intraoperative blood loss [300 (IQR 100-800) mL vs. 500 (IQR 175-738) mL, p = 0.51] between both groups. During a 16-month (range, 15-23 months) follow-up period, two cases progressed in the oblique group and three cases progressed in the traditional group. CONCLUSIONS: The modified IVC oblique occlusion technique procedure is relatively safe and effective in RARN with IVC thrombectomy. The IVC oblique occlusion technique may play a role in the protection of renal function.

Non-Covalent Dimer as Donor Chromophore for Constructing Artificial Light-Harvesting System in Water.

Dynamic emissive materials in aqueous media have received much attention owing to their ease of preparation, tunable luminescence and environmental friendliness. However, hydrophobic fluorophores usually suffer from aggregation-caused quenching in water. In this work, we constructed an artificial light-harvesting system by using a non-covalent aggregation-induced emission dimer as antenna and energy donor. The dimer is quadruple hydrogen bonded from a ureidopyrimidinone derivative (M) containing a tetraphenylethylene group. The dispersed nano-assemblies based on the dimer in aqueous media were fabricated with the help of surfactant. By loading a hydrophobic acceptor molecule DBT into the nano-assemblies, man-made light-harvesting nanoparticles were fabricated, showing considerable energy transfer efficiency and a relatively high antenna effect. Additionally, the fluorescence color of the system can be gradually tuned by varying the content of the acceptors. This study provides a general way for the construction of an aqueous light-harvesting system based on a supramolecular dimer, which is important for potential application in luminescent materials.

Role for Biochemical Assays and Kayser-Fleischer Rings in Diagnosis of Wilson's Disease.

BACKGROUND & AIMS: Wilson disease is an autosomal recessive disorder that impairs copper homeostasis and is caused by homozygous or compound heterozygous mutations in ATP7B, which encodes a copper-transporting P-type ATPase. Patients have variable clinical manifestations and laboratory test results, resulting in diagnostic dilemmas. We aimed to identify factors associated with symptoms and features of Wilson disease from a large cohort, over 15 years. METHODS: We collected data from 715 patients (529 with symptoms, 146 without symptoms, and 40 uncategorized) and a genetic confirmation of Wilson's disease (mean age of diagnosis, 18.84 years), recruited from 3 hospitals in China from 2004 through 2019. We analyzed clinical data along with serum levels of ceruloplasmin (available from 636 patients), 24-hr urinary copper excretion (collected from 131 patients), Kayser-Fleisher rings (copper accumulation in eyes, with neurologic data from 355 patients), and magnetic resonance imaging (MRI) abnormalities. Differences among the groups were analyzed using 1-way analysis of variance followed by Tukey multiple comparison test. RESULTS: Of the 529 patients with symptoms, 121 had hepatic features, 355 had neurologic features, 28 had osteomuscular features (premature osteoarthritis, skeletal deformities, and pathological bone fractures), and 25 had psychiatric symptoms. Age of onset was significantly younger in patients with hepatic (16.94 ± 1.03 years; P = .0105) or osteomuscular features (13 ± 1.33 years; P = .0001) than patients with neurological features (19.48 ± 0.46 years). Serum levels of ceruloplasmin differed among asymptomatic patients and patients with osteomuscular or neurologic symptoms of Wilson disease. Serum levels of ceruloplasmin ranged from 18.93 mg/L to approximately 120.00 mg/L (quantiles of 0.025 to approximately 0.975). Fifty-one of 131 patients (39%) had urinary copper excretion levels below 100 µg/24 hr; there was significant variation in levels of urinary copper excretion between patients older than 14 years vs 14 years or younger. Of the 355 patients with neurologic features, 244 patients (69%) had abnormal findings from MRI and Kayser-Fleisher rings; only 1 patient with abnormal findings from brain MRI was negative for Kayser-Fleisher rings. CONCLUSIONS: Serum level of ceruloplasmin, 24-hour urinary copper excretion, and Kayser-Fleisher rings can be used to identify patients who might have Wilson disease. Patients with serum levels of ceruloplasmin below 120 mg/L and children with urinary copper excretion above 40 µg should undergo genetic testing for Wilson's disease. Patients with movement disorders and brain MRI abnormalities without Kayser-Fleisher rings are not likely to have Wilson disease.

MicroRNA miR-326 regulates TH-17 differentiation and is associated with the pathogenesis of multiple sclerosis.

Interleukin 17 (IL-17)-producing T helper cells (T(H)-17 cells) are increasingly recognized as key participants in various autoimmune diseases, including multiple sclerosis. Although sets of transcription factors and cytokines are known to regulate T(H)-17 differentiation, the role of noncoding RNA is poorly understood. Here we identify a T(H)-17 cell-associated microRNA, miR-326, whose expression was highly correlated with disease severity in patients with multiple sclerosis and mice with experimental autoimmune encephalomyelitis (EAE). In vivo silencing of miR-326 resulted in fewer T(H)-17 cells and mild EAE, and its overexpression led to more T(H)-17 cells and severe EAE. We also found that miR-326 promoted T(H)-17 differentiation by targeting Ets-1, a negative regulator of T(H)-17 differentiation. Our data show a critical role for microRNA in T(H)-17 differentiation and the pathogenesis of multiple sclerosis.

Isolation and genomic analysis of temperate phage 5W targeting multidrug-resistant Acinetobacter baumannii.

Temperate phages are potential therapeutic agents, but only a few temperate phages infecting multidrug-resistant Acinetobacter baumannii have been identified. In this study, we isolated 5W, a temperate phage that infects multidrug-resistant A. baumannii, from pond water using the enrichment method. A member of the Siphoviridae family, 5W has a narrow host range and infected only four of 19 A. baumannii clinical isolates. It exhibited rapid adsorption (> 90% in 6 min), a latency period of 20 min, and a burst size of ~ 180 plaque-forming units (PFU/cell). 5W contains a linear double-stranded DNA (dsDNA) genome of 43,032 bp with a GC content of 39.85%. The 5W genome contains 61 open reading frames, including lysogen-forming genes, but lacks any known virulence and antibiotic resistance genes. The lysin of 5W is an N-acetyl-ß-D-muramidase belonging to the GH_108 family. The α-helical structure and highly positively charged amino acids in the C-terminal region indicate potential antibacterial activity against A. baumannii, and the M/S subunits of the restriction endonuclease are inserted into the lysogenic gene cluster. Comparative genome analysis revealed high similarity with two different prophages in A. baumannii ABCR01, suggesting that 5W may be derived from recombination of other prophages.

CHIP control degradation of mutant ETF:QO through ubiquitylation in late-onset multiple acyl-CoA dehydrogenase deficiency.

Late-onset multiple acyl-CoA dehydrogenase deficiency (MADD) is the most common form of lipid storage myopathy. The disease is mainly caused by mutations in electron-transfer flavoprotein dehydrogenase gene (ETFDH), which leads to decreased levels of ETF:QO in skeletal muscle. However, the specific underlying mechanisms triggering such degradation remain unknown. We constructed expression plasmids containing wild type ETF:QO and mutants ETF:QO-A84T, R175H, A215T, Y333C, and cultured patient-derived fibroblasts containing the following mutations in ETFDH: c.250G>A (p.A84T), c.998A>G (p.Y333C), c.770A>G (p.Y257C), c.1254_1257delAACT (p. L418TfsX10), c.524G>A (p.R175H), c.380T>A (p.L127P), and c.892C>T (p.P298S). We used in vitro expression systems and patient-derived fibroblasts to detect stability of ETF:QO mutants then evaluated their interaction with Hsp70 interacting protein CHIP with active/inactive ubiquitin E3 ligase carboxyl terminus using western blot and immunofluorescence staining. This interaction was confirmed in vitro and in vivo by co-immunoprecipitation and immunofluorescence staining. We confirmed the existence two ubiquitination sites in mutant ETF:QO using mass spectrometry (MS) analysis. We found that mutant ETF:QO proteins were unstable and easily degraded in patient fibroblasts and in vitro expression systems by ubiquitin-proteasome pathway, and identified the specific ubiquitin E3 ligase as CHIP, which forms complex to control mutant ETF:QO degradation through poly-ubiquitination. CHIP-dependent degradation of mutant ETF:QO proteins was confirmed by MS and site-directed mutagenesis of ubiquitination sites. Hsp70 is directly involved in this process as molecular chaperone of CHIP. CHIP plays an important role in ubiquitin-proteasome pathway dependent degradation of mutant ETF:QO by working as a chaperone-assisted E3 ligase, which reveals CHIP's potential role in pathological mechanisms of late-onset MADD.