Correction to: Increased RTN3 phenocopies nonalcoholic fatty liver disease by inhibiting the AMPK-IDH2 pathway.

[This corrects the article DOI: 10.1002/mco2.226.].

RTN3 deficiency exacerbates cisplatin-induced acute kidney injury through the disruption of mitochondrial stability.

Reticulum 3 (RTN3) is an endoplasmic reticulum (ER) protein that has been reported to act in neurodegenerative diseases and lipid metabolism. However, the role of RTN3 in acute kidney injury (AKI) has not been explored. Here, we employed public datasets, patient data, and animal models to explore the role of RTN3 in AKI. The underlying mechanisms were studied in primary renal tubular epithelial cells and in the HK2 cell line. We found reduced expression of RTN3 in AKI patients, cisplatin-induced mice, and cisplatin-treated HK2 cells. RTN3-null mice exhibit more severe AKI symptoms and kidney fibrosis after cisplatin treatment. Mitochondrial dysfunction was also found in cells with RTN3 knockdown or knockout. A mechanistic study revealed that RTN3 can interact with HSPA9 in kidney cells. RTN3 deficiency may disrupt the RTN3-HSPA9-VDAC2 complex and affect MAMs during ER-mitochondrion contact, which further leads to mitochondrial dysfunction and exacerbates cisplatin-induced AKI. Our study indicated that RTN3 was important in the kidney and that a decrease in RTN3 in the kidney might be a risk factor for the aggravation of AKI.

Reticulon 3 regulates sphingosine-1-phosphate synthesis in endothelial cells to control blood pressure.

The discovery of the endothelium as a major regulator of vascular tone triggered intense research among basic and clinical investigators to unravel the physiologic and pathophysiologic significance of this phenomenon. Sphingosine-l-phosphate (S1P), derived from the vascular endothelium, is a significant regulator of blood pressure. However, the mechanisms underlying the regulation of S1P biosynthetic pathways in arteries remain to be further clarified. Here, we reported that Reticulon 3 (RTN3) regulated endothelial sphingolipid biosynthesis and blood pressure. We employed public datasets, patients, and mouse models to explore the pathophysiological roles of RTN3 in blood pressure control. The underlying mechanisms were studied in human umbilical vein endothelial cells (HUVECs). We reported that increased RTN3 was found in patients and that RTN3-null mice presented hypotension. In HUVECs, RTN3 can regulate migration and tube formation via the S1P signaling pathway. Mechanistically, RTN3 can interact with CERS2 to promote the selective autophagy of CERS2 and further influence S1P signals to control blood pressure. We also identified an RTN3 variant (c.116C>T, p.T39M) in a family with hypertension. Our data provided the first evidence of the association between RTN3 level changes and blood pressure anomalies and preliminarily elucidated the importance of RTN3 in S1P metabolism and blood pressure regulation.

A compound heterozygous mutation of ERCC8 is responsible for a family with Cockayne syndrome.

BACKGROUND: Cockayne syndrome is an inherited heterogeneous defect in transcription-coupled DNA repair (TCR) cause severe clinical syndromes, which may affect the nervous system development of infants and even lead to premature death in some cases. ERCC8 diverse critical roles in the nucleotide excision repair (NER) complex, which is one of the disease-causing genes of Cockayne syndrome. METHODS AND RESULTS: The mutation of ERCC8 in the patient was identified and validated using WES and Sanger sequencing. Specifically, a compound heterozygous mutation (c.454_460dupGTCTCCA p. T154Sfs*13 and c.755_759delGTTTT p.C252Yfs*3) of ERCC8 (CSA) was found, which could potentially be the genetic cause of Cockayne syndrome in the proband. CONCLUSION: In this study, we identified a novel heterozygous mutation of ERCC8 in a Chinese family with Cockayne syndrome, which enlarging the genetic spectrum of the disease.

Identification of A Novel Variant of Filamin A Destroying the Attraction Between Benzene Rings and Sulfhydryl in Developmental Dysplasia of the Hip.

Developmental dysplasia of the hip (DDH), characterized by acetabular deformity that manifests from loose ligaments to complete dislocation of the hip, can cause notable pain and dysfunction and lead to hip dislocation, secondary fractures, scoliosis, and osteoarthritis of hip. Variants in FLNA may produce a spectrum of malformations in multiple organs, especially the skeleton. This study aimed to identify the genetic etiologies of DDH patients and provide genetic testing information for further diagnosis and treatment of DDH. We recruited a Chinese woman with DDH and her family members. Whole-exome sequencing was used to identify the patient's genetic etiologies. Protein models were used to analyze the pathogenic mechanism of the identified variants. A novel variant (c.3493T>G, p.C1165G) of FLNA was detected. The structural models of the mutant FLNA protein indicated that the variant would lose its sulfhydryl side chain and destroy the attraction between benzene rings and sulfhydryl. We reported a novel variant (c.3493T>G, p.C1165G) of FLNA in a Chinese woman with DDH. Our research outcome enriches the gene pool for hip dysplasia and emphasizes the pathogenicity of sulfhydryl side chain disruption in FLNA.

Case report: Identification of novel fibrillin-2 variants impacting disulfide bond and causing congenital contractural arachnodactyly.

Background: Congenital contractural arachnodactyly (CCA) is an autosomal dominant connective tissue disorder with clinical features of arthrogryposis, arachnodactyly, crumpled ears, scoliosis, and muscular hypoplasia. The heterozygous pathogenic variants in FBN2 have been shown to cause CCA. Fibrillin-2 is related to the elasticity of the tissue and has been demonstrated to play an important role in the constitution of extracellular microfibrils in elastic fibers, providing strength and flexibility to the connective tissue that sustains the body's joints and organs. Methods: We recruited two Chinese families with arachnodactyly and bilateral arthrogryposis of the fingers. Whole-exome sequencing (WES) and co-segregation analysis were employed to identify their genetic etiologies. Three-dimensional protein models were used to analyze the pathogenic mechanism of the identified variants. Results: We have reported two CCA families and identified two novel missense variants in FBN2 (NM_001999.3: c.4093T>C, p.C1365R and c.2384G>T, p.C795F). The structural models of the mutant FBN2 protein in rats exhibited that both the variants could break disulfide bonds. Conclusion: We detected two FBN2 variants in two families with CCA. Our description expands the genetic profile of CCA and emphasizes the pathogenicity of disulfide bond disruption in FBN2.

Increased RTN3 phenocopies nonalcoholic fatty liver disease by inhibiting the AMPK-IDH2 pathway.

Reticulon 3 (RTN3), an endoplasmic reticulum protein, is crucial in neurodegenerative and kidney diseases. However, the role of RTN3 in liver tissues has not been described. Here, we employed public datasets, patients, and several animal models to explore the role of RTN3 in nonalcoholic fatty liver disease (NAFLD). The underlying mechanisms were studied in primary hepatocytes and L02 cells in vitro. We found an increased expression of RTN3 in NAFLD patients, high-fat diet mice, and oxidized low-density lipoprotein-treated L02 cells. The RTN3 transgenic mice exhibited the phenotypes of fatty liver and lipid accumulation. Single-cell RNA sequencing analysis indicated that increased RTN3 might induce mitochondrial dysfunction. We further showed this in primary hepatocytes, the L02 cell line, and the Caenorhabditis elegans strain. Mechanistically, RTN3 regulated these events through its interactions with glucose-regulated protein 78 (GRP78), which further inhibited the adenosine 5 monophosphate-activated protein kinase (AMPK)-isocitrate dehydrogenase 2 (IDH2) pathway. In the end, knockout of RTN3 relieved fatty liver and mitochondrial dysfunction. Our study indicated that RTN3 was important in NAFLD and lipid catabolism and that an increase in RTN3 in the liver might be a risk factor for nonalcoholic steatohepatitis and NAFLD.

ZPA Regulatory Sequence Variants in Chinese Patients With Preaxial Polydactyly: Genetic and Clinical Characteristics.

Preaxial polydactyly (PPD) is a common congenital abnormality with an incidence of 0.8-1.4% in Asians, characterized by the presence of extra digit(s) on the preaxial side of the hand or foot. PPD is genetically classified into four subtypes, PPD type I-IV. Variants in six genes/loci [including GLI family zinc finger 3 (GLI3), ZPA regulatory sequence (ZRS), and pre-ZRS region] have been identified in PPD cases. Among these loci, ZRS is, perhaps, the most special and well known, but most articles only reported one or a few cases. There is a lack of reports on the ZRS-variant frequency in patients with PPD. In this study, we recruited 167 sporadic or familial cases (including 154 sporadic patients and 13 families) with PPD from Central-South China and identified four ZRS variants in four patients (2.40%, 4/167), including two novel variants (ZRS131A > T/chr7:g.156584439A > T and ZRS474C > G/chr7:g.156584096C > G) and two known variants (ZRS428T > A/chr7:g.156584142T > A and ZRS619C > T/chr7:g.156583951C > T). ZRS131A > T and ZRS428T > A were detected in PPD I cases and ZRS474C > G and ZRS619C > T combinedly acted to cause PPD II. The detectable rate of ZRS variants in PPD I was 1.60% (2/125), while PPD II was significantly higher (9.52%, 2/21). Three bilateral PPD cases harbored ZRS variants (13.64%, 3/22), suggesting that bilateral PPD was more possibly caused by genetic etiologies. This study identified two novel ZRS variants, further confirmed the association between ZRS and PPD I and reported a rare PPD II case resulted from the compound heterozygote of ZRS. This investigation preliminarily evaluated a ZRS variants rate in patients with PPD and described the general picture of PPD in Central-South China.

Loss of RTN3 phenocopies chronic kidney disease and results in activation of the IGF2-JAK2 pathway in proximal tubular epithelial cells.

Reticulon 3 (RTN3) is an endoplasmic reticulum protein that has previously been shown to play roles in neurodegenerative diseases, but little is known about its function in the kidneys. The aim of the present study was to clarify the roles of RTN3 in chronic kidney disease (CKD) and kidney fibrosis. In this study, RTN3 levels were measured in kidney tissues from healthy controls and CKD or kidney fibrosis patients. An RTN3-null mouse model was generated to explore the pathophysiological roles of RTN3 in the kidneys. The underlying mechanisms were studied in primary proximal tubular epithelial cells and HEK293 cells in vitro. The results showed that (1) a reduction in RTN3 in mice induces CKD and kidney fibrosis; (2) decreased RTN3 expression is found in patients with CKD; (3) RTN3 plays critical roles in regulating collagen biosynthesis and mitochondrial function; and (4) mechanistically, RTN3 regulates these phenotypes by interacting with GC-Rich Promoter Binding Protein 1 (GPBP1), which activates the IGF2-JAK2-STAT3 pathway. Our study indicates that RTN3 might play crucial roles in CKD and kidney fibrosis and that a reduction in RTN3 in the kidneys might be a risk factor for CKD and kidney fibrosis.

Case Report: Identification of the First Synonymous Variant of Myosin Binding Protein C3 (c.24A>C, p.P8P) Altering RNA Splicing in a Cardiomyopathy and Sudden Cardiac Death Case.

Background: Sudden cardiac death (SCD), based on sudden cardiac ejection cessation, is an unexpected death. Primary cardiomyopathies, including dilated cardiomyopathy (DCM), are one of main causes of SCD. The DCM is characterized by a cardiac dilatation and a reduced systolic function with a prevalence of 1/250 in adults. The DCM has been reported with more than 60 disease-causing genes, and MYBPC3 variants are one of the most common and well-known causes of DCM. Methods: We identified a 29-year-old female who died of SCD. We performed a whole-exome sequencing (WES) to detect her genetic etiology and used minigene modeling and immunohistochemistry staining to verify the pathogenicity. Results: We determined that the woman died of SCD caused by DCM due to an identified novel synonymous variant of MYBPC3 (NM_000256.3: c.24A>C, p.P8P) in the deceased. The variant can result in abnormal splicing, which was confirmed by minigene models and immunohistochemistry staining. Conclusion: We may have identified the first deleterious synonymous variant of MYBPC3 in an SCD case and verified its significant impact on RNA splicing. Our description enriched the spectrum of MYBPC3 variants and emphasized the significance of synonymous variants that are always disregarded in genetic screening.

Genetic analysis combined with 3D-printing assistant surgery in diagnosis and treatment for an X-linked hypophosphatemia patient.

BACKGROUND: Hypophosphatemia is mainly characterized by hypophosphatemia and a low level of 1alpha,25-Dihydroxyvitamin D2 (1,25-(OH)2 D2) and/or 1alpha,25-Dihydroxyvitamin D3 (1,25-(OH)2 D3) in the blood. Previous studies have demonstrated that variants in PHEX and FGF23 are primarily responsible for this disease. Although patients with variants of these two genes share almost the same symptoms, they exhibit the different hereditary pattern, X-link dominant and autosome dominant, respectively. Three-dimensional (3D) printing is a method which can accurately reconstruct physical objects, and its applications in orthopedics can contribute to realizing a more accurate surgical performance and a better outcome. METHODS: An X-linked hypophosphatemia (XLH) family was recruited, with four patients across three generations. We screened candidate genes and filtered a duplication variant in PHEX. Variant analysis and co-segregation confirmation were then performed. Before the operation of our patient, a digital model of our patient's leg had been rebuilt upon the CT scan data, and a polylactic acid (PLA) model had been 3D-printed. RESULTS: A novel duplication PHEX variant c.574dupG (p.A192GfsX20) was identified in a family with XLH. Its pathogenicity was confirmed by the co-segregation assay and online bioinformatics database. The preoperative plan was made with the help of the PLA model. Then, arch osteotomy and transverse osteotomy were performed under the guidance of the previous simulation. The appearance of the surgical-intervened leg was satisfactory. CONCLUSIONS: This study identified a novel PHEX variant and showed that 3D printing tech is a very promising approach for corrective osteotomies.

Case Report: A Novel CACNA1S Mutation Associated With Hypokalemic Periodic Paralysis in a Chinese Family.

Hypokalemic periodic paralysis (HypoPP) is a rare autosomal dominant disorder characterized by episodic flaccid paralysis with concomitant hypokalemia. More than half of patients were associated with mutations in CACNA1S that encodes the alpha-1-subunit of the skeletal muscle L-type voltage-dependent calcium channel. Mutations in CACNA1S may alter the structure of CACNA1S and affect the functions of calcium channels, which damages Ca2+-mediated excitation-contraction coupling. In this research, we identified and described a Chinese HypoPP patient with a novel frameshift mutation in CACNA1S [NM_000069.2: c.1364delA (p.Asn455fs)] by targeted sequencing. This study would expand the spectrum of CACNA1S mutations, further our understanding of HypoPP, and provided a new perspective for selecting effective treatments.

Case Report: A Novel Gross Deletion in PAX3 (10.26 kb) Identified in a Chinese Family With Waardenburg Syndrome by Third-Generation Sequencing.

Waardenburg syndrome (WS) is a group of autosomal-dominant hereditary conditions with a global incidence of 1/42,000. WS can be categorized into at least four types: WS1-4, and these are characterized by heterochromia iridis, white forelock, prominent nasal root, dystopia canthorum, hypertrichosis of the medial part of the eyebrows, and deaf-mutism. WS3 is extremely rare, with a unique phenotype (upper limb abnormality). Heterozygous mutations of PAX3 are commonly associated with WS1, whereas partial or total deletions of PAX3 are often observed in WS3 cases. Deletions, together with insertions, translocations, inversions, mobile elements, tandem duplications, and complexes, constitute structural variants (SVs), which can be fully and accurately detected by third-generation sequencing (TGS), a new generation of high-throughput DNA sequencing technology. In this study, after failing to identify the causative gene by Sanger sequencing, SNP-array, and whole-exome sequencing (WES), we finally detected a heterozygous gross deletion of PAX3 (10.26kb, chr2: 223153899-223164405) in a WS family by TGS. Our description would enrich the genetic map of WS and help us to further understand this disease. Our findings also demonstrated the value of TGS in clinical genetics researches.

A novel POF1B variant in a Chinese patient is associated with premature ovarian failure.

A novel mutation of POF1B was identified in a patient with premature ovarian failure.

Case Report: A Homozygous Mutation (p.Y62X) of Phospholipase D3 May Lead to a New Leukoencephalopathy Syndrome.

Leukodystrophies are a heterogeneous group of inherited disorders with highly variable clinical manifestations and pathogenetic backgrounds. At present, variants in more than 20 genes have been described and may be responsible for different types of leukodystrophies. Members of the phospholipase D family of enzymes catalyze the hydrolysis of membrane phospholipids. Meanwhile, phospholipase D3 (PLD3) has also been found to exhibit single stranded DNA (ssDNA) acid 5' exonuclease activity. Variants in phospholipase D3 (PLD3) may increase the risk of Alzheimer's disease and spinocerebellar ataxia, but this hypothesis has not been fully confirmed. In this study, we identified a novel homozygous mutation (NM_012268.3: c.186C>G/ p.Y62X) of PLD3 in a consanguineous family with white matter lesions, hearing and vision loss, and kidney disease by whole exome sequencing. Real-time PCR revealed that the novel mutation may lead to non-sense-mediated messenger RNA (mRNA) decay. This may be the first case report on the homozygous mutation of PLD3 in patients worldwide. Our studies indicated that homozygous mutation of PLD3 may result in a novel leukoencephalopathy syndrome with white matter lesions, hearing and vision loss, and kidney disease.

Whole-Exome Sequencing Identifies a Novel TRPM4 Mutation in a Chinese Family with Atrioventricular Block.

Atrioventricular block (AVB) is a leading cause of sudden cardiac death, and most of AVB cases are presented as autosomal dominant. The electrocardiogram of AVB patients presents an abnormal progressive cardiac conduction disorder between atria and ventricles. Transient receptor potential melastatin 4 (TRPM4) is a nonselective Ca2+-activated cation channel gene defined as a novel disease-causing gene of AVB. So far, 47 mutations of TRPM4 have been recorded in Human Gene Mutation Database. The aim of this study was to explore the relationship between TRPM4 mutation and pathogenesis of AVB. We investigated a Chinese family with AVB by whole-exome sequencing. An arrhythmia-related gene filtering strategy was used to analyze the disease-causing mutations. Three different bioinformatics programs were used to predict the effects of the mutation result. A novel mutation of TRPM4 was identified (c.2455C>T/p.R819C) and cosegregated in the affected family members. The three bioinformatics programs predicted that the novel mutation may lead to damage. Our study will contribute to expand the spectrum of TRPM4 mutations and supply accurate genetic testing information for further research and the clinical therapy of AVB.

Identification of Two Novel Frameshift Mutations in Exostosin 1 in Two Families with Multiple Osteochondromas.

Multiple osteochondromas (MO) is an autosomal dominant hereditary disorder, which typically manifests as skeletal dysplasia, mainly involving long bones and knees, ankles, elbows, wrists, shoulders, and pelvis. Previous studies have demonstrated that mutations in exostosin glycosyl transferase-1 (EXT1) and exostosin glycosyl transferase-2 (EXT2) were the main cause of MO. In this study, we enrolled 2 families with MO. Sanger sequencing revealed 2 novel frameshift mutations - c.1432_1433insCCCCCCT; p.Lys479Profs*44 and c.1431_1431delC; p.S478PfsX10 - in the EXT1 gene detected in 2 families, respectively. Both novel mutations, located in the conserved domain of EXT1 and predicted to be disease causing by informatics programs, were absent in our 200 control cohorts and other public databases. Our study expanded the spectrum of EXT1 mutations and contributed to genetic diagnosis and counseling of patients with MO.

Identification of a novel mutation in the C6 gene of a Han Chinese C6SD child with meningococcal disease.

Deficiency of the sixth complement component (C6D) is a genetic disease associated with increased susceptibility to Neisseria meningitides infection. Individuals with C6D usually present with recurrent meningococcal disease (MD). According to the patients' C6 levels, C6D is divided into complete genetic deficiency of C6 and subtotal deficiency of C6 (C6SD). The present study reported on a Han Chinese pediatric patient with MD, in whom further investigation revealed a C6SD genetic lesion. A heterozygote nonsense mutation (c.1062C>G/p.Y354*) in the C6 gene was identified by Sanger sequencing. The mutation alters the tyrosine codon at position 354 to a termination codon and results in a truncated protein. In conclusion, the genetic lesion of a pediatric patient with C6SD who was diagnosed due to having MD was investigated and a novel pathogenic mutation in the C6 gene was identified. The study confirmed the clinical diagnosis for this patient with C6SD and also expanded the spectrum of C6 mutations.

A Novel COMP Mutated Allele Identified in a Chinese Family with Pseudoachondroplasia.

Pseudoachondroplasia (PSACH) is an autosomal dominant skeletal dysplasia with an estimated incidence of ~1/60000 that is characterized by disproportionate short stature, brachydactyly, joint laxity, and early-onset osteoarthritis. COMP encodes the cartilage oligomeric matrix protein, which is expressed predominantly in the extracellular matrix (ECM) surrounding the cells that make up cartilage, ligaments, and tendons. Mutations in COMP are known to give rise to PSACH. In this study, we identified a novel nucleotide mutation (NM_000095.2: c.1317C>G, p.D439E) in COMP responsible for PSACH in a Chinese family by employing whole-exome sequencing (WES) and built the structure model of the mutant protein to clarify its pathogenicity. The novel mutation cosegregated with the affected individuals. Our study expands the spectrum of COMP mutations and further provides additional genetic testing information for other PSACH patients.

GLIS Family Zinc Finger 1 was First Linked With Preaxial Polydactyly I in Humans by Stepwise Genetic Analysis.

Background: Preaxial polydactyly (PPD) is one of the most common developmental malformations, with a prevalence of 0.8-1.4% in Asians. PPD is divided into four types, PPD I-IV, and PPD I is the most frequent type. Only six loci (GLI1, GLI3, STKLD1, ZRS, pre-ZRS, and a deletion located 240 kb from SHH) have been identified in non-syndromic PPD cases. However, pathogenesis of most PPD patients has never been investigated. This study aimed to understand the genetic mechanisms involved in the etiology of PPD I in a family with multiple affected members. Methods: We recruited a PPD I family (PPD001) and used stepwise genetic analysis to determine the genetic etiology. In addition, for functional validation of the identified GLIS1 variant, in vitro studies were conducted. GLIS1 variants were further screened in additional 155 PPD cases. Results: We identified a GLIS1 variant (NM_147193: c.1061G > A, p.R354H) in the PPD001 family. In vitro studies showed that this variant decreased the nuclear translocation of GLIS1 and resulted in increased cell viability and migration. RNA sequencing revealed abnormal TBX4 and SFRP2 expression in 293T cells transfected with mutant GLIS1. Additionally, we identified a GLIS1 variant (c.664G > A, p.D222N) in another PPD case. Conclusion: We identified two GLIS1 variants in PPD I patients and first linked GLIS1 with PPD I. Our findings contributed to future molecular and clinical diagnosis of PPD and deepened our knowledge of this disease.