Hypospadias is a common congenital malformation of the male external genitalia, with severe cases presenting considerable surgical and long-term challenges. Despite the clinical importance of severe hypospadias demonstrated by prolonged hospital stays, repeated surgeries, and substantial costs, the genetic etiology of severe hypospadias remains incompletely understood, particularly in diverse populations. To determine the molecular basis, we performed whole-exome sequencing (WES) on 30 Chinese patients from southeastern China with confirmed 46,XY karyotypes. Our analysis identified clinically relevant genetic variants, including single-nucleotide variants (SNVs) and copy number variations (CNVs), with subsequent phenotypic correlation. Clinically relevant genetic variants were identified in 33.3% (10/30) of cases, including novel SNVs in gonadal regulators (nuclear receptor subfamily 5 group A member 1 [NR5A1] c.1344dupC/c.244+1G>T and SRY-box 3 [SOX3] c.1273G>C), morphogenetic modulators (GLI family zinc finger 3 [GLI3] c.4731delA and aristaless-related homeobox [mARX] c.644C>G), and syndromic genes (patched domain containing 1 [PTCHD1] c.667G>A and euchromatic histone lysine methyltransferase 1 [EHMT1] c.3081C>T). Additionally, recurrent CNVs at 22q12.3 and a novel CNV exon 18 deletion in myelin regulatory factor (MYRF) and 18q11.2 were identified. Mutation carriers showed a significantly higher frequency of cryptorchidism (40.0% vs 5.0%, P < 0.01) and a higher prevalence of ≥3 associated malformations (80.0% vs 35.0%, P < 0.05) than non-carriers, highlighting genotype-phenotype correlations. The 33.3% diagnostic yield tripled conventional estimates, demonstrating WES efficacy in identifying SNVs and CNVs in severe phenotypes. These findings reveal the genetic heterogeneity of severe hypospadias and support WES utility in uncovering novel variants and structural genomic alterations.

Recovery from cuprizone induced incontinence is not dependent upon remyelination.

MYRF controls mesothelium specification, signaling, and plasticity in lung development.

Myelin, along with the oligodendrocytes (OLs) that produce it, is essential for proper central nervous system (CNS) function in vertebrates. Although the accurate targeting of myelin to axons and its maintenance are critical for CNS performance, the molecular pathways that regulate these processes remain poorly understood. Through a combination of zebrafish genetics, mouse models, and primary OL cultures, we find that FBXW7, a recognition subunit of an E3 ubiquitin ligase complex, is a regulator of adult myelination in the CNS. Loss of Fbxw7 in myelinating OLs results in increased myelin sheath lengths with no change in myelin thickness. As the animals age, they develop progressive abnormalities including myelin outfolds, disrupted paranodal organization, and ectopic ensheathment of neuronal cell bodies with myelin. Through biochemical studies we find that FBXW7 directly binds and degrades the N-terminus of Myelin Regulatory Factor (N-MYRF), to control the balance between OL myelin growth and homeostasis.

MYRF gene mutations can lead to the development of Cardio-Urogenital Syndrome (CUGS), characterized by congenital heart disease, abnormalities in the internal and external reproductive organs, and ocular anomalies. CUGS can manifest with various types of congenital heart diseases, such as Tetralogy of Fallot, Scimitar syndrome, Hypoplastic Left Heart Syndrome, Atrial Septal Defect, Ventricular Septal Defect, Dextrocardia, Aortic Arch Anomalies, and Pulmonary Vein Anomalies et al. Male patients (46,XY) may present with unilateral cryptorchidism, ambiguous genitalia, or even typical female genitalia. The most common ocular issues consistent with high myopia and nanophthalmos. To date, there have been reports of over 30 cases of MYRF gene mutations worldwide. A 5-month-old female infant was admitted to Beijing Children's Hospital affiliated with Capital Medical University due to "rapid breathing since birth." No targeted therapy was administered prior to admission despite persistent tachypnea. Four months before admission, the infant was seen at a local hospital due to abnormalities in the external reproductive organs. Chromosomal analysis revealed 46, XY, and whole exome gene testing showed a MYRF gene mutation, identified heterozygosity for de novo mutations in the MYRF gene, splice site variant, respectively, that was not found in the gnomAD database, clinically presenting as Cardio-Urogenital Syndrome (GUGS; OMIM: 618280), inherited in an autosomal dominant (AD) manner. Family history was negative. The infant's showed severely breathing progressively, leading to admission to our hospital. Electrocardiography showed pathological Q waves in leads I, avL, and V4-V6. Echocardiography revealed congenital heart disease of anomalous origin of the left coronary artery from the pulmonary artery, severe left ventricular enlargement (left ventricular end-diastolic diameter 35.2mm), mild-to-moderate mitral regurgitation, and slightly reduced left ventricular systolic function (EF: 54%, which is just below the normal threshold 55%). Subsequent coronary angiography indicated the left coronary artery originating from the main pulmonary artery. The infant underwent further surgical treatment which confirmed the prior diagnosis and had a good postoperative recovery. This case represents an exceptionally rare instance of MYRF-CUGS combined with coronary artery anomaly. This case report enriches the clinical phenotype spectrum of CUGS caused by MYRF gene mutations and improves recognition among clinicians.

Neuroaxonal injury is a major driver of irreversible disability in demyelinating conditions. Accurate assessment of the association between demyelination and axonal pathology is critical for evaluating and developing effective therapeutic approaches. Measuring neurofilament light chain (NfL) in the blood could putatively allow longitudinal monitoring of neuroaxonal injury at ‘single protein resolution’ with high pathological specificity. Here, we demonstrate a robust association between blood and tissue NfL-based assessment of neuroaxonal injury and severity of inflammatory demyelination in experimental autoimmune encephalitis (EAE).In EAE, high levels of NfL were evident at the peak of demyelination and correlated with tissue evidence of NfL loss when using antibodies that target the same NfL epitopes. In addition, we validate the longitudinal NfL dynamics in relation to de- and remyelination in an inducible genetic model of inflammatory-independent myelin loss. Through inducible knockout of myelin regulatory factor (Myrf) in proteolipid protein (PLP) expressing cells in Myrffl/fl PLP1-CreERT (MyrfΔiPLP) mice, serum NfL peaked at the time of demyelination and reduced following effective remyelination. In people with multiple sclerosis, the most common demyelinating condition, we confirmed the association between NfL and myelin breakdown proteins in two independent cohorts using Olink proximity extension assays, the ReBUILD clinical trial and the multiple sclerosis participants in the UK-Biobank.Our study provides a translational framework to understand the biology behind NfL changes in the context of de- and remyelination and reveals novel aspects related to monitoring potentially reversible neuroaxonal pathology in humans and rodents.

Myelin regulatory factor (Myrf) is a critical transcription factor in early retinal and retinal pigment epithelial development, and human variants in MYRF are a cause for nanophthalmos. Single cell RNA sequencing (scRNAseq) was performed on Myrf conditional knockout mice (Rx > Cre Myrffl/fl) at 3 developmental timepoints. Myrf was expressed specifically in the RPE, and expression was abrogated in Rx > Cre Myrffl/fl eyes. scRNAseq analysis revealed a loss of RPE cells at all timepoints resulting from cell death. GO-term analysis in the RPE revealed downregulation of melanogenesis and anatomic structure morphogenesis pathways, which were supported by electron microscopy and histologic analysis. Novel structural target genes including Ermn and Upk3b, along with macular degeneration and inherited retinal disease genes were identified as downregulated, and a strong upregulation of TGFß/BMP signaling and effectors was observed. Regulon analysis placed Myrf downstream or parallel to Pax6 and Mitf and upstream of Sox10 in RPE differentiation. Together, these results suggest a strong role for MYRF in the RPE maturation by regulating melanogenesis, cell survival, and cell structure, in part acting through suppression of TGFß signaling and activation of Sox10.

46,XY differences/disorders of sex development (DSD) are genetically heterogeneous conditions characterized by atypical development of the reproductive system. MYRF, a gene encoding a transcription factor, has been identified as a potential causative gene for DSD and cardiac urogenital syndrome (CUGS). This study aims to delineate the clinical manifestations of patients with 46,XY DSD and MYRF mutations, encompassing both from our cohort and cases reported in the literature. Patients with 46,XY DSD were recruited from Peking Union Medical College Hospital and identified through a comprehensive review of published literature. Whole-exome sequencing was conducted to elucidate the genetic etiology. Comprehensive clinical data, including physical examination findings, hormonal profiles, and imaging results, were retrospectively gathered from medical records and published sources. Three of our patients with 46,XY DSD were found to harbor heterozygous loss-of-function mutations in the MYRF gene, including the recurrent variant c.789dup and two novel variants c.1915C>T and c.2154del. The patients exhibited underdeveloped testicular tissue, inadequate masculinization, and the persistence of Müllerian ducts. A review of the literature indentified 31 MYRF-linked 46,XY DSD patients and two 46,XX DSD patients. Among these, 11 cases presented with isolated testicular dysgenesis, 20 cases exhibited severe cardiopulmonary issues, and the majority of patients had congenital diaphragmatic hernia. Genetic analysis revealed 26 distinct MYRF variants among these patients, including 10 missense, 8 frameshift, 5 nonsense, and 3 splice site alterations, affecting critical domains of the MYRF gene. Our study broadens the spectrum of MYRF mutations in 46,XY DSD patients and highlighting the gene's indispensable role in gonadal development. However, a clear genotype-phenotype correlation in MYRF-related DSD remain elusive.

Cardiac-Urogenital Syndrome (CUGS) is a recently identified genetic disease characterized by urogenital, diaphragmatic, ophthalmic, and cardiac abnormalities caused by heterozygous pathogenic variants in the Myelin Regulatory Factor (MYRF) gene. The complete spectrum of disease characteristics and prevalence is not yet defined. This report documents the first known cases of anomalous origin of the left coronary artery from the pulmonary artery (ALCAPA) in MYRF-associated Cardiac-Urogenital Syndrome (MYRF-CUGS).

Background/AimsTo distinguish the clinical feature of nanophthalmos (NNO) caused by mutations in protease serine 56 (PRSS56), membrane-type frizzled-related protein (MFRP), myelin regulatory factor (MYRF) and transmembrane protein 98 (TMEM98) and...

Myelin, along with the oligodendrocytes (OLs) that produce it, is essential for proper central nervous system (CNS) function in vertebrates. Although the accurate targeting of myelin to axons and its maintenance are critical for CNS performance, the molecular pathways that regulate these processes remain poorly understood. Through a combination of zebrafish genetics, mouse models, and primary OL cultures, we found FBXW7, a recognition subunit of an E3 ubiquitin ligase complex, is a regulator of adult myelination in the CNS. Loss of Fbxw7 in myelinating OLs resulted in increased myelin sheath lengths with no change in myelin thickness. As the animals aged, they developed progressive abnormalities including myelin outfolds, disrupted paranodal organization, and ectopic ensheathment of neuronal cell bodies with myelin. Through biochemical studies we found that FBXW7 directly binds and degrades the N-terminal of Myelin Regulatory Factor (N-MYRF), to control the balance between oligodendrocyte myelin growth and homeostasis.

Preventing production of new oligodendrocytes impairs remyelination and sustains behavioural deficits after demyelination

Nanophthalmos-Associated MYRF gene mutation facilitates intraocular inflammation in mice

Growing evidence indicates that non‐neuronal oligodendrocyte plays an important role in Amyotrophic lateral sclerosis (ALS) and other neurodegenerative diseases. In patient's brain, the impaired myelin structure is a pathological feature with the observation of TDP‐43 in cytoplasm of oligodendrocyte. However, the mechanism underlying the gain of function by TDP‐43 in oligodendrocytes, which are vital for the axonal integrity, remains unclear. Recently, we found that the primate‐specific cleavage of truncated TDP‐43 fragments occurred in cytoplasm of monkey neural cells. This finding opened up the avenue to investigate the myelin integrity affected by pathogenic TDP‐43 in oligodendrocytes. In current study, we demonstrated that the truncated TDP‐35 in oligodendrocytes specifically, could lead to the dysfunctional demyelination in corpus callosum of monkey. As a consequence of the interaction of myelin regulatory factor with the accumulated TDP‐35 in cytoplasm, the downstream myelin‐associated genes expression was downregulated at the transcriptional level. Our study aims to investigate the potential effect on myelin structure injury, affected by the truncated TDP‐43 in oligodendrocyte, which provided the additional clues on the gain of function during the progressive pathogenesis and symptoms in TDP‐43 related diseases.

Myrf regulates RPE development, melanogenesis, and is important for cell structure and survival, in part through regulation of Ermn , Upk3b and Sox10, and BMP/TGFb signaling.

To explore the genetic basis for a fetus with Cardiac-urogenital syndrome (CUGS). A fetus with congenital heart disease identified at the Maternal Fetal Medical Center for Fetal Heart Disease, Beijing Anzhen Hospital Affiliated to Capital Medical University in January 2019 was selected as the study subject. Clinical data of the fetus was collected. Copy number variation sequencing (CNV-seq) and trio-whole exome sequencing (trio-WES) were carried out for the fetus and its parents. Candidate variants were verified by Sanger sequencing. Detailed fetal echocardiographic examination had revealed hypoplastic aortic arch. The results of trio-WES revealed that the fetus has harbored a de novo splice variant of the MYRF gene (c.1792-2A>C), for which both parents were of the wild-type. Sanger sequencing confirmed the variant to be de novo. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was rated as likely pathogenic. CNV-seq has identified no chromosomal anomalies. And the fetus was diagnosed with Cardiac-urogenital syndrome. The de novo splice variant of the MYRF gene probably underlay the abnormal phenotype in the fetus. Above finding has enriched the spectrum of MYRF gene variants.

BackgroundOligodendrocytes have robust regenerative ability and are key players in remyelination during physiological and pathophysiological states. However, the mechanisms of brain microenvironmental cue in regulation of the differentiation of oligodendrocytes still needs to be further investigated.ResultsWe demonstrated that myelin-associated glycoprotein (MAG) was a novel receptor for angiopoietin-like protein 2 (ANGPTL2). The binding of ANGPTL2 to MAG efficiently promoted the differentiation of oligodendrocytes in vitro, as evaluated in an HCN cell line. Angptl2-null mice had a markedly impaired myelination capacity in the early stage of oligodendrocyte development. These mice had notably decreased remyelination capacities and enhanced motor disability in a cuprizone-induced demyelinating mouse model, which was similar to the Mag-null mice. The loss of remyelination ability in Angptl2-null/Mag-null mice was similar to the Angptl2-WT/Mag-null mice, which indicated that the ANGPTL2-mediated oligodendrocyte differentiation effect depended on the MAG receptor. ANGPTL2 bound MAG to enhance its phosphorylation level and recruit Fyn kinase, which increased Fyn phosphorylation levels, followed by the transactivation of myelin regulatory factor (MYRF).ConclusionOur study demonstrated an unexpected cross-talk between the environmental protein (ANGPTL2) and its surface receptor (MAG) in the regulation of oligodendrocyte differentiation, which may benefit the treatment of many demyelination disorders, including multiple sclerosis.

The atypical protein tyrosine phosphatase enzyme, dual-specificity phosphate 15 (DUSP15) is thought to be activated by myelin regulatory factor (MyRF) and to have a role in oligodendrocyte differentiation. Here, we assess whether Dusp15 is reduced in the hippocampus of mice with conditional knock-out of Myrf in oligodendrocyte precursor cells. Using quantitative polymerase chain reaction (qPCR) we found that Dusp15 expression was indeed lower in these mice. Alterations in myelin have been associated with Alzheimer's disease (AD), autism spectrum disorder (ASD) and attention deficit/hyperactivity disorder (ADHD). Symptoms of these disorders can include impairments of object recognition and attention. We, therefore tested the mice in the object recognition task (ORT) and 5-choice serial reaction time task (5CSRTT). However, we did not find behavioural impairments indicating that attentional abilities and object recognition are not impacted by reduced oligodendrogenesis and hippocampal Dusp15 expression. Gaining insight into the role of newly formed oligodendrocytes and Dusp15 expression is helpful for the development of well targeted treatments for myelin dysregulation.

HIV-associated neurocognitive disorders (HAND) remain pervasive even with increased efficacy/use of antiretroviral therapies. Opioid use/abuse among HIV + individuals is documented to exacerbate CNS deficits. White matter (WM) alterations, including myelin pallor, and volume/structural alterations detected by diffusion tensor imaging are common observations in HIV + individuals, and studies in non-human primates suggest that WM may harbor virus. Using transgenic mice that express the HIV-1 Tat protein, we examined in vivo effects of 2-6weeks of Tat and morphine exposure on WM using genomic and biochemical methods. RNA sequencing of striatal tissue at 2weeks revealed robust changes in mRNAs associated with oligodendrocyte precursor populations and myelin integrity, including those for transferrin, the atypical oligodendrocyte marker N-myc downstream regulated 1 (Ndrg1), and myelin regulatory factor (Myrf/Mrf), an oligodendrocyte-specific transcription factor with a significant role in oligodendrocyte differentiation/maturation. Western blots conducted after 6-weeks exposure in 3 brain regions (striatum, corpus callosum, pre-frontal cortex) revealed regional differences in the effect of Tat and morphine on Myrf levels, and on levels of myelin basic protein (MBP), whose transcription is regulated by Myrf. Responses included individual and interactive effects. Although baseline and post-treatment levels of Myrf and MBP differed between brain regions, post-treatment MBP levels in striatum and pre-frontal cortex were compatible with changes in Myrf activity. Additionally, the Myrf regulatory ubiquitin ligase Fbxw7 was identified as a novel target in our model. These results suggest that Myrf and Fbxw7 contribute to altered myelin gene regulation in HIV.

BackgroundCardiac‐urogenital syndrome [MIM # 618280] is a newly described very rare syndrome associated with pathogenic variants in the myelin regulatory factor (MYRF) gene that leads to loss of protein function. MYRF is a transcription factor previously associated only with the control of myelin‐related gene expression. However, it is also highly expressed in other tissues and associated with various organ anomalies. The clinical picture is primarily dominated by complex congenital cardiac developmental defects, pulmonary hypoplasia, congenital diaphragmatic hernia, and urogenital malformations.Case PresentationWe present case reports of two siblings of unrelated parents in whom whole‐exome sequencing was indicated due to familial occurrence of extensive developmental defects. A new, previously undescribed splicing pathogenic variant c.1388+2T>G in the MYRF gene has been identified in both patients. Both parents are unaffected, tested negative, and have another healthy daughter. The identical de novo event in siblings suggests gonadal mosaicism, which can mimic recessive inheritance.ConclusionsTo our knowledge, this is the first published case of familial cardiac‐urogenital syndrome indicating gonadal mosaicism.