Customizing carrier screening in the Chinese population: Insights from a 334-gene panel.

OBJECTIVE: This study aimed to evaluate the yield and applicability of expanded carrier screening and propose carrier rate screening thresholds suitable for the Chinese population by comparing the current screening panel with the American College of Medical Genetics and Genomics recommended panel of 113 genes. METHODS: Using targeted next-generation sequencing, a customized panel with 334 genes was performed on 2168 individuals without clinical phenotypes for expanded carrier screening purpose. Variant interpretation followed the American College of Medical Genetics and Genomics guidelines. Carrier rates were calculated for each identified variant and each gene. At-risk couple rates were also assessed. The yield of expanded carrier screening was evaluated through calculating cumulative carrier rate. RESULTS: Overall, 65.87% of the individuals were found to be carriers of at least 1 disease causing variants. The overall at-risk couple rate was 11.76%, of which the GJB2:c.109G > A related at-risk couple rate was 5.78%. The cumulative carrier rate of 334-panel was 65.53%. When screened genes with gene carrier rate ≥1/1000, the expanded carrier screening can cover over 90% of the cumulative carrier rate and at-risk couples. A total of 86 genes overlapped with American College of Medical Genetics and Genomics Tier-3 genes and were attributed to the cumulative carrier rate of 47.33%. CONCLUSION: Expanded carrier screening using the 334-gene panel showed high screening efficiency. A threshold of gene carrier rate ≥1/1000 is recommended for selecting carrier screening genes in the Chinese Han population. This study highlights the importance of customizing screening panels based on the ACMG Tier-3 genes in conjunction with population-specific carrier frequencies to improve the accuracy and effectiveness of expanded carrier screening.

Genetic etiology of agenesis of the corpus callosum: a retrospective single-center cohort analysis of 114 fetuses.

PURPOSE: The identification and prognosis of the agenesis of the corpus callosum (ACC) for prenatal consultation are complex and currently unclear. This study aims to explore the correlated genetic mutations of prenatal ACC. METHODS: We retrospectively analyzed 114 prenatal cases of ACC. All cases (n = 114) were subjected to chromosomal microarray analysis (CMA), and 66 CMA-negative cases underwent prenatal exome sequencing (pES) for further analysis. RESULTS: CMA was diagnosed positively in 15/114 (13.2%) cases and pES was diagnosed positively in 24/66 (36.4%) CMA-negative cases. The detection rate of genetic causes between complete and partial ACCs was not significantly different (P > 0.05). Between isolated and non-isolated (other anomalies present) ACCs, the diagnostic rate of pES in non-isolated cases was significantly higher (P < 0.001), while CMA results did not differ (P > 0.05). The diagnostic rate of CMA was significantly increased in cases combined with intracranial and extracranial malformations (P = 0.014), while no CMA positivity was detected in cases combined with only intracranial malformations. CONCLUSION: For fetuses with prenatal ACC, further pES analysis should be recommended after negative CMA results. Chromosome abnormalities are less likely to occur when ACC with only intracranial malformations combined.

Identification of copy number variations among fetuses with isolated ultrasound soft markers in pregnant women not of advanced maternal age.

BACKGROUND: Pathogenic (P) copy number variants (CNVs) may be associated with second-trimester ultrasound soft markers (USMs), and noninvasive prenatal screening (NIPS) can enable interrogate the entire fetal genome to screening of fetal CNVs. This study evaluated the clinical application of NIPS for detecting CNVs among fetuses with USMs in pregnant women not of advanced maternal age (AMA). RESULTS: Fetal aneuploidies and CNVs were identified in 6647 pregnant women using the Berry Genomics NIPS algorithm.Those with positive NIPS results underwent amniocentesis for prenatal diagnosis. The NIPS and prenatal diagnosis results were analyzed and compared among different USMs. A total of 96 pregnancies were scored positive for fetal chromosome anomalies, comprising 37 aneuploidies and 59 CNVs. Positive predictive values (PPVs) for trisomy 21, trisomy 18, trisomy 13, and sex chromosome aneuploidies were 66.67%, 80.00%, 0%, and 30.43%, respectively. NIPS sensitivity for aneuploidies was 100%. For CNVs, the PPVs were calculated as 35.59% and false positive rate of 0.57%. There were six P CNVs, two successfully identified by NIPS and four missed, of which three were below the NIPS resolution limit and one false negative. The incidence of aneuploidies was significantly higher in fetuses with absent or hypoplastic nasal bone, while that of P CNVs was significantly higher in fetuses with aberrant right subclavian artery (ARSA), compared with other groups. CONCLUSIONS: NIPS yielded a moderate PPV for CNVs in non-AMA pregnant women with fetal USM. However, NIPS showed limited ability in identifying P CNVs. Positive NIPS results for CNVs emphasize the need for further prenatal diagnosis. We do not recommend the use of NIPS for CNVs screening in non-AMA pregnant women with fetal USM, especially in fetuses with ARSA.

Maternal CHD7 gonosomal mosaicism in a fetus with CHARGE syndrome.

Parental mosaicism is important in families with de novo mutations. Herein, we report a case of fetal CHARGE syndrome (CS) with a CHD7 variant inherited from maternal CHD7 gonosomal mosaicism. The variant was detected through trio-based whole-exome sequencing and Sanger sequencing. High-depth whole-exome sequencing was performed for the identification of parental mosaicism. A novel heterozygous CHD7 nonsense mutation (c.5794G>T/ p.E1932*) was detected in the tissue from the aborted fetus. The parents were wild-type, indicating that the mutation was a de novo variant. The mutation was suspected to be the cause of the fetal CS. However, high-depth whole-exome sequencing revealed maternal gonosomal mosaicism at a variant allele frequency of 3.2%-23.3%. The variant was identified in various tissues (peripheral blood, hair follicles, buccal epithelia, and pharyngeal epithelia) from the asymptomatic mother. We confirmed maternal CHD7 gonosomal mosaicism as a genetic cause of fetal CS. Our results emphasize the importance of clinical analysis in accurately determining the parents' status in detecting the CHD7 de novo variant in fetal CS, as this analysis has vital implications for evaluating the recurrence risk for genetic counseling.

Fresh Washed Microbiota Transplantation Alters Gut Microbiota Metabolites to Ameliorate Sleeping Disorder Symptom of Autistic Children.

Accumulating studies have raised concerns about gut dysbiosis associating autism spectrum disorder (ASD) and its related symptoms. However, the effect of gut microbiota modification on the Chinese ASD population and its underlying mechanism were still elusive. Herein, we enrolled 24 ASD children to perform the first course of fresh washed microbiota transplantation (WMT), 18 patients decided to participate the second course, 13 of which stayed to participate the third course, and there were 8 patients at the fourth course. Then we evaluated the effects of fresh WMT on these patients and their related symptoms. Our results found that the sleeping disorder symptom was positively interrelated to ASD, fresh WMT significantly alleviated ASD and its sleeping disorder and constipation symptoms. In addition, WMT stably and continuously downregulated Bacteroides/Flavonifractor/Parasutterella while upregulated Prevotella_9 to decrease toxic metabolic production and improve detoxification by regulating glycolysis/myo-inositol/D-glucuronide/D-glucarate degradation, L-1,2-propanediol degradation, fatty acid ß-oxidation. Thus, our results suggested that fresh WMT moderated gut microbiome to improve the behavioral and sleeping disorder symptoms of ASD via decrease toxic metabolic production and improve detoxification. Which thus provides a promising gut ecological strategy for ASD children and its related symptoms treatments.

Maternal, neonatal, pregnancy outcome characteristics of pregnant women with high plasma cell-free DNA concentration in non-invasive prenatal screening: a retrospective analysis.

Objective: Cell-free DNA (cfDNA) is a useful biomarker in various clinical contexts. Herein, we aimed to identify maternal characteristics and pregnancy outcomes associated with a failed NIPS test due to high cfDNA concentrations. Methods: A retrospective study of cases with high plasma cfDNA concentration in pregnant women in which NIPS test was performed (from 174,318 cases). We reported the detection of 126 cases (118 with complete clinical information) in which the high amount of cfDNA did not allow the performance of NIPS and study the possible causes of this result. Results: 622 (0.35%) of 174,318 pregnant women had failed the NIPS test, including 126 (20.3%) cases with high plasma cfDNA concentrations. The failed NIPS due to high plasma cfDNA concentrations was associated with maternal diseases and treatment with low-molecular-weight heparin (LMWH). Further follow-up of the 118 pregnant women in the case group revealed that the pregnancy outcomes included 31 premature deliveries, 21 abortions. The cfDNA concentrations of pregnant women with preterm deliveries were 1.15 (0.89, 1.84), which differed significantly from those who had full-term deliveries. Conclusions: Among pregnant women with high cfDNA concentrations, systemic autoimmune diseases, pregnancy complications and LMWH were associated with increased incidence of failed NIPS test. High maternal cfDNA concentrations may not be associated with chromosomal abnormalities in the fetus. However, they should be alerted to the possibility of preterm births and stillbirths. Further clinical studies on pregnant women with high cfDNA concentrations are required.

Defining the scope of extended NIPS in Western China: evidence from a large cohort of fetuses with normal ultrasound scans.

BACKGROUND: Standard noninvasive prenatal screening(NIPS) is an accurate and reliable method to screen for common chromosome aneuploidies, such as trisomy 21, 18 and 13. Extended NIPS has been used in clinic for not only aneuploidies but also copy number variants(CNVs). Here we aim to define the range of chromosomal abnormalities that should be able to identify by NIPS in order to be an efficient extended screening test for chromosomal abnormalities. METHODS: A prospective study was conducted, involving pregnant women without fetal sonographic structural abnormalities who underwent amniocentesis. Prenatal samples were analyzed using copy number variation sequencing(CNV-seq) to identify fetal chromosomal abnormalities. RESULTS: Of 28,469 pregnancies included 1,022 (3.59%) were identified with clinically significant fetal chromosome abnormalities, including 587 aneuploidies (2.06%) and 435 (1.53%) pathogenic (P) / likely pathogenic (LP) CNVs. P/LP CNVs were found in all chromosomes, but the distribution was not uniform. Among them, P/LP CNVs in chromosomes 16, 22, and X exhibited the highest frequencies. In addition, P/LP CNVs were most common on distal ends of the chromosomes and in low copy repeat regions. Recurrent microdeletion/microduplication syndromes (MMS) accounted for 40.69% of total P/LP CNVs. The size of most P/LP CNVs (77.47%) was < 3 Mb. CONCLUSIONS: In addition to aneuploidies, the scope of extended NIPS should include the currently known P/LP CNVs, especially the regions with recurrent MMS loci, distal ends of the chromosomes, and low copy repeat regions. To be effective detection should include CNVs of < 3 Mb. Meanwhile, sufficient preclinical validation is still needed to ensure the clinical effect of extended NIPS.

A novel mutation in CFAP47 causes male infertility due to multiple morphological abnormalities of the sperm flagella.

Introduction: A previous study suggested that loss of CFAP47 function is involved in multiple morphological abnormalities of the sperm flagella (MMAF) in humans and mice. However, the comprehensive role of CFAP47 in spermatogenesis is largely unknown. Methods: Whole-exome sequencing (WES) was conducted to identify pathogenic variant in two patients with MMAF. The functional effect of the identified mutations was investigated by immunofluorescence staining and western blotting. Intracytoplasmic sperm injection (ICSI) was used to assist fertilization for the patient with MMAF. Results: In this study, we identified a novel missense mutation (c.1414G>A; p.V472M) in CFAP47 in two unrelated patients with oligoasthenoteratozoospermia. Intriguingly, in addition to the MMAF phenotype very analogous to the previous report, the two patients notably presented abnormal morphology of sperm heads, the sperm mitochondrial sheath was obviously disorganized, and the sperm annulus were almost defective. Further functional experiments confirmed that the expression of CFAP47 was markedly reduced in the spermatozoa of the patients. Mechanism analysis suggested that CFAP47 might regulate the expression of CFAP65, CFAP69 and SEPTIN4 through their physical interactions and thus modulating sperm morphogenesis. Conclusion: we revealed a novel mutation in CFAP47 and further expanded the phenotype and mutation spectrum of CFAP47, as well as the potential mechanism of CFAP47 manipulating spermatogenesis, finally providing important guidance for genetic counseling and targeted treatment for CFAP47 mutation-related male infertility.

Maternal and fetal factors influencing fetal fraction: A retrospective analysis of 153,306 pregnant women undergoing noninvasive prenatal screening.

Background: Genetic factors are important causes of birth defects. Noninvasive prenatal screening (NIPS) is widely used for prenatal screening of trisomy 21, trisomy 18, and trisomy 13, which are the three most common fetal aneuploidies. Fetal fraction refers to the proportion of cell-free fetal DNA in maternal plasma, which can influence the accuracy of NIPS. Elucidating the factors that influence fetal fraction can provide guidance for the interpretation of NIPS results and genetic counseling. However, there is currently no broad consensus on the known factors that influence fetal fraction. Objective: The study aimed to explore the maternal and fetal factors influencing fetal fraction. Methods: A total of 153,306 singleton pregnant women who underwent NIPS were included. Data on gestational age; maternal age; body mass index (BMI); z-scores for chromosomes 21, 18, and 13; and fetal fraction in NIPS were collected from the study population, and the relationships between fetal fraction and these factors were examined. The relationship between fetal fraction and different fetal trisomy types was also analyzed. Results: The results showed that the median gestational age, maternal age, and BMI of the pregnant women were 18 (16, 20) weeks, 29 (25, 32) years, and 22.19 (20.40, 24.24) kg/m2, respectively. The median fetal fraction was 11.62 (8.96, 14.7)%. Fetal fraction increased with gestational age and decreased with maternal age and BMI (P < 0.001). Fetal fraction of fetuses with trisomies 21, 18, and 13 was similar to that of the NIPS-negative group. The z-scores of pregnant women with trisomy 21 and 18 fetuses were positively correlated with fetal fraction, but not with that of the trisomy 13 cases. Conclusions: The factors that influence fetal fraction need to be taken into consideration before NIPS for quality control and after NIPS for result interpretation.

Anemia Is Associated with Disease Severity, Hepatic Complications, and Progression of Wilson Disease: A Retrospective Cohort Study.

INTRODUCTION: Anemia is a common manifestation of chronic liver diseases. It is a predictor of severe disease, a high risk of complications, and poor outcomes in various liver diseases. However, it remains unclear whether anemia serves as a similar indicator in patients with Wilson disease (WD). Therefore, this study aimed to investigate the relationship between anemia and severity, hepatic complications, and the progression of WD. METHODS: Medical data were collected retrospectively from January 1, 2016, to December 31, 2020. Univariate and multivariate analyses were carried out to investigate the relationship between anemia and liver-associated disease severity, hepatic complications, and the progression of WD. RESULTS: A total of 288 WD patients (48 with and 240 without anemia) were enrolled in the study. Multivariate linear regression revealed that WD patients with anemia had significantly higher levels of bilirubin, alanine transaminase, prothrombin time, international normalized ratio, type Ⅳ collagen, and hyaluronic acid and significantly lower levels of albumin, total cholesterol, and high-density lipoprotein-cholesterol (all p < 0.05). Multivariate logistic regression showed that anemia was a risk factor for gastric varices and ascites (all p < 0.05). Fully adjusted Cox regression revealed that anemia was an independent risk factor for advanced Child-Pugh classification (p = 0.034). CONCLUSIONS: Anemia was common in WD patients and was associated with greater disease severity, a higher risk of hepatic complications, and a faster progression.

Reduced type 3 innate lymphoid cells related to worsening kidney function in renal dysfunction.

Intestinal mucosa barrier injury and immunity imbalance contribute to chronic kidney disease (CKD) progression. Type 3 innate lymphoid cells (ILC3s) are essential for normal intestinal homeostasis. Nevertheless, the relationship between ILC3s and CKD remains largely unknown. The aim of this study was to investigate the relationship linking ILC3s to clinical indicators among patients with renal dysfunction. The levels of circulating ILC3s and dendritic cells, as well as their subsets, in patients with renal dysfunction and healthy controls were determined through flow cytometry. The levels of human plasma granulocyte-macrophage colony-stimulating factor (GM-CSF) were measured using enzyme-linked immunosorbent assay. Renal function was evaluated by measuring the estimated glomerular filtration rate (eGFR), as well as the levels of serum creatinine, blood urea nitrogen (BUN), and uric acid. The results revealed that the proportion of peripheral ILC3s was significantly decreased in patients with renal dysfunction. This reduction was positively associated with the levels of eGFR, and inversely associated with the levels of BUN and uric acid. Similarly, the percentage of circulating C-C motif chemokine receptor 6-positive (CCR6 +) ILC3s was also obviously reduced, and demonstrated positive and negative associations with the levels of eGFR and BUN, respectively. Furthermore, the levels of CCR6 + ILC3s correlated positively with those of GM-CSF, as well as type 1 conventional dendritic cells (cDC1s), which also decreased in parallel with kidney function. Thus, the reduction of ILC3s, particularly CCR6 + ILC3s, was related to worsening kidney function in patients with renal dysfunction. This effect may delay renal function impairment by regulating cDC1s via the secretion of GM-CSF, indicating that CCR6 + ILC3s may serve as efficient biomarkers for evaluating kidney function.

Identification of POLR3B biallelic mutations-associated hypomyelinating leukodystrophy-8 in two siblings.

POLR3B gene encodes the 2nd largest catalytic subunit and affects the function of RNA polymerase III enzymes in transcription. Bi-allelic variants in POLR3B pathogenically cause hypomyelinating leukodystrophy-8 (HLD8). Herein, we recruited a family with two patients, who presented clinically with cerebellar atrophy, intellectual disability, hypogonadotropic hypogonadism, and visual problems. We identified the two affected siblings carrying the compound heterozygous variations (c.165_167del; c.1615G>T) in POLR3B by trio-whole-exome sequencing (trio-WES). The qPCR and western blot showed that both transcriptional and translational levels of the mutation (c.165_167del, p.I55_K56delinsM) were sharply attenuated. Following that, a thorough functional examination of a zebrafish line disrupted for human POLR3B validated the pathogenic effects of the two mutations. Our research broadens the spectrum of HLD8-related pathogenic POLR3B mutations and provides new molecular and animal evidence.

Type 3 innate lymphoid cells as an indicator of renal dysfunction and serum uric acid in hyperuricemia.

BACKGROUND: Type 3 innate lymphoid cells (ILC3s) are a newly identified group of innate immune cells that participate in the progression of several metabolic diseases by secreting interleukin (IL)-17 and IL-22. These cytokines are associated with hyperuricemia (HUA) severity and development; however, the relationship between ILC3s and HUA remains unclear. OBJECTIVES: To determine the characteristics of circulating ILC3s in patients with HUA. MATERIAL AND METHODS: Type 3 innate lymphoid cells and their subsets were detected using flow cytometry in peripheral blood mononuclear cells (PBMCs) of 80 HUA patients and 30 healthy controls (HC). Plasma levels of IL-17A and IL-22 were measured with enzyme-linked immunosorbent assay (ELISA). Clinical data of enrolled subjects were collected from electronic medical records. RESULTS: In patients with HUA, the frequency of circulating ILC3s was elevated and positively correlated with levels of serum uric acid and serum creatinine (Scr). Although there was no significant difference in the plasma concentration of IL-17A between the patients with HUA and healthy controls, positive correlations between plasma IL-17A and the concentration of serum uric acid and frequency of circulating ILC3s were observed in the patients with HUA. CONCLUSIONS: In patients with HUA, positive correlations were detected between circulating ILC3 levels, plasma IL-17A and serum uric acid. Therefore, ILC3s and IL-17A may be useful indicators of disease severity, and are potential new therapeutic targets in HUA.

Novel biallelic mutations in TTC29 cause asthenoteratospermia and male infertility.

BACKGROUND: Multiple morphological abnormalities of the sperm flagella (MMAF), which is characterized as asthenoteratospermia involving absent, short, bent, coiled, and/or irregular-caliber flagella, is a rare recessive inherited disorder associated with male infertility. To date, genetic causes of MMAF cases are not fully explored. METHODS: Whole-exome sequencing was conducted to identify pathogenic variants in a patient with MMAF. The functional effect of the identified mutations was investigated by immunofluorescence staining and western blotting. Intracytoplasmic sperm injection was used to assist fertilization for the patient with MMAF. RESULTS: We identified novel biallelic mutations, a splicing variant NC_000004.12:g.146937593C>T (c.254+1G>A), and a nonsense mutation NM_001300761.4:c.1185C>G (NP_001287690.1:p.Tyr395*), in TTC29 from an infertile patient. In addition to the typical MMAF phenotype, the patient also presented aberrant morphology of sperm heads. Further functional experiments confirmed the absence of TTC29 expression in the spermatozoa. We also explored the specific expression pattern of TTC29 in human and mouse spermatogenesis. The outcome of intracytoplasmic sperm injection in the patient was unsuccessful, while additional female risk factors should not be excluded. CONCLUSIONS: Our study revealed the novel biallelic mutations in TTC29 in a MMAF patient, which findings expand the mutational spectrum of TTC29 and further contribute to the diagnosis, genetic counseling, and prognosis of male infertility.

Coordination-Driven Self-Assembly of Iron Oxide Nanoparticles for Tumor Microenvironment-Responsive Magnetic Resonance Imaging.

Accurate diagnosis of diseases located in deep tissues is always challenging. The "always-on" probe often leads to false-positive signals due to nonspecific interaction of nanoprobes. Thus, stimuli-responsive nanoprobes are highly desirable, which, however, require complicated surface modification so as to achieve trigger-induced signal changes. Here pH-triggered switchable magnetic resonance imaging (MRI) nanoprobes were constructed by coordination-driven self-assembly of monodispersed iron oxide nanoparticles (MIONPs) with simple amino acid derivatives, which displayed typical T2-weighted MRI features, yet, were turned into T1-weighted MRI under slightly acidic conditions at the tumor site. The dynamic assembly and disassembly properties of MIONPs afford T2/T1 switchable contrast imaging, enabling selective "turn-on" signals at the tumor site with high specificity.

Impaired Function of a Rare Mutation in the MMUT Gene Causes Methylmalonic Acidemia in a Chinese Patient.

Methylmalonic acidemia (MMA) is an autosomal recessive metabolic disorder mainly caused by mutations in the methylmalonyl coenzyme A mutase (MCM) gene (MMUT) and leads to the reduced activity of MCM. In this study, a 3-year-old girl was diagnosed with carnitine deficiency secondary to methylmalonic acidemia by tandem mass spectrometry (MS/MS) and gas chromatography/mass spectrometry (GS/MS). Whole-exome sequencing (WES) was performed on the patient and identified two compound heterozygous mutations in MMUT: c.554C>T (p. S185F) and c.729-730insTT (p. D244Lfs ∗ 39). Bioinformatics analysis predicted that the rare missense mutation of c.554C>T would be damaging. Moreover, this rare mutation resulted in the reduced levels of MMUT mRNA and MMUT protein. Collectively, our findings provide a greater understanding of the effects of MMUT variants and will facilitate the diagnosis and treatment of patients with MMA.

Long-read Oxford nanopore sequencing reveals a de novo case of complex chromosomal rearrangement involving chromosomes 2, 7, and 13.

BACKGROUND: Complex chromosomal rearrangements (CCRs) are associated with high reproductive risk, infertility, abnormalities in offspring, and recurrent miscarriage in women. It is essential to accurately characterize apparently balanced chromosome rearrangements in unaffected individuals. METHODS: A CCR young couple who suffered two spontaneous abortions and underwent labor induction due to fetal chromosomal abnormalities was studied using long-read sequencing(LRS), single-nucleotide polymorphism (SNP) array, G-banding karyotype analysis (550-band resolution), and Sanger sequencing. RESULTS: SNP analysis of the amniotic fluid cells during the third pregnancy revealed a 9.9-Mb duplication at 7q21.11q21.2 and a 24.8-Mb heterozygous deletion at 13q21.1q31.1. The unaffected female partner was a carrier of a three-way CCR [46,XX,? ins(7;13)(q21.1;q21.1q22)t(2;13)(p23;q22)]. Subsequent LRS analysis revealed the exact breakpoint locations on the derivative chromosomes and the specific method of chromosome rearrangement, indicating that the CCR carrier was a more complex structural rearrangement comprising five breakpoints. Furthermore, LRS detected an inserted fragment of chromosome 13 in chromosome 7. CONCLUSIONS: LRS is effective for analyzing the complex structural variations of the human genome and may be used to clarify the specific CCRs for effective genetic counseling and appropriate intervention.

Identification of a Novel Missense Mutation of the PHEX Gene in a Large Chinese Family with X-Linked Hypophosphataemia.

X-linked hypophosphataemia (XLH) is an X-linked dominant rare disease that refers to the most common hereditary hypophosphatemia (HH) caused by mutations in the phosphate-regulating endopeptidase homolog X-linked gene (PHEX; OMIM: * 300550). However, mutations that have already been reported cannot account for all cases of XLH. Extensive genetic analysis can thus be helpful for arriving at the diagnosis of XLH. Herein, we identified a novel heterozygous mutation of PHEX (NM_000444.5: c.1768G > A) in a large Chinese family with XLH by whole-exome sequencing (WES). In addition, the negative effect of this mutation in PHEX was confirmed by both bioinformatics analysis and in vitro experimentation. The three-dimensional protein-model analysis predicted that this mutation might impair normal zinc binding. Immunofluorescence staining, qPCR, and western blotting analysis confirmed that the mutation we detected attenuated PHEX protein expression. The heterozygous mutation of PHEX (NM_000444.5: c.1768G > A) identified in this study by genetic and functional experiments constitutes a novel genetic cause of XLH, but further study will be required to expand its use in clinical and molecular diagnoses of XLH.

CEP128 is involved in spermatogenesis in humans and mice.

Centrosomal proteins are necessary components of the centrosome, a conserved eukaryotic organelle essential to the reproductive process. However, few centrosomal proteins have been genetically linked to fertility. Herein we identify a homozygous missense variant of CEP128 (c.665 G > A [p.R222Q]) in two infertile males. Remarkably, male homozygous knock-in mice harboring the orthologous CEP128R222Q variant show anomalies in sperm morphology, count, and motility. Moreover, Cep128 knock-out mice manifest male infertility associated with disrupted sperm quality. We observe defective sperm flagella in both homozygous Cep128 KO and KI mice; the cilia development in other organs is normal-suggesting that CEP128 variants predominantly affected the ciliogenesis in the testes. Mechanistically, CEP128 is involved in male reproduction via regulating the expression of genes and/or the phosphorylation of TGF-ß/BMP-signalling members during spermatogenesis. Altogether, our findings unveil a crucial role for CEP128 in male fertility and provide important insights into the functions of centrosomal proteins in reproductive biology.