Anomalous thermal transport under high pressure in boron arsenide.

High pressure represents extreme environments and provides opportunities for materials discovery1-8. Thermal transport under high hydrostatic pressure has been investigated for more than 100 years and all measurements of crystals so far have indicated a monotonically increasing lattice thermal conductivity. Here we report in situ thermal transport measurements in the newly discovered semiconductor crystal boron arsenide, and observe an anomalous pressure dependence of the thermal conductivity. We use ultrafast optics, Raman spectroscopy and inelastic X-ray scattering measurements to examine the phonon bandstructure evolution of the optical and acoustic branches, as well as thermal conductivity under varied temperatures and pressures up to 32 gigapascals. Using atomistic theory, we attribute the anomalous high-pressure behaviour to competitive heat conduction channels from interactive high-order anharmonicity physics inherent to the unique phonon bandstructure. Our study verifies ab initio theory calculations and we show that the phonon dynamics-resulting from competing three-phonon and four-phonon scattering processes-are beyond those expected from classical models and seen in common materials. This work uses high-pressure spectroscopy combined with atomistic theory as a powerful approach to probe complex phonon physics and provide fundamental insights for understanding microscopic energy transport in materials of extreme properties.

Deficiency of primate-specific SSX1 induced asthenoteratozoospermia in infertile men and cynomolgus monkey and tree shrew models.

Primate-specific genes (PSGs) tend to be expressed in the brain and testis. This phenomenon is consistent with brain evolution in primates but is seemingly contradictory to the similarity of spermatogenesis among mammals. Here, using whole-exome sequencing, we identified deleterious variants of X-linked SSX1 in six unrelated men with asthenoteratozoospermia. SSX1 is a PSG expressed predominantly in the testis, and the SSX family evolutionarily expanded independently in rodents and primates. As the mouse model could not be used for studying SSX1, we used a non-human primate model and tree shrews, which are phylogenetically similar to primates, to knock down (KD) Ssx1 expression in the testes. Consistent with the phenotype observed in humans, both Ssx1-KD models exhibited a reduced sperm motility and abnormal sperm morphology. Further, RNA sequencing indicated that Ssx1 deficiency influenced multiple biological processes during spermatogenesis. Collectively, our experimental observations in humans and cynomolgus monkey and tree shrew models highlight the crucial role of SSX1 in spermatogenesis. Notably, three of the five couples who underwent intra-cytoplasmic sperm injection treatment achieved a successful pregnancy. This study provides important guidance for genetic counseling and clinical diagnosis and, significantly, describes the approaches for elucidating the functions of testis-enriched PSGs in spermatogenesis.

Bi-allelic human TEKT3 mutations cause male infertility with oligoasthenoteratozoospermia owing to acrosomal hypoplasia and reduced progressive motility.

Oligoasthenoteratozoospermia (OAT) can result in male infertility owing to reduced sperm motility and abnormal spermatozoan morphology. The Tektins are a family of highly conserved filamentous proteins expressed in the axoneme and associated structures in many different metazoan species. Earlier studies on mice identified Tektin3 (Tekt3) as a testis-enriched gene, and knockout of Tekt3 resulted in asthenozoospermia in the mice. Here, whole-exome sequencing of 100 males with asthenozoospermia from unrelated families was performed, followed by Sanger sequencing, leading to the identification of TEKT3 as a candidate gene in two of these patients and their associated family members. In total, three mutations in the TEKT3 gene were identified in both these patients, including one homozygous deletion-insertion mutation (c.543_547delinsTTGAT: p.Glu182*) and one compound heterozygous mutation (c.[548G > A]; [752A > C], p.[Arg183Gln]; [Gln251Pro]). Both of these mutations resulted in the complete loss of TEKT3 expression. The patients were both found to produce sperm that, although those showed no apparent defects in the flagellar structure, had reduced progressive motility. In contrast to mice, most sperm from these two patients exhibited acrosomal hypoplasia, although this did not prevent the use of the sperm for in vitro fertilization through an ICSI approach. TEKT3 was found to bind to other TEKT proteins, suggesting that these proteins form a complex within human spermatozoa. Overall, these results suggest that a loss of TEKT3 function can contribute to OAT incidence in humans. TEKT3 deficiencies can reduce sperm motility and contribute to severe acrosomal hypoplasia in spermatozoa, compromising their normal function.

Bi-allelic variants in chromatoid body protein TDRD6 cause spermiogenesis defects and severe oligoasthenoteratozoospermia in humans.

BACKGROUND: The association between the TDRD6 variants and human infertility remains unclear, as only one homozygous missense variant of TDRD6 was found to be associated with oligoasthenoteratozoospermia (OAT). METHODS: Whole-exome sequencing and Sanger sequencing were employed to identify potential pathogenic variants of TDRD6 in infertile men. Histology, immunofluorescence, immunoblotting and ultrastructural analyses were conducted to clarify the structural and functional abnormalities of sperm in mutated patients. Tdrd6-knockout mice were generated using the CRISPR-Cas9 system. Total RNA-seq and single-cell RNA-seq (scRNA-seq) analyses were used to elucidate the underlying molecular mechanisms, followed by validation through quantitative RT-PCR and immunostaining. Intracytoplasmic sperm injection (ICSI) was also used to assess the efficacy of clinical treatment. RESULTS: Bi-allelic TDRD6 variants were identified in five unrelated Chinese individuals with OAT, including homozygous loss-of-function variants in two consanguineous families. Notably, besides reduced concentrations and impaired motility, a significant occurrence of acrosomal hypoplasia was detected in multiple spermatozoa among five patients. Using the Tdrd6-deficient mice, we further elucidate the pivotal role of TDRD6 in spermiogenesis and acrosome identified. In addition, the mislocalisation of crucial chromatoid body components DDX4 (MVH) and UPF1 was also observed in round spermatids from patients harbouring TDRD6 variants. ScRNA-seq analysis of germ cells from a patient with TDRD6 variants revealed that TDRD6 regulates mRNA metabolism processes involved in spermatid differentiation and cytoplasmic translation. CONCLUSION: Our findings strongly suggest that TDRD6 plays a conserved role in spermiogenesis and confirms the causal relationship between TDRD6 variants and human OAT. Additionally, this study highlights the unfavourable ICSI outcomes in individuals with bi-allelic TDRD6 variants, providing insights for potential clinical treatment strategies.

Bi-allelic mutations of DNAH10 cause primary male infertility with asthenoteratozoospermia in humans and mice.

Multiple morphological abnormalities of the sperm flagella (MMAF)-induced asthenoteratozoospermia is a common cause of male infertility. Previous studies have identified several MMAF-associated genes, highlighting the condition's genetic heterogeneity. To further define the genetic causes underlying MMAF, we performed whole-exome sequencing in a cohort of 643 Chinese MMAF-affected men. Bi-allelic DNAH10 variants were identified in five individuals with MMAF from four unrelated families. These variants were either rare or absent in public population genome databases and were predicted to be deleterious by multiple bioinformatics tools. Morphological and ultrastructural analyses of the spermatozoa obtained from men harboring bi-allelic DNAH10 variants revealed striking flagellar defects with the absence of inner dynein arms (IDAs). DNAH10 encodes an axonemal IDA heavy chain component that is predominantly expressed in the testes. Immunostaining analysis indicated that DNAH10 localized to the entire sperm flagellum of control spermatozoa. In contrast, spermatozoa from the men harboring bi-allelic DNAH10 variants exhibited an absence or markedly reduced staining intensity of DNAH10 and other IDA components, including DNAH2 and DNAH6. Furthermore, the phenotypes were recapitulated in mouse models lacking Dnah10 or expressing a disease-associated variant, confirming the involvement of DNAH10 in human MMAF. Altogether, our findings in humans and mice demonstrate that DNAH10 is essential for sperm flagellar assembly and that deleterious bi-allelic DNAH10 variants can cause male infertility with MMAF. These findings will provide guidance for genetic counseling and insights into the diagnosis of MMAF-associated asthenoteratozoospermia.

Deleterious variants in X-linked CFAP47 induce asthenoteratozoospermia and primary male infertility.

Asthenoteratozoospermia characterized by multiple morphological abnormalities of the flagella (MMAF) has been identified as a sub-type of male infertility. Recent progress has identified several MMAF-associated genes with an autosomal recessive inheritance in human affected individuals, but the etiology in approximately 40% of affected individuals remains unknown. Here, we conducted whole-exome sequencing (WES) and identified hemizygous missense variants in the X-linked CFAP47 in three unrelated Chinese individuals with MMAF. These three CFAP47 variants were absent in human control population genome databases and were predicted to be deleterious by multiple bioinformatic tools. CFAP47 encodes a cilia- and flagella-associated protein that is highly expressed in testis. Immunoblotting and immunofluorescence assays revealed obviously reduced levels of CFAP47 in spermatozoa from all three men harboring deleterious missense variants of CFAP47. Furthermore, WES data from an additional cohort of severe asthenoteratozoospermic men originating from Australia permitted the identification of a hemizygous Xp21.1 deletion removing the entire CFAP47 gene. All men harboring hemizygous CFAP47 variants displayed typical MMAF phenotypes. We also generated a Cfap47-mutated mouse model, the adult males of which were sterile and presented with reduced sperm motility and abnormal flagellar morphology and movement. However, fertility could be rescued by the use of intra-cytoplasmic sperm injections (ICSIs). Altogether, our experimental observations in humans and mice demonstrate that hemizygous mutations in CFAP47 can induce X-linked MMAF and asthenoteratozoospermia, for which good ICSI prognosis is suggested. These findings will provide important guidance for genetic counseling and assisted reproduction treatments.

Novel MEIOB pathogenic variants including a homozygous non-canonical splicing variant, cause meiotic arrest and human non-obstructive azoospermia.

Non-obstructive azoospermia (NOA) is the most severe form of human male infertility, and the genetic causes of NOA with meiotic arrest remain largely unclear. In this study, we identified novel compound heterozygous MEIOB variants (c.814C > T: p.R272X and c.976G > A: p.A326T) and a previously undescribed homozygous non-canonical splicing variant of MEIOB (c.528 + 3A > C) in two NOA-affected individuals from two irrelevant Chinese families. MEIOB missense variant (p.A326T) significantly reduced protein abundance and nonsense variant (p.R272X) produced a truncated protein. Both of two variants impaired the MEIOB-SPATA22 interaction. The MEIOB non-canonical splicing variant resulted in whole Exon 6 skipping by minigene assay, which was predicted to produce a frameshift truncated protein (p.S111Rfs*32). Histological and immunostaining analysis indicated that both patients exhibited a similar phenotype as we previously reported in Meiob mutant mice, that is, absence of spermatids in seminiferous tubules and meiotic arrest. Our study identified three novel pathogenic variants of MEIOB in NOA patients, extending the mutation spectrum of the MEIOB and highlighting the contribution of meiotic recombination related genes in human fertility.

TARGET OF MONOPTEROS: key transcription factors orchestrating plant development and environmental response.

Plants have an incredible ability to sustain root and vascular growth after initiation of the embryonic root and the specification of vascular tissue in early embryos. Microarray assays have revealed that a group of transcription factors, TARGET OF MONOPTEROS (TMO), are important for embryonic root initiation in Arabidopsis. Despite the discovery of their auxin responsiveness early on, their function and mode of action remained unknown for many years. The advent of genome editing has accelerated the study of TMO transcription factors, revealing novel functions for biological processes such as vascular development, root system architecture, and response to environmental cues. This review covers recent achievements in understanding the developmental function and the genetic mode of action of TMO transcription factors in Arabidopsis and other plant species. We highlight the transcriptional and post-transcriptional regulation of TMO transcription factors in relation to their function, mainly in Arabidopsis. Finally, we provide suggestions for further research and potential applications in plant genetic engineering.

High sensitivity distributed dynamic pressure sensor based on dual-linear frequency modulated optical frequency domain reflectometry.

In this Letter, we propose and experimentally demonstrate a highly sensitive distributed dynamic pressure sensor based on a dual-linear frequency modulated optical frequency domain reflectometry (OFDR) and a coating thickness-enhanced single-mode fiber (SMF). A dual-sideband linear frequency modulation (LFM) signal is used to interrogate the sensing fiber, which allows us to obtain a dual-sideband Rayleigh backscattering signal. Due to the opposite slopes of the two LFM sidebands, the Rayleigh backscattering spectra of the two sidebands drift in opposite directions when the fiber is disturbed. By subtracting the frequency shifts of the two spectra, we can double the system's sensitivity. We further enhance the sensitivity by using an SMF with a coating thickness of 200 µm. This results in a pressure sensitivity of 3979 MHz/MPa, a measurement accuracy of 0.76 kPa, and a spatial resolution of 35 cm over a 500 m optical fiber. Our system successfully detected a dynamic pressure change at a sampling rate of 1.25 kHz, demonstrating the sensor's excellent dynamic measuring capabilities.

Achieving High Proton Conductivity in MIL-91(Al) Aerogel.

The processability and sustainability of proton conductors are two important indicators of their application. Here, MIL-91(Al) with an intrinsic proton conduction framework originating from protonated phosphonate groups was cross-linked with poly(vinyl alcohol) (PVA) to obtain MIL-91(Al) aerogel through freeze-drying. This simple and inexpensive strategy not only facilitated the processing of MIL-91(Al) powder but also resulted in a molded MIL-91(Al) aerogel having a high proton conductivity of 1.02 × 10-2 S cm-1 at 70 °C and 100% relative humidity. Furthermore, MIL-91(Al) aerogel was recyclable and reusable, in line with the principles of environmental protection and sustainability. To the best of our knowledge, this is the first example of using a metal-organic framework aerogel as a proton conductor, which may develop a new model system in this field.

Dyslipidemia and hyperuricemia: a cross-sectional study of residents in Wuhu, China.

BACKGROUND: While dyslipidemia has been recognized as a potential risk factor for hyperuricemia, there is currently a dearth of large-scale data specifically focused on studying the relationship between these two conditions. To address this gap, the present study analyzed a dataset of 298,891 physical examination records to investigate in greater detail the clinical classification and compositional relationship between hyperuricemia and dyslipidemia. METHODS: For this investigation, a cross-sectional research design was utilized to analyze physical examination data that was gathered from Yijishan Hospital in Wuhu, China between 2011 and 2016. Logistic regression was employed to examine the association between hyperuricemia and dyslipidemia. Furthermore, the association between hyperuricemia and dyslipidemia was evaluated based on the clinical classifications of dyslipidemia and its components. RESULTS: A total of 298,891 participants from China (124,886 [41.8%] females) were included in the study, with an age range of 18 to 90 years (mean [SD]: 47.76 [13.54] years). In multivariate analysis, the odds of hyperuricemia was 1.878 times higher in patients with dyslipidemia compared to those without dyslipidemia (95% confidence interval [CI]: 1.835-1.922). In the clinical classification of dyslipidemia, individuals with hypertriglyceridemia and mixed hyperlipidemia had 1.753 times (95% CI: 1.706-1.802) and 1.925 times (95% CI: 1.870-1.982) higher odds of hyperuricemia, respectively, compared to those without dyslipidemia. Among the components of dyslipidemia, the odds ratios for hyperuricemia in individuals in the fourth quartile compared to those in the first quartile were 3.744 (95% CI: 3.636-3.918) for triglycerides, 1.518 (95% CI: 1.471-1.565) for total cholesterol, and 1.775 (95% CI: 1.718 - 1.833) for non-high-density lipoprotein cholesterol. CONCLUSIONS: Dyslipidemia has been independently linked with hyperuricemia. Moreover, the elevation of triglycerides or total cholesterol levels, including conditions such as hypertriglyceridemia and mixed hyperlipidemia, have been observed to have a positive association with the development of hyperuricemia.

Homozygous variants in AKAP3 induce asthenoteratozoospermia and male infertility.

BACKGROUND: As a common type of asthenoteratozoospermia, multiple morphological abnormalities of the sperm flagella (MMAF) can cause male infertility. Previous studies have revealed genetic factors as a major cause of MMAF. The known MMAF-associated genes are involved in the mitochondrial sheath, outer dense fibre or axoneme of the sperm flagella. These findings indicate the genetic heterogeneity of MMAF. METHODS AND RESULTS: Here, we conducted genetic analyses using whole-exome sequencing in a cohort of 150 Han Chinese men with asthenoteratozoospermia. Homozygous deleterious variants of AKAP3 (A-kinase anchoring protein 3) were identified in two MMAF-affected men from unrelated families. One AKAP3 variant was a frameshift (c.2286_2287del, p.His762Glnfs*22) and the other variant was a missense mutation (c.44G>A, p.Cys15Tyr), which was predicted to be damaging by multiple bioinformatics tools. Further western blotting and immunofluorescence assays revealed the absence of AKAP3 in the spermatozoa from the man harbouring the homozygous frameshift variant, whereas the expression of AKAP3 was markedly reduced in the spermatozoa of the man with the AKAP3 missense variant p.Cys15Tyr. Notably, the clinical outcomes after intracytoplasmic sperm injection (ICSI) were divergent between these two cases, suggesting a possibility of AKAP3 dosage-dependent prognosis of ICSI treatment. CONCLUSIONS: Our study revealed AKAP3 as a novel gene involved in human asthenoteratozoospermia.

Biallelic mutations in CFAP54 cause male infertility with severe MMAF and NOA.

BACKGROUND: Spermatogenic impairments can lead to male infertility by different pathological conditions, such as multiple morphological abnormalities of the sperm flagella (MMAF) and non-obstructive azoospermia (NOA). Genetic factors are involved in impaired spermatogenesis. METHODS AND RESULTS: Here, we performed genetic analyses through whole-exome sequencing in a cohort of 334 Han Chinese probands with severe MMAF or NOA. Biallelic variants of CFAP54 were identified in three unrelated men, including one homozygous frameshift variant (c.3317del, p.Phe1106Serfs*19) and two compound heterozygous variants (c.878G>A, p.Arg293His; c.955C>T, p.Arg319Cys and c.4885C>T, p.Arg1629Cys; c.937G>A, p.Gly313Arg). All of the identified variants were absent or extremely rare in the public human genome databases and predicted to be damaging by bioinformatic tools. The men harbouring CFAP54 mutations exhibited abnormal sperm morphology, reduced sperm concentration and motility in ejaculated semen. Significant axoneme disorganisation and other ultrastructure abnormities were also detected inside the sperm cells from men harbouring CFAP54 mutations. Furthermore, immunofluorescence assays showed remarkably reduced staining of four flagellar assembly-associated proteins (IFT20, IFT52, IFT122 and SPEF2) in the spermatozoa of CFAP54-deficient men. Notably, favourable clinical pregnancy outcomes were achieved with sperm from men carrying CFAP54 mutations after intracytoplasmic sperm injection treatment. CONCLUSION: Our genetic analyses and experimental observations revealed that biallelic deleterious mutations of CFAP54 can induce severe MMAF and NOA in humans.

Dendrobium officinale Polysaccharides as a Natural Functional Component for Acetic-Acid-Induced Gastric Ulcers in Rats.

Dendrobium officinale is an important edible and medicinal plant, with the Dendrobium officinale polysaccharide (DOP) being its primary active constituent, known for its diverse biological activities. In this study, DOP was extracted and characterized for its structural properties. The potential of DOP to ameliorate gastric ulcers (GUs) was investigated using an acetic-acid-induced GU model in rats. The results demonstrated that DOP exerted a multifaceted protective effect against GU, mitigating the deleterious impact on food intake and body weight in rats. DOP exhibited its protective action by attenuating cellular damage attributed to oxidative stress and inflammatory reactions mediated by enhanced activities of SOD, GSH, and GSH-PX, coupled with a downregulation in the expression of pro-inflammatory cytokines, including IL-1ß, IL-6, and TNF-α. Furthermore, DOP effectively inhibited apoptosis in gastric mucosa cells of acetic-acid-induced GU rat models and facilitated the self-repair of damaged tissues. Remarkably, the DOP-200 and DOP-400 groups outperformed omeprazole in reducing the expression of IL-6 and malondialdehyde (MDA) in tissues, as well as IL-1ß, IL-6, and TNF-α in serum. These groups also exhibited an improved expression of SOD in tissues and SOD, GSH, and GSH-PX in serum. A Western blot analysis of gastric mucosa demonstrated that the DOP-200 and DOP-400 groups significantly reduced the expression of NF-κBp65, phosphorylated NF-κBp65, FoxO3a, and Bim. The observed antagonism to GU appeared to be associated with the NF-κB cell pathway. Additionally, qRT-PCR results indicate that DOP reduced the mRNA transcription levels of IL-6, and TNF-α, which shows that the healing of GU is related to the reduction in the inflammatory reaction by DOP. However, the expression of EGF and VEGF decreased, suggesting that the mechanism of DOP inhibiting GU may not be directly related to EGF and VEGF, or there is an uncertain competitive relationship between them, so further research is needed.

The mutation rate of rpoB gene showed an upward trend with the increase of MIRU10, MIRU39 and QUB4156 repetitive number.

BACKGROUND: Mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) is a frequently used typing method for identifying the Beijing genotype of Mycobacterium tuberculosis (Mtb), which is easily transformed into rifampicin (RIF) resistance. The RIF resistance of Mtb is considered to be highly related with the mutation of rpoB gene. Therefore, this study aimed to analyze the relationship between the repetitive number of MIRU loci and the mutation of rpoB gene. METHODS: An open-source whole-genome sequencing data of Mtb was used to detect the mutation of rpoB gene and the repetitive number of MIRU loci by bioinformatics methods. Cochran-Armitage analysis was performed to analyze the trend of the rpoB gene mutation rate and the repetitive number of MIRU loci. RESULTS: Among 357 rifampicin-resistant tuberculosis (RR-TB), 304 strains with mutated rpoB genes were detected, and 6 of 67 rifampicin susceptible strains were detected mutations. The rpoB gene mutational rate showed an upward trend with the increase of MIRU10, MIRU39, QUB4156 and MIRU16 repetitive number, but only the repetitive number of MIRU10, MRIU39 and QUB4156 were risk factors for rpoB gene mutation. The Hunter-Gaston discriminatory index (HGDI) of MIRU10 (0.65) and QUB4156 (0.62) was high in the overall sample, while MIRU39 (0.39) and MIRU16 (0.43) showed a moderate discriminatory Power. CONCLUSION: The mutation rate of rpoB gene increases with the addition of repetitive numbers of MIRU10, QUB4156 and MIRU39 loci.

Characterization of Highbush Blueberry (Vaccinium corymbosum L.) Anthocyanin Biosynthesis Related MYBs and Functional Analysis of VcMYB Gene.

As one of the most important transcription factors regulating plant anthocyanin biosynthesis, MYB has attracted great attentions. In this study, we identified fifteen candidate anthocyanin biosynthesis related MYB (ABRM) proteins, including twelve R2R3-MYBs and three 1R-MYBs, from highbush blueberry. The subcellular localization prediction results showed that, with the exception of VcRVE8 (localized in chloroplast and nucleus), all of the blueberry ABRMs were nucleus-localized. The gene structure analysis revealed that the exon numbers of the blueberry ABRM genes varied greatly, ranging between one and eight. There are many light-responsive, phytohormone-responsive, abiotic stress-responsive and plant growth and development related cis-acting elements in the promoters of the blueberry ABRM genes. It is noteworthy that almost all of their promoters contain light-, ABA- and MeJA-responsive elements, which is consistent with the well-established results that anthocyanin accumulation and the expression of MYBs are influenced significantly by many factors, such as light, ABA and JA. The gene expression analysis revealed that VcMYB, VcMYB6, VcMYB23, VcMYBL2 and VcPH4 are expressed abundantly in blueberry fruits, and VcMYB is expressed the highest in the red, purple and blue fruits among all blueberry ABRMs. VcMYB shared high similarity with functionally proven ABRMs from many other plant species. The gene cloning results showed that VcMYB had three variable transcripts, but only the transient overexpression of VcMYB-1 promoted anthocyanin accumulation in the green fruits. Our study can provide a basis for future research on the anthocyanin biosynthesis related MYBs in blueberry.

Loss of DRC1 function leads to multiple morphological abnormalities of the sperm flagella and male infertility in human and mouse.

Motile cilia and flagellar defects can result in primary ciliary dyskinesia, which is a multisystemic genetic disorder that affects roughly 1:10 000 individuals. The nexin-dynein regulatory complex (N-DRC) links neighboring doublet microtubules within flagella, serving as a central regulatory hub for motility in Chlamydomonas. Herein, we identified two homozygous DRC1 variants in human patients that were associated with multiple morphological abnormalities of the sperm flagella (MMAF) and male infertility. Drc1-/-, Drc1R554X/R554X and Drc1W244X/W244X mice on the C57BL/6 background suffered from pre-pubertal mortality. However, when the ICR background was introduced, some of these mice were able to survive and recapitulate the MMAF phenotypes detected in human patients. By analyzing these animals, we determined that DRC1 is an essential regulator of N-DRC assembly in cilia and flagella. When DRC1 is absent, this results in the shortening of cilia and consequent impairment of their motility. Damage associated with DRC1 deficiency in sperm flagella was more pronounced than in cilia, as manifested by complete axoneme structural disorder in addition to the loss of the DRC structure. Altogether, these findings suggest that DRC1 is required for the structural stability of flagella but not cilia, emphasizing the key role of this protein in mammalian species.

Bi-allelic DNAH8 Variants Lead to Multiple Morphological Abnormalities of the Sperm Flagella and Primary Male Infertility.

Sperm malformation is a direct factor for male infertility. Multiple morphological abnormalities of the flagella (MMAF), a severe form of asthenoteratozoospermia, are characterized by immotile spermatozoa with malformed and/or absent flagella in the ejaculate. Previous studies indicated genetic heterogeneity in MMAF. To further define genetic factors underlying MMAF, we performed whole-exome sequencing in a cohort of 90 Chinese MMAF-affected men. Two cases (2.2%) were identified as carrying bi-allelic missense DNAH8 variants, variants which were either absent or rare in the control human population and were predicted to be deleterious by multiple bioinformatic tools. Re-analysis of exome data from a second cohort of 167 MMAF-affected men from France, Iran, and North Africa permitted the identification of an additional male carrying a DNAH8 homozygous frameshift variant. DNAH8 encodes a dynein axonemal heavy-chain component that is expressed preferentially in the testis. Hematoxylin-eosin staining and electron microscopy analyses of the spermatozoa from men harboring bi-allelic DNAH8 variants showed a highly aberrant morphology and ultrastructure of the sperm flagella. Immunofluorescence assays performed on the spermatozoa from men harboring bi-allelic DNAH8 variants revealed the absent or markedly reduced staining of DNAH8 and its associated protein DNAH17. Dnah8-knockout male mice also presented typical MMAF phenotypes and sterility. Interestingly, intracytoplasmic sperm injections using the spermatozoa from Dnah8-knockout male mice resulted in good pregnancy outcomes. Collectively, our experimental observations from humans and mice demonstrate that DNAH8 is essential for sperm flagellar formation and that bi-allelic deleterious DNAH8 variants lead to male infertility with MMAF.

Bi-allelic Loss-of-function Variants in CFAP58 Cause Flagellar Axoneme and Mitochondrial Sheath Defects and Asthenoteratozoospermia in Humans and Mice.

Multiple morphological abnormalities of the sperm flagella (MMAF) is a severe form of asthenoteratozoospermia. Although recent studies have revealed several MMAF-associated genes and demonstrated MMAF to be a genetically heterogeneous disease, at least one-third of the cases are still not well understood for their etiology. Here, we identified bi-allelic loss-of-function variants in CFAP58 by using whole-exome sequencing in five (5.6%) unrelated individuals from a cohort of 90 MMAF-affected Chinese men. Each of the men harboring bi-allelic CFAP58 variants presented typical MMAF phenotypes. Transmission electron microscopy demonstrated striking flagellar defects with axonemal and mitochondrial sheath malformations. CFAP58 is predominantly expressed in the testis and encodes a cilia- and flagella-associated protein. Immunofluorescence assays showed that CFAP58 localized at the entire flagella of control sperm and predominantly concentrated in the mid-piece. Immunoblotting and immunofluorescence assays showed that the abundances of axoneme ultrastructure markers SPAG6 and SPEF2 and a mitochondrial sheath protein, HSP60, were significantly reduced in the spermatozoa from men harboring bi-allelic CFAP58 variants. We generated Cfap58-knockout mice via CRISPR/Cas9 technology. The male mice were infertile and presented with severe flagellar defects, consistent with the sperm phenotypes in MMAF-affected men. Overall, our findings in humans and mice strongly suggest that CFAP58 plays a vital role in sperm flagellogenesis and demonstrate that bi-allelic loss-of-function variants in CFAP58 can cause axoneme and peri-axoneme malformations leading to male infertility. This study provides crucial insights for understanding and counseling of MMAF-associated asthenoteratozoospermia.

Prevalence and Associations of Beta2-Microglobulin Mutations in MSI-H/dMMR Cancers.

Microsatellite instability (MSI) has emerged as an important predictor of sensitivity for immunotherapy-based strategies. ß-2-Microglobulin (B2M) contains microsatellites within the coding regions and is prone to somatic changes in MSI/mismatch repair deficiency (MSI/dMMR) tumors. To delineate prevalence and associations of B2M mutations in MSI-H/dMMR cancers, we investigated the mutational profile of B2M and clinical and pathological features in gastric cancer (GC), colorectal cancer (CRC), and endometrial cancer (EC) with a high incidence of microsatellite instability-high (MSI-H)/dMMR. Formalin-fixed paraffin-embedded (FFPE) tumor tissues along with matched normal tissues were collected from 108 MSI/dMMR patients with GC, CRC, and EC. Genomic profiling of tissue and blood samples were assessed next-generation sequencing (NGS). Immunohistochemistry (IHC) was used to examine the presence or absence of B2M protein. Alternations in the exonic microsatellite regions of B2M were observed at various but high frequencies (57.5% in CRC, 23.9% in GC, and 13.6% in EC) and in different forms. NGS assay revealed that genes involved in chromatin regulation, the PI3K pathway, the WNT pathway, and mismatch repair were extensively altered in the MSI-H cohort. Signature 6 and 26, 2 of 4 mutational signatures associated with defective DNA mismatch repair, featured with high numbers of small insertion/deletions (INDEL) dominated in all 3 types of cancer. Alternations in the exonic microsatellite regions of B2M were observed at various but high frequencies (57.5% in CRC, 23.9% in GC, and 13.6% in EC) and in different forms. Tumor mutational burden (TMB) was significantly higher in the patients carrying MSI-H/dMMR tumors with B2M mutation than that in patients with wild-type B2M (P = .026).The frame shift alteration occurring at the exonic microsatellite sties caused loss of function of B2M gene. In addition, a case with CRC carrying indels in B2M gene resisted the ICI treatment was reported. In conclusion, patients carrying MSI-H/dMMR tumors with B2M mutation showed significantly higher TMB. Prescription of ICIs should be thoroughly evaluated for these patients.