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Sperm with normal morphology and motility are essential for successful fertilization, and the strong attachment of the sperm head-tail coupling apparatus to the nuclear envelope during spermatogenesis is required to ensure the integrity of sperm for capacitation and fertilization. Here, we report that Arrdc5 is associated with spermatogenesis. The Arrdc5 knockout mouse model showed male infertility characterized by a high bent-head rate and reduced motility in sperm, which led to capacitation defects and subsequent fertilization failure. Through mass spectrometry, we found that ARRDC5 affects spermatogenesis by affecting NDC1 and SUN5. We further found that ARRDC5 might affect the vesicle-trafficking protein SEC22A-mediated transport and localization of NDC1, SUN5 and other head-tail coupling apparatus-related proteins that are responsible for initiating the attachment of the sperm head and tail. We finally performed intracytoplasmic sperm injection as a way to explore therapeutic strategies. Our findings demonstrate the essential role and the underlying molecular mechanism of ARRDC5 in anchoring the sperm head to the tail during spermatogenesis.
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Infertilidade Masculina , Sêmen , Humanos , Animais , Camundongos , Masculino , Sêmen/metabolismo , Espermatogênese/genética , Espermatozoides/metabolismo , Cabeça do Espermatozoide/metabolismo , Proteínas/metabolismo , Infertilidade Masculina/genética , Infertilidade Masculina/metabolismo , Camundongos Knockout , Proteínas de Membrana/metabolismoRESUMO
The accumulation and storage of maternal mRNA is crucial for oocyte maturation and embryonic development. PATL2 is an oocyte-specific RNA-binding protein, and previous studies have confirmed that PATL2 mutation in humans and knockout mice cause oocyte maturation arrest or embryonic development arrest, respectively. However, the physiological function of PATL2 in the process of oocyte maturation and embryonic development is largely unknown. Here, we report that PATL2 is highly expressed in growing oocytes and couples with EIF4E and CPEB1 to regulate maternal mRNA expression in immature oocytes. The germinal vesicle oocytes from Patl2-/- mice exhibit decreasing maternal mRNA expression and reduced levels of protein synthesis. We further confirmed that PATL2 phosphorylation occurs in the oocyte maturation process and identified the S279 phosphorylation site using phosphoproteomics. We found that the S279D mutation decreased the protein level of PATL2 and led to subfertility in Palt2S279D knock-in mice. Our work reveals the previously unrecognized role of PATL2 in regulating the maternal transcriptome and shows that phosphorylation of PATL2 leads to the regulation of PATL2 protein levels via ubiquitin-mediated proteasomal degradation in oocytes.
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Fator de Iniciação 4E em Eucariotos , Proteínas Nucleares , RNA Mensageiro Estocado , Proteínas de Ligação a RNA , Animais , Feminino , Humanos , Camundongos , Gravidez , Fator de Iniciação 4E em Eucariotos/metabolismo , Homeostase , Camundongos Knockout , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo , Proteínas Nucleares/metabolismo , Oócitos/metabolismo , RNA Mensageiro/metabolismo , RNA Mensageiro Estocado/metabolismo , Proteínas de Ligação a RNA/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Fertilization is a fundamental process of development, and the blocking mechanisms act at the zona pellucida (ZP) and plasma membrane of the egg to prevent any additional sperm from binding, permeating and fusing after fertilization. In clinical practice, some couples undergoing recurrent IVF failures that mature oocytes had abnormal fertilization for unknown reason. Ovastacin encoded by ASTL cleave the ZP protein ZP2 and play a key role in preventing polyspermy. Here, we identified bi-allelic variants in ASTL that are mainly characterized by fertilization problems in humans. All four independent affected individuals had bi-allelic frameshift variants or predicted damaging missense variants, which follow a Mendelian recessive inheritance pattern. The frameshift variants significantly decreased the quantity of ASTL protein in vitro. And all missense variants affected the enzymatic activity that cleaves ZP2 in mouse egg in vitro. Three knock-in female mice (corresponding to three missense variants in patients) all show subfertility due to low embryo developmental potential. This work presents strong evidence that pathogenic variants in ASTL cause female infertility and provides a new genetic marker for the diagnosis of fertilization problems.
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Infertilidade Feminina , Sêmen , Humanos , Masculino , Feminino , Camundongos , Animais , Glicoproteínas da Zona Pelúcida/genética , Glicoproteínas da Zona Pelúcida/metabolismo , Sêmen/metabolismo , Oócitos/metabolismo , Infertilidade Feminina/genética , Fertilização/genética , Metaloproteases/genéticaRESUMO
Capacity-limited visual working memory (VWM) requires that individuals have sufficient memory space and the ability to filter distractors. Negative emotional states are known to impact VWM storage, yet their influence on distractor filtering within VWM remains underexplored. We conducted direct neural measurement of participants (n = 56) who conducted a lateralized change detection task with distractors, while manipulating the emotional state by presenting neutral or negative images before each trial. We found a detrimental effect of distractors on memory accuracy under both neutral and negative emotional states. Using the event-related potential (ERP) component, contralateral delay activity (CDA; sensitive to VWM load), to observe the VWM load in each condition, we found that in the neutral state, the participants showed significantly higher late CDA amplitudes when remembering 4 targets compared with 2 targets and 2 targets with 2 distractors but no significant difference when remembering 2 targets compared with 2 targets with 2 distractors. In the negative state, no significant CDA amplitude differences were evident when remembering 4 targets and 2 targets, but CDA was significantly higher when remembering 2 targets with 2 distractors compared with 2 targets. These results suggest that the maximum number of items participants could store in VWM was lower under negative emotional states than under neutral emotional states. Importantly, the participants could filter out distractors when in a neutral emotional state but not in a negative emotional state, indicating that negative emotional states impair their ability to filter out distractors in VWM.
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Eletroencefalografia , Emoções , Potenciais Evocados , Memória de Curto Prazo , Humanos , Feminino , Masculino , Memória de Curto Prazo/fisiologia , Emoções/fisiologia , Adulto Jovem , Potenciais Evocados/fisiologia , Adulto , Atenção/fisiologia , Encéfalo/fisiologia , AdolescenteRESUMO
Layered indium selenide (InSe) is a new 2D semiconductor material with high carrier mobility, widely adjustable bandgap, and high ductility. However, its ion storage behavior and related electrochemical reaction mechanism are rarely reported. In this study, InSe nanoflakes encapsulated in conductive polypyrrole (InSe@PPy) are designed in consideration of restraining the severe volume change in the electrochemical reaction and increasing conductivity via in situ chemical oxidation polymerization. Density functional theory calculations demonstrate that the construction of heterostructure can generate an internal electric field to accelerate electron transfer via additional driving forces, offering synergistically enhanced structural stability, electrical conductivity, and Na+ diffusion process. The resulting InSe@PPy composite shows outstanding electrochemical performance in the sodium ion batteries system, achieving a high reversible capacity of 336.4 mA h g-1 after 500 cycles at 1 A g-1 and a long-term cyclic stability with capacity of 274.4 mA h g-1 after 2800 cycles at 5 A g-1 . In particular, the investigation of capacity fluctuation within the first cycling reveals the alternating significance of intercalation and conversion reactions and evanescent alloying reaction. The combined reaction mechanism of insertion, conversion, and alloying of InSe@PPy is revealed by in situ X-ray diffraction, ex situ electrochemical impedance spectroscopy, and transmission electron microscopy.
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The two-stage preference design (TSPD) enables inference for treatment efficacy while allowing for incorporation of patient preference to treatment. It can provide unbiased estimates for selection and preference effects, where a selection effect occurs when patients who prefer one treatment respond differently than those who prefer another, and a preference effect is the difference in response caused by an interaction between the patient's preference and the actual treatment they receive. One potential barrier to adopting TSPD in practice, however, is the relatively large sample size required to estimate selection and preference effects with sufficient power. To address this concern, we propose a group sequential two-stage preference design (GS-TSPD), which combines TSPD with sequential monitoring for early stopping. In the GS-TSPD, pre-planned sequential monitoring allows investigators to conduct repeated hypothesis tests on accumulated data prior to full enrollment to assess study eligibility for early trial termination without inflating type I error rates. Thus, the procedure allows investigators to terminate the study when there is sufficient evidence of treatment, selection, or preference effects during an interim analysis, thereby reducing the design resource in expectation. To formalize such a procedure, we verify the independent increments assumption for testing the selection and preference effects and apply group sequential stopping boundaries from the approximate sequential density functions. Simulations are then conducted to investigate the operating characteristics of our proposed GS-TSPD compared to the traditional TSPD. We demonstrate the applicability of the design using a study of Hepatitis C treatment modality.
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Preferência do Paciente , Projetos de Pesquisa , Humanos , Tamanho da Amostra , Resultado do TratamentoRESUMO
Currently, purely organic compounds showing ambient phosphorescence with high efficiency (ΦP ) and ultra-long lifetime (τP ) are quite rare and often need to be achieved in hydrophilic poly(vinyl alcohol)-based hosts. This severely limits their applications. Here, we provide a solution to this issue by constructing an ortho-linked donor-acceptor (D-A) dyad whose D moiety has not only a long-lived T1 state to achieve a long τP , but also a Tn state that is close to the S1 state of the dyad to trigger effective spin-orbit charge transfer intersystem crossing (SOCT-ISC). The rationality of this strategy was validated by a new phosphor OF-BCz that is able to show a τP of 1.92â s and a ΦP of 30 % even in a less rigid matrix of poly(methyl methacrylate) (PMMA). Excitingly, OF-BCz exhibited its potential as both a photocuring initiator and an in situ quality indicator, allowing for the visual detection of defects in photolithographic patterning.
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Oocyte maturation defects are major phenotypes resulting in female infertility. Although many genetic factors have been found to be responsible for these phenotypes, the underlying pathogenic genes and variants remain to be identified. The anaphase promoting complex or cyclosome (APC/C) is known to be essential in the metaphase-to-anaphase transition. In this study, we identified two homozygous missense variants (c.986A > G, p.Y329C and c.988C > T, p.R330C) in CDC23 that are responsible for female infertility characterized by oocyte maturation defects in three infertile individuals. CDC23 (cell division cycle 23) is one of the core subunits of the APC/C. In vitro experiments showed that the variant c.986A > G (p.Y329C) led to a decrease in CDC23 protein level and the variant c.988C > T (p.R330C) changed the localization of CDC23 in HeLa cells and mouse oocytes. In vivo studies showed that Cdc23Y329C/Y329C mice successfully mimicked the patients' phenotype by causing low expression of CDC23 and APC4 and the accumulation of securin and cyclin B1 in oocytes. AZ3146 treatment was able to rescue the phenotype. Taken together, our findings reveal the important roles of CDC23 in human oocyte maturation and provide a new genetic marker for female infertility.
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Proteínas de Ciclo Celular , Infertilidade Feminina , Humanos , Feminino , Animais , Camundongos , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Células HeLa , Infertilidade Feminina/genética , Ciclossomo-Complexo Promotor de Anáfase , OócitosRESUMO
Asthenozoospermia is one of the main factors leading to male infertility, but the genetic mechanisms have not been fully elucidated. Variants in the androglobin (ADGB) gene were identified in an infertile male characterized by asthenozoospermia. The variants disrupted the binding of ADGB to calmodulin. Adgb-/- male mice were infertile due to reduced sperm concentration (< 1 × 106 /mL) and motility. Spermatogenesis was also abnormal, with malformation of both elongating and elongated spermatids, and there was an approximately twofold increase in apoptotic cells in the cauda epididymis. These exacerbated the decline in sperm motility. It is surprising that ICSI with testicular spermatids allows fertilization and eventually develops into blastocyst. Through mass spectrometry, we identified 42 candidate proteins that are involved in sperm assembly, flagella formation, and sperm motility interacting with ADGB. In particular, CFAP69 and SPEF2 were confirmed to bind to ADGB. Collectively, our study suggests the potential important role of ADGB in human fertility, revealing its relevance to spermatogenesis and infertility. This expands our knowledge of the genetic causes of asthenozoospermia and provides a theoretical basis for using ADGB as an underlying genetic marker for infertile males.
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Astenozoospermia , Infertilidade Masculina , Animais , Humanos , Masculino , Camundongos , Astenozoospermia/genética , Infertilidade Masculina/genética , Infertilidade Masculina/metabolismo , Sêmen/metabolismo , Motilidade dos Espermatozoides/genética , Espermatozoides/metabolismoRESUMO
Normal oocyte maturation is an important requirement for the success of human reproduction, and defects in this process will lead to female infertility and repeated IVF/ICSI failures. In order to identify genetic factors that are responsible for oocyte maturation defect, we used whole exome sequencing in the affected individual with oocyte maturation defect from a consanguineous family and identified a homozygous variant c.853_861del (p.285_287del) in ZFP36L2. ZFP36L2 is a RNA-binding protein, which regulates maternal mRNA decay and oocyte maturation. In vitro studies showed that the variant caused decreased protein levels of ZFP36L2 in oocytes due to mRNA instability and might lead to the loss of its function to degrade maternal mRNAs. Previous study showed that the pathogenic variants in ZFP36L2 were associated with early embryonic arrest. In contrast, we identified a novel ZFP36L2 variant in the affected individual with oocyte maturation defect, which further broadened the mutational and phenotypic spectrum of ZFP36L2, suggesting that ZFP36L2 might be a genetic diagnostic marker for the affected individuals with oocyte maturation defect.
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Infertilidade Feminina , Feminino , Humanos , Infertilidade Feminina/genética , Infertilidade Feminina/patologia , Oócitos/metabolismo , Oogênese/genética , Mutação , Homozigoto , Fatores de Transcrição/genéticaRESUMO
Ovarian dysfunction, including premature ovarian insufficiency and decreased ovarian reserve, affects the ovarian reserve and is one of the leading causes of female infertility. More and more cases of ovarian dysfunction are associated with genetic factors. Here, we identified eight potential variants in five genes (MSH4, HFM1, SYCE1, FSHR, and C14orf39) from six independent families by exome sequencing. The splice-site variants in SYCE1 and MSH4 affected canonical splicing isoforms, leading to missing protein domains or premature termination. Our findings expand the mutational spectrum of ovarian dysfunction and provide potential biomarkers for future genetic counseling and for more personalized treatments. Exome sequencing was shown to be a useful tool to better dissect the genetic basis for ovarian dysfunction and yielded a genetic diagnosis in about 5.0% (6/124) of cases in a cohort of 124 patients with ovarian dysfunction.
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Menopausa Precoce , Insuficiência Ovariana Primária , Humanos , Feminino , Insuficiência Ovariana Primária/diagnóstico , Insuficiência Ovariana Primária/genética , Menopausa Precoce/genética , Mutação , Testes GenéticosRESUMO
STUDY QUESTION: Can new genetic factors responsible for male infertility be identified, especially for those characterized by asthenospermia despite normal sperm morphology? SUMMARY ANSWER: We identified the novel pathogenetic gene IQ motif and ubiquitin-like domain-containing (IQUB) as responsible for male infertility characterized by asthenospermia, involving sperm radial spoke defects. WHAT IS KNOWN ALREADY: To date, only a few genes have been found to be responsible for asthenospermia with normal sperm morphology. Iqub, encoding the IQUB protein, is highly and specifically expressed in murine testes and interacts with the proteins radial spoke head 3 (RSPH3), CEP295 N-terminal like (CEP295NL or DDC8), glutathione S-transferase mu 1 (GSTM1) and outer dense fiber of sperm tails 1 (ODF1) in the yeast two-hybrid system. STUDY DESIGN, SIZE, DURATION: The IQUB variant was identified by whole-exome sequencing in a cohort of 126 male infertility patients with typical asthenospermia recruited between 2015 and 2020. Knockout (KO) and knockin (KI) mouse models, scanning and transmission electron microscopy (TEM), and other functional assays were performed, between 2019 and 2021. PARTICIPANTS/MATERIALS, SETTING, METHODS: The IQUB variant was identified by whole-exome sequencing and confirmed by Sanger sequencing. Iqub KO and KI mice were constructed to mimic the phenotype of the affected individual. After recapitulating the phenotype of human male infertility, scanning and TEM were performed to check the ultrastructure of the sperm. Western blot and co-immunoprecipitation were performed to clarify the pathological mechanism of the IQUB variant. MAIN RESULTS AND THE ROLE OF CHANCE: We identified a homozygous nonsense IQUB variant (NM_001282855.2:c.942T> G(p.Tyr314*)) from an infertile male. Iqub KO and KI mice mimicked the infertility phenotype and confirmed IQUB to be the pathogenetic gene. Scanning and TEM showed that sperm of both the mouse models and the affected individual had radial spoke defects. The functional assay suggested that IQUB may recruit calmodulin in lower Ca2+ environments to facilitate the normal assembly of radial spokes by inhibiting the activity of RSPH3/p-ERK1/2 (a nontypical AKAP (A-Kinase Anchoring Protein) forming by RSPH3 and phosphorylation of extracellular signal-regulated kinase 1 and 2 (p-ERK1/2)). LIMITATIONS, REASONS FOR CAUTION: Additional cases are needed to confirm the genetic contribution of IQUB variants to male infertility. In addition, because no IQUB antibody is available for immunofluorescence and the polyclonal antibody we generated was only effective in western blotting, immunostaining for IQUB was not performed in this study. Therefore, this study lacks direct in vivo proof to confirm the effect of the variant on IQUB protein level. WIDER IMPLICATIONS OF THE FINDINGS: Our results suggest a causal relation between IQUB variants and male infertility owing to asthenospermia, and partly clarify the pathological mechanism of IQUB variants. This expands our knowledge of the genes involved in human sperm asthenospermia and potentially provides a new genetic marker for male infertility. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research and Development Program of China (2021YFC2700100), the National Natural Science Foundation of China (32130029, 82171643, 81971450, 82001538, and 81971382) and the Guangdong Science and Technology Department Guangdong-Hong Kong-Macao Joint Innovation Project (2020A0505140003). There are no competing interests to declare. TRIAL REGISTRATION NUMBER: N/A.
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Astenozoospermia , Infertilidade Masculina , Humanos , Masculino , Animais , Camundongos , Fosforilação , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases , Sêmen/metabolismo , Camundongos Knockout , Infertilidade Masculina/patologia , Espermatozoides/metabolismo , Astenozoospermia/metabolismoRESUMO
Matcha, a powder processed from tea leaves, has a unique green tea flavor and appealing color, in addition to many other sought after functional properties for a wide range of formulated food applications (e.g., dairy products, bakery products, and beverage). The properties of matcha are influenced by cultivation method and processing post-harvest. The transition from drinking tea infusion to eating whole leaves provides a healthy option for the delivery of functional component and tea phenolics in various food matrix. The aim of this review is to describe the physico-chemical properties of matcha, the specific requirements for tea cultivation and industrial processing. The quality of matcha mainly depends on the quality of fresh tea leaves, which is affected by preharvest factors including tea cultivar, shading treatment, and fertilization. Shading is the key measure to increase greenness, reduce bitterness and astringency, and enhance umami taste of matcha. The potential health benefits of matcha and the gastrointestinal fate of main phenolics in matcha are covered. The chemical compositions and bioactivities of fiber-bound phenolics in matcha and other plant materials are discussed. The fiber-bound phenolics are considered promising components which endow matcha with boosted bioavailability of phenolics and health benefits through modulating gut microbiota.
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In visual working memory (VWM) tasks, participants' performances can be improved through the use of dimension-based retro-cues, which direct internal attention to prioritize a particular dimension (e.g., color or orientation) of VWM representations even after the stimuli disappear. This phenomenon is known as the dimension-based retro-cue benefit (RCB). The present study investigates whether sustained attention is required for the dimension-based RCB by inserting interference or interruption between the retro-cue and the test array to distract attention. We tested the effects of perceptual interference or cognitive interruption on dimension-based RCB when the interference (Experiments 1 and 2 with masks) or interruption (Experiments 3 and 4 with an odd-even task) occurred concurrently with the stages for the maintenance of prioritized information (long cue-and-interference/interruption interstimulus interval, e.g., Experiments 1 and 3) or the deployment of attention (short cue-and-interference/interruption interstimulus interval, e.g., Experiments 2 and 4). Our results demonstrate that perceptual interference or cognitive interruption attenuates the dimension-based RCB. These findings suggest that sustained attention is necessary for the effective prioritization of a specific dimension of VWM representations.
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Sinais (Psicologia) , Memória de Curto Prazo , Humanos , Percepção VisualRESUMO
Environmental temperature and humidity play a vital role in plant-pathogen interactions, which profoundly affect the occurrence of crop diseases. However, the specific methods and mechanisms through which intermittent changes in temperature and humidity mitigate plant diseases remain unclear. In this study, six temperature and humidity combinations were set, the disease severity of tomatoes and biomass of Botrytis cinerea were analyzed, and the infection process of pathogens was observed using an optical microscope. Furthermore, dual RNA-seq analysis was performed to explore the interactions between plants and pathogens. Results showed that the 24 hours postinoculation (hpi)-12 h day (regulation was performed at 24 hpi for 12 h after inoculation during the day) treatment reduced the gray mold severity and biomass of B. cinerea in plants by the greatest amount and effectively inhibited the growth of mycelia. The 24 hpi-12 h day treatment induced the upregulation of light reactions, photorespiration, and Calvin cycle-related genes in tomatoes, whereas fungal genes related to the biosynthesis of sesquiterpene botrydial and polyketide botcinic acid were downregulated. Overall, we identified the optimal combination of temperature and humidity changes to inhibit the development of tomato gray mold and preliminarily explored the interactions between tomato and B. cinerea under temperature and humidity changes. This work has practical importance and provides a theoretical basis for the ecological control of plant diseases.
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Solanum lycopersicum , Umidade , Solanum lycopersicum/genética , Temperatura , Regulação para CimaRESUMO
Temperature and humidity play an important role in plant-pathogen interactions. However, regulating the temperature and humidity specifically to inhibit the development of plant diseases remains unclear. In this study, we explored the influence of intermittent temperature and humidity variation on tomato gray mold. Intermittent regulation of temperature and humidity (increasing temperature with decreasing humidity for different periods within 24 h) inhibited the disease severity of plants and the infection process of Botrytis cinerea. The 4-h treatment (increasing temperature accompanied by decreasing humidity for 4 h and recovering for 4 h, and so on) effectively inhibited the development of tomato gray mold, reduced the biomass of B. cinerea, delayed the differentiation time of mycelia, and inhibited the accumulation of hydrogen peroxide in tomato leaves at the later stage of infection. The increased expressions of heat-shock protein (HSP) genes HSP20, HSP70, HSP90, BAG6, and BAG7 in tomato were mainly caused by environmental changes and environment-plant-pathogen interactions, and the increased expression of the latter was greater than that of the former in the 2-h (increasing temperature accompanied by decreasing humidity for 2 h and recovering for 2 h, and so on) and 4-h treatments. Pathogen infection induced the expression of defense-related genes in tomato, and the increase in the expressions of FLS2, FEI1, PI2, Pti5, and WRKY75 induced by B. cinerea in the 4-h treatment was greater than that under unregulated temperature and humidity conditions. In general, intermittent temperature and humidity variation can effectively inhibit the development of tomato gray mold, and the 4-h treatment had the best inhibitory effect.
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Solanum lycopersicum , Umidade , Solanum lycopersicum/genética , Temperatura , FungosRESUMO
PURPOSE: The genetic causes of oocyte maturation arrest leading to female infertility are largely unknown, and no population-based genetic analysis has been applied in cohorts of patients with infertility. We aimed to identify novel pathogenic genes causing oocyte maturation arrest by using a gene-based burden test. METHODS: Through comparison of exome sequencing data from 716 females with infertility characterized by oocyte maturation arrest and 3539 controls, we performed a gene-based burden test and identified a novel pathogenic gene LHX8. Splicing event was evaluated using a minigene assay, expression of LHX8 protein was assessed in HeLa cells, and nuclear subcellular localization was determined in both HeLa cells and mouse oocytes. RESULTS: A total of 5 heterozygous loss-of-function LHX8 variants were identified from 6 independent families (c.389+1G>T, c.412C>T [p.Arg138∗], c.282C>A [p.Cys94∗]; c.257dup [p.Tyr86∗]; and c.180del, [p.Ser61Profs∗30]). All the identified variants in LHX8 produced truncated LHX8 protein and resulted in loss of LHX8 nuclear localization in both HeLa cells and mouse oocytes. CONCLUSION: By combining genetic evidence and functional evaluations, we identified a novel pathogenic gene LHX8 and established the causative relationship between LHX8 haploinsufficiency and female infertility characterized by oocyte maturation arrest.
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Infertilidade Feminina , Feminino , Humanos , Camundongos , Animais , Infertilidade Feminina/genética , Infertilidade Feminina/patologia , Células HeLa , Oogênese/genética , Oócitos , Sequenciamento do ExomaRESUMO
Peptidyl arginine deiminase, type VI (PADI6) is a member of the subcortical maternal complex (SCMC), which plays vital roles in mammalian embryogenesis. Most mutations in SCMC members have been reported to cause human embryonic arrest, and a total of 15 mutations in PADI6 have been shown to be responsible for early embryonic arrest according to previous studies. However, the genetic factors behind this phenotype remain to be understood in further detail. Here, we identified 13 novel mutations and 4 previously reported mutations of PADI6 in 14 patients who were diagnosed with abnormal embryonic development caused by early arrest, embryonic fragmentation, and recurrent implantation failure. Most of the mutations were predicted by in silico analysis to be deleterious or damaging to the function of PADI6. In addition, the total and East Asian population frequencies of the mutations were low or absent in the gnomAD database. Our study expands the mutational spectrum in PADI6 and will provide precise targets for genetic counseling in the future.
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Mamíferos , Oócitos , Animais , Desenvolvimento Embrionário , Feminino , Humanos , Mutação , Fenótipo , Gravidez , Proteína-Arginina Desiminase do Tipo 6RESUMO
STUDY QUESTION: Are there new genetic factors responsible for male infertility with normal sperm quantity and morphology? SUMMARY ANSWER: We identified the bi-allelic variants in KCNU1 and confirmed it a novel pathogenetic gene for male infertility mainly due to impaired sperm acrosome reactions (ARs). WHAT IS KNOWN ALREADY: Until now, the underlying genetic determinants for male affected individuals exhibiting normal sperm quantity and morphology have been largely unknown. Potassium/calcium-activated channel subfamily U member 1 (KCNU1) is a sperm-specific potassium channel. The Kcnu1 null mutation in male mice causes infertility due to the impaired progressive motility and AR. STUDY DESIGN, SIZE, DURATION: We recruited a cohort of 126 male infertility individuals with typical asthenospermia or fertilization failure and focused on two infertile males from two consanguineous families from 2015 to 2020; whole-exome sequencing and homozygosity mapping were performed. We identified a homozygous missense variant (c.2144A>G, p.His715Arg) and a homozygous donor splice-site variant (c.1295 + 3A>C, p.Val405Glyfs*8) in KCNU1. Then, we generated a knock-in (KI) mouse model in September 2020 and have now carried out functional studies and possible treatment strategies. PARTICIPANTS/MATERIALS, SETTING, METHODS: The affected individuals with infertility were recruited from the Shanghai Ninth Hospital affiliated to Shanghai Jiao Tong University. Genomic DNA from the affected individual was extracted from peripheral blood. Whole-exome sequencing, homozygosity mapping and in silico analyses were used to screen and identify KCNU1 variants, and the variants were confirmed by Sanger sequencing. We used C57BL/6N mouse to construct KI mouse model to mimic the reproductive phenotype in vivo. We performed functional experiments by western blotting, AR assay and immunofluorescent Staining. Finally, we performed IVF and ICSI to explore the treatment strategies. MAIN RESULTS AND THE ROLE OF CHANCE: We identified a homozygous missense variant (c.2144A>G, p.His715Arg) and a homozygous donor splice-site variant (c.1295 + 3A>C, p.Val405Glyfs*8) in KCNU1 in two infertile males. We demonstrated that the splice-site variant affected normal alternative splicing of KCNU1, thus leading to the loss of function of KCNU1. Meanwhile, the missense pathogenic variant reduced the KCNU1 protein levels in sperm of both the affected individual and the KI mouse model, resulting in impaired ARs and male infertility. Intracytoplasmic sperm injection was able to rescue the deficiencies. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The exact molecular mechanism of KCNU1 and pathways need to be further explore in the future. WIDER IMPLICATIONS OF THE FINDINGS: This is the first report that establishes a causal relationship between KCNU1 deficiency and male infertility, confirming the critical role of KCNU1 in human reproduction. Our findings expand our knowledge of the genes that play critical roles in the human sperm AR and provide a new genetic marker for infertility. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the SHIPM-pi fund no. JY201801 from the Shanghai Institute of Precision Medicine, Ninth People's Hospital Shanghai Jiao Tong University School of Medicine, the National Natural Science Foundation of China (81725006, 81771649, 81822019, 81771581, 81971450, 81971382, 82001538 and 82071642). The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.
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Reação Acrossômica , Infertilidade Masculina , Canais de Potássio Ativados por Cálcio de Condutância Alta , Reação Acrossômica/genética , Animais , China , Humanos , Infertilidade Masculina/genética , Canais de Potássio Ativados por Cálcio de Condutância Alta/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sêmen , EspermatozoidesRESUMO
Most objects show high degrees of spatial regularity (e.g. beach umbrellas appear above, not under, beach chairs). The spatial regularities of real-world objects benefit visual working memory (VWM), but the mechanisms behind this spatial regularity effect remain unclear. The "encoding specificity" hypothesis suggests that spatial regularity will enhance the visual encoding process but will not facilitate the integration of information online during VWM maintenance. The "perception-alike" hypothesis suggests that spatial regularity will function in both visual encoding and online integration during VWM maintenance. We investigated whether VWM integrates sequentially presented real-world objects by focusing on the existence of the spatial regularity effect. Throughout five experiments, we manipulated the presentation (simultaneous vs. sequential) and regularity (with vs. without regularity) of memory arrays among pairs of real-world objects. The spatial regularity of memory objects presented simultaneously, but not sequentially, improved VWM performance. We also examined whether memory load, verbal suppression and masking, and memory array duration hindered the spatial regularity effect in sequential presentation. We found a stable absence of the spatial regularity effect, suggesting that the participants were unable to integrate real-world objects based on spatial regularities online. Our results support the encoding specificity hypothesis, wherein the spatial regularity of real-world objects can enhance the efficiency of VWM encoding, but VWM cannot exploit spatial regularity to help organize sampled sequential information into meaningful integrations.