NLRP14 Safeguards Calcium Homeostasis via Regulating the K27 Ubiquitination of Nclx in Oocyte-to-Embryo Transition.

Sperm-induced Ca2+ rise is critical for driving oocyte activation and subsequent embryonic development, but little is known about how lasting Ca2+ oscillations are regulated. Here it is shown that NLRP14, a maternal effect factor, is essential for keeping Ca2+ oscillations and early embryonic development. Few embryos lacking maternal NLRP14 can develop beyond the 2-cell stage. The impaired developmental potential of Nlrp14-deficient oocytes is mainly caused by disrupted cytoplasmic function and calcium homeostasis due to altered mitochondrial distribution, morphology, and activity since the calcium oscillations and development of Nlrp14-deficient oocytes can be rescued by substitution of whole cytoplasm by spindle transfer. Proteomics analysis reveal that cytoplasmic UHRF1 (ubiquitin-like, containing PHD and RING finger domains 1) is significantly decreased in Nlrp14-deficient oocytes, and Uhrf1-deficient oocytes also show disrupted calcium homeostasis and developmental arrest. Strikingly, it is found that the mitochondrial Na+ /Ca2+ exchanger (NCLX) encoded by Slc8b1 is significantly decreased in the Nlrp14mNull oocyte. Mechanistically, NLRP14 interacts with the NCLX intrinsically disordered regions (IDRs) domain and maintain its stability by regulating the K27-linked ubiquitination. Thus, the study reveals NLRP14 as a crucial player in calcium homeostasis that is important for early embryonic development.

Nitrogen-doped hydrochar prepared by biomass and nitrogen-containing wastewater for dye adsorption: Effect of nitrogen source in wastewater on the adsorption performance of hydrochar.

Dye wastewater has become one of the main risk sources of environmental pollution due to its high toxicity and difficulty in degradation. Hydrochar prepared by hydrothermal carbonization (HTC) of biomass has abundant surface oxygen-containing functional groups, and therefore is used as an adsorbent to remove water pollutants. The adsorption performance of hydrochar can be enhanced after improving its surface characteristics through nitrogen-doping (N-doping). In this study, wastewater rich in nitrogen sources such as urea, melamine and ammonium chloride were selected as the water source for the preparation of HTC feedstock. The N atoms were doped in the hydrochar with a content of 3.87%-5.70%, and mainly in the form of pyridinic-N, pyrrolic-N and graphitic-N, which changed the acidity and basicity of the hydrochar surface. The N-doped hydrochar adsorbed methylene blue (MB) and congo red (CR) in wastewater through pore filling, Lewis acid-base interaction, hydrogen bond, and π-π interaction, and the maximum adsorption capacities of those were obtained with 57.52 mg/g and 62.19 mg/g, respectively. However, the adsorption performance of N-doped hydrochar was considerably affected by the acid-base property of the wastewater. In a basic environment, the surface carboxyl of the hydrochar exhibited a high negative charge and thus an enhanced electrostatic interaction with MB. Whereas, the hydrochar surface was positively charged in an acid environment by binding H+, resulting in an enhanced electrostatic interaction with CR. Therefore, the adsorption efficiency of MB and CR by N-doped hydrochar can be tuned by adjusting the nitrogen source and the pH of the wastewater.

SRSF10 is essential for progenitor spermatogonia expansion by regulating alternative splicing.

Alternative splicing expands the transcriptome and proteome complexity and plays essential roles in tissue development and human diseases. However, how alternative splicing regulates spermatogenesis remains largely unknown. Here, using a germ cell-specific knockout mouse model, we demonstrated that the splicing factor Srsf10 is essential for spermatogenesis and male fertility. In the absence of SRSF10, spermatogonial stem cells can be formed, but the expansion of Promyelocytic Leukemia Zinc Finger (PLZF)-positive undifferentiated progenitors was impaired, followed by the failure of spermatogonia differentiation (marked by KIT expression) and meiosis initiation. This was further evidenced by the decreased expression of progenitor cell markers in bulk RNA-seq, and much less progenitor and differentiating spermatogonia in single-cell RNA-seq data. Notably, SRSF10 directly binds thousands of genes in isolated THY+ spermatogonia, and Srsf10 depletion disturbed the alternative splicing of genes that are preferentially associated with germ cell development, cell cycle, and chromosome segregation, including Nasp, Bclaf1, Rif1, Dazl, Kit, Ret, and Sycp1. These data suggest that SRSF10 is critical for the expansion of undifferentiated progenitors by regulating alternative splicing, expanding our understanding of the mechanism underlying spermatogenesis.

Effect of corn stover hydrochar on anaerobic digestion performance of its associated wastewater.

Hydrothermal carbonation (HTC) is an effective method to enhance the fuel quality of biomass in a subcritical water environment, but generates large amounts of wastewater (HTCWW), which was converted through anaerobic digestion (AD) into methane in this study. However, the toxic and refractory substances contained in HTCWW tended to cause operation instability of the AD system. The solid product in HTC of corn stover (CS), named CS hydrochar, was modified with KOH immersion and then added to the AD reactor to improve the methanogenic performance. The results showed that the optimum dosage of modified hydrochar (MCH) was 15 g/L, and the COD removal rate was increased by 19.3% and methane yield was increased by 42.3%-301 mL/g-COD, as the pore and the oxygen-containing functional groups of MCH provided colonization points for microorganisms, and also enhanced the electron transfer efficiency among methanogenic archaea. In addition, the increased alkalinity of MCH due to alkaline modification increased the pH buffering capability, and accelerated the consumption of acetic acid and butyric acid in the early AD stage (0-8 days) and propionic acid in the late AD stage (12-18 days), which then alleviated the organic acid accumulation and reduced the lag period by 2 days. The adverse effects of toxic and refractory substances of HTCWW on the AD performance were also decreased due to the adsorption of MCH at the beginning of the AD process, and latterly the adsorbed substances could be degraded by the microorganisms colonized on the MCH surface. The finding of this study showed AD is a feasible method to recover organic energy contained in HTCWW, and the associated hydrochar can be used as an effective promoter for the AD of HTCWW.

Maternal EHMT2 is essential for homologous chromosome segregation by regulating Cyclin B3 transcription in oocyte meiosis.

During oocyte growth, various epigenetic modifications are gradually established, accompanied by accumulation of large amounts of mRNAs and proteins. However, little is known about the relationship between epigenetic modifications and meiotic progression. Here, by using Gdf9-Cre to achieve oocyte-specific ablation of Ehmt2 (Euchromatic-Histone-Lysine-Methyltransferase 2) from the primordial follicle stage, we found that female mutant mice were infertile. Oocyte-specific knockout of Ehmt2 caused failure of homologous chromosome separation independent of persistently activated SAC during the first meiosis. Further studies revealed that lacking maternal Ehmt2 affected the transcriptional level of Ccnb3, while microinjection of exogenous Ccnb3 mRNA could partly rescue the failure of homologous chromosome segregation. Of particular importance was that EHMT2 regulated ccnb3 transcriptions by regulating CTCF binding near ccnb3 gene body in genome in oocytes. In addition, the mRNA level of Ccnb3 significantly decreased in the follicles microinjected with Ctcf siRNA. Therefore, our findings highlight the novel function of maternal EHMT2 on the metaphase I-to-anaphase I transition in mouse oocytes: regulating the transcription of Ccnb3.

Conceptual Compression via Deep Structure and Texture Synthesis.

Existing compression methods typically focus on the removal of signal-level redundancies, while the potential and versatility of decomposing visual data into compact conceptual components still lack further study. To this end, we propose a novel conceptual compression framework that encodes visual data into compact structure and texture representations, then decodes in a deep synthesis fashion, aiming to achieve better visual reconstruction quality, flexible content manipulation, and potential support for various vision tasks. In particular, we propose to compress images by a dual-layered model consisting of two complementary visual features: 1) structure layer represented by structural maps and 2) texture layer characterized by low-dimensional deep representations. At the encoder side, the structural maps and texture representations are individually extracted and compressed, generating the compact, interpretable, inter-operable bitstreams. During the decoding stage, a hierarchical fusion GAN (HF-GAN) is proposed to learn the synthesis paradigm where the textures are rendered into the decoded structural maps, leading to high-quality reconstruction with remarkable visual realism. Extensive experiments on diverse images have demonstrated the superiority of our framework with lower bitrates, higher reconstruction quality, and increased versatility towards visual analysis and content manipulation tasks.

Properties and the Application of Sludge-Based Biochar in the Removal of Phosphate and Methylene Blue from Water: Effects of Acid Treating.

Sludge-based biochar could be used to remove phosphate and methylene blue (MB) from water. It is a highly efficient way to treat the sludge and contaminated water synergistically. The high ash content in sludge greatly influenced the adsorption property of the resultant biochar. In this work, the influence of carbonization-activation and acid treating on the adsorption performance of the sludge-based biochar was evaluated. The composition, structure, and surface properties of biochar were improved after acid treating. The biochar was obtained in a sequence of carbonization-activation first and then acid treating, providing the optimal adsorption property. Zn550-H and Zn750-H showed excellent adsorption capacity to phosphate and MB, respectively. The adsorption process was well described by the pseudo-first-order and pseudo-second-order kinetic models. Isothermal studies implied that it was controlled by multiple processes. What is more, sludge-based biochar performed well in the adsorption of phosphate and MB from weakly acidic to alkaline conditions, which was beneficial to utilize the sludge-based biochar in water remediation practically.

Energy conversion performance in co-hydrothermal carbonization of sewage sludge and pinewood sawdust coupling with anaerobic digestion of the produced wastewater.

Energy conversion and utilization of sewage sludge (SS) and lignocellulosic biomass are an important measure to deal with environmental pollution and resource utilization. Addressing the waste by-product in a clean way is essential. In this study, solid char fuel (hydrochar) was obtained through co-hydrothermal carbonization of SS with pinewood sawdust (PS), and methane gas was obtained through anaerobic digestion (AD) of hydrothermal carbonization wastewater (HTCWW). The energy conversion performance of the feedstock organics under different HTC conditions (temperature of 160 °C, 220 °C, and 280 °C; reaction time of 0, 2, and 4 h; feedstock liquid-solid mass ratio of 4:1, 10:1, and 16:1), and the mass and energy yields of hydrochar and methane and their influencing factors were emphasized. More than 60% of the energy in SS and PS can be recovered by coupling the HTC-AD process. With the increase in hydrothermal reaction temperature and reaction time, the mass yield of hydrochar decreased, but the higher heating value increased. The maximum energy yield of hydrochar was 86.47% under the HTC temperature of 160 °C, liquid-solid ratio of 10:1, and reaction time of 2 h. The HTCWW obtained at a lower temperature (160 °C) showed the highest cumulative methane yield of 304.16 mL-CH4/g-COD.

DFT study on the effect of functional groups of carbonaceous surface on ammonium adsorption from water.

Density functional theory (DFT) was used to study the adsorption of ammonium ion on carbon materials. The effects of single and multiple adjacent functional groups of carbon structures on ammonium ion adsorption were emphasized. The electrostatic potential, adsorption energy, charge transfer, molecular orbital, and dipole moment of different configurations were analyzed. Results showed that the carbonyl group was more likely to adsorb ammonium ion than lactone, carboxyl, and hydroxyl. When the carbon material contained multiple adjacent functional groups at the same time, the adsorption of ammonium ion can be promoted or inhibited due to the interaction among functional groups. The effect of functional groups on the adsorption of π bond in carbon materials was related to the electronegativity of functional groups, i.e., greater electronegativity led to smaller adsorption energy of π bond. Carbon material itself is nonpolar and hydrophobic, so adding oxygen-containing functional groups can increase the dipole moment of carbon material molecules, thereby enhancing its polarity and adsorption capacity.

Single cell RNA sequencing techniques and applications in research of ovary development and related diseases.

The ovary is a highly organized composite of germ cells and various types of somatic cells, whose communications dictate ovary development to generate functional oocytes. The differences between individual cells might have profound effects on ovary functions. Single cell RNA sequencing techniques are promising approaches to explore the cell type composition of organisms, the dynamics of transcriptomes, the regulatory network between genes and the signaling pathways between cell types at the single cell resolution. In this review, we provide an overview of the currently available single cell RNA sequencing techniques including Smart-seq2 and Drop-seq, as well as their applications in biological and clinical research to provide a better understanding on the molecular mechanisms underlying ovary development and associated diseases.

The redistribution and migration mechanism of nitrogen in the hydrothermal co­carbonization process of sewage sludge and lignocellulosic wastes.

Blending lignocellulosic wastes (such as cornstalk, CS) into sewage sludge (SS) for hydrothermal carbonization (HTC) could contribute to the importance of the hydrothermal solid product (hydrochar) as a substitute for fossil fuel. However, the interactions between SS and CS changed the fate of Nitrogen (N), affecting the clean combustion utilization of hydrochar. This study focused on the influence of SS-CS interactions on the redistribution and migration behavior of N during the co-HTC process by tuning the mass ratio of SS to CS (SS:CS), reaction temperature, and residence time. Under the hydrothermal condition of 220 °C, 2 h, and SS:CS = 1:1, the high heating value of hydrochar and the energy recovery efficiency (ERE) respectively reached 15.89 MJ/kg and 71.19%. Further raising the temperature to 250 °C, the hydrochar was enhanced in the coalification degree, whereas ERE decreased to 61.86%. Part of the amino-N in sludge organics was fractured during the co-HTC process and reacted with carbohydrate and intermediate products, such as 5-hydroxymethylfurfural, which degraded from CS, to generate heterocyclic-N compounds (including pyridine, pyrrole, and pyrazine). The remaining amino-N formed pyridine-N, pyrrole-N, and quaternary-N through various solid-solid conversions. The heterocyclic-N polymerized and formed melanoidins, which thereafter polymerized with aromatic clusters to form the N-containing polyaromatic char. Therefore, the N retention rate (NRR) was enhanced and showed a synergistic effect. NRR was increased by raising the proportion of CS or extending time, reaching 57.02% at SS:CS = 1:1 and 8 h. Conversely, rising temperatures resulted in a downward trend of NRR with a phased increase at 220 °C-250 °C.

Nuclear and cytoplasmic quality of oocytes derived from serum-free culture of secondary follicles in vitro.

In vitro culture of follicles is a promising technology to generate large quantities of mature oocytes and it could offer a novel option of assisted reproductive technologies. Here we described a 2-dimensional follicular serum-free culture system with 3-dimensional effect that can make secondary follicles develop into antral follicles (78.52%), generating developmentally mature oocytes in vitro (66.45%). The oocytes in this serum-free system completed the first meiosis; spindle assembly and chromosome congression in most oocytes matured from follicular culture were normal. However, these oocytes showed significantly lower activation and embryonic development rates, and their ability to produce Ca2+ oscillations was also lower in response to parthenogenetic activation, after which a 2-cell embryonic developmental block occurred. Oocytes matured from follicular culture displayed increased abnormal mitochondrial distribution and increased reactive oxygen species levels when compared to in vivo matured oocytes. These data are important for understanding the reasons for reduced developmental potential of oocytes matured from follicular culture, and for further improving the cultivation system.

Single-cell RNA sequencing reveals species-specific time spans of cell cycle transitions in early oogenesis.

Oogenesis is a highly regulated process and its basic cellular events are evolutionarily conserved. However, the time spans of oogenesis differ substantially among species. To explore these interspecies differences in oogenesis, we performed single-cell RNA-sequencing on mouse and monkey female germ cells and downloaded the single-cell RNA-sequencing data of human female germ cells. The cell cycle analyses indicate that the period and extent of cell cycle transitions are significantly different between the species. Moreover, hierarchical clustering of critical cell cycle genes and the interacting network of cell cycle regulators also exhibit distinguished patterns across species. We propose that differences in the regulation of cell cycle transitions may underlie female germ cell developmental allochrony between species. A better understanding of the cell cycle transition machinery will provide new insights into the interspecies differences in female germ cell developmental time spans.

PRC2 and EHMT1 regulate H3K27me2 and H3K27me3 establishment across the zygote genome.

The formation of zygote is the beginning of mammalian life, and dynamic epigenetic modifications are essential for mammalian normal development. H3K27 di-methylation (H3K27me2) and H3K27 tri-methylation (H3K27me3) are marks of facultative heterochromatin which maintains transcriptional repression established during early development in many eukaryotes. However, the mechanism underlying establishment and regulation of epigenetic asymmetry in the zygote remains obscure. Here we show that maternal EZH2 is required for the establishment of H3K27me3 in mouse zygotes. However, combined immunostaining with ULI-NChIP-seq (ultra-low-input micrococcal nuclease-based native ChIP-seq) shows that EZH1 could partially safeguard the role of EZH2 in the formation of H3K27me2. Meanwhile, we identify that EHMT1 is involved in the establishment of H3K27me2, and that H3K27me2 might be an essential prerequisite for the following de novo H3K27me3 modification on the male pronucleus. In this work, we clarify the establishment and regulatory mechanisms of H3K27me2 and H3K27me3 in mouse zygotes.

Single-cell RNA sequencing reveals regulation of fetal ovary development in the monkey (Macaca fascicularis).

Germ cells are vital for reproduction and heredity. However, the mechanisms underlying female germ cell development in primates, especially in late embryonic stages, remain elusive. Here, we performed single-cell RNA sequencing of 12,471 cells from whole fetal ovaries, and explored the communications between germ cells and niche cells. We depicted the two waves of oogenesis at single-cell resolution and demonstrated that progenitor theca cells exhibit similar characteristics to Leydig cells in fetal monkey ovaries. Notably, we found that ZGLP1 displays differentially expressed patterns between mouse and monkey, which is not overlapped with NANOG in monkey germ cells, suggesting its role in meiosis entry but not in activating oogenic program in primates. Furthermore, the majority of germ cell clusters that sharply express PRDM9 and SPO11 might undergo apoptosis after cyst breakdown, leading to germ cell attrition. Overall, our work provides new insights into the molecular and cellular basis of primate fetal ovary development at single-cell resolution.

High-throughput sequencing reveals landscapes of female germ cell development.

Female germ cell development is a highly complex process that includes meiosis initiation, oocyte growth recruitment, oocyte meiosis retardation and resumption and final meiotic maturation. A series of coordinated molecular signaling factors ensure successful oogenesis. The recent rapid development of high-throughput sequencing technologies allows for the dynamic omics in female germ cells, which is essential for further understanding the regulatory mechanisms of molecular events comprehensively. In this review, we summarize the current literature of multi-omics sequenced by epigenome-, transcriptome- and proteome-associated technologies, which provide valuable information for understanding the regulation of key events during female germ cell development.

RNA-Seq transcriptome reveals different molecular responses during human and mouse oocyte maturation and fertilization.

BACKGROUND: Female infertility is a worldwide concern and the etiology of infertility has not been thoroughly demonstrated. Although the mouse is a good model system to perform functional studies, the differences between mouse and human also need to be considered. The objective of this study is to elucidate the different molecular mechanisms underlying oocyte maturation and fertilization between human and mouse. RESULTS: A comparative transcriptome analysis was performed to identify the differentially expressed genes and associated biological processes between human and mouse oocytes. In total, 8513 common genes, as well as 15,165 and 6126 uniquely expressed genes were detected in human and mouse MII oocytes, respectively. Additionally, the ratios of non-homologous genes in human and mouse MII oocytes were 37 and 8%, respectively. Functional categorization analysis of the human MII non-homologous genes revealed that cAMP-mediated signaling, sister chromatid cohesin, and cell recognition were the major enriched biological processes. Interestingly, we couldn't detect any GO categories in mouse non-homologous genes. CONCLUSIONS: This study demonstrates that human and mouse oocytes exhibit significant differences in gene expression profiles during oocyte maturation, which probably deciphers the differential molecular responses to oocyte maturation and fertilization. The significant differences between human and mouse oocytes limit the generalizations from mouse to human oocyte maturation. Knowledge about the limitations of animal models is crucial when exploring a complex process such as human oocyte maturation and fertilization.

Single-cell RNA sequencing reveals the landscape of early female germ cell development.

Meiosis initiation is a crucial step for the production of haploid gametes, which occurs from anterior to posterior in fetal ovaries. The asynchrony of the transition from mitosis to meiosis results in heterogeneity in the female germ cell populations, which limits the studies of meiosis initiation and progression at a higher resolution level. To dissect the process of meiosis initiation, we investigated the transcriptional profiles of 19 363 single germ cells collected from E12.5, E14.5, and E16.5 mouse fetal ovaries. Clustering analysis identified seven groups and defined dozens of corresponding transcription factors, providing a global view of cellular differentiation from primordial germ cells toward meiocytes. Furthermore, we explored the dynamics of gene expression within the developmental trajectory with special focus on the critical state of meiosis. We found that meiosis initiation occurs as early as E12.5 and the cluster of oogonia_4 is the critical state between mitosis and meiosis. Our data provide key insights into the transcriptome features of peri-meiotic female germ cells, which offers new information not only on meiosis initiation and progression but also on screening pathogenic mutations in meiosis-associated diseases.

Deletion of BAF250a affects oocyte epigenetic modifications and embryonic development.

BRG1-associated factor 250a (BAF250a) is a component of the SWI/SNF adenosine triphosphate-dependent chromatin remodeling complex, which has been shown to control chromatin structure and transcription. BAF250a was reported to be a key component of the gene regulatory machinery in embryonic stem cells controlling self-renewal, differentiation, and cell lineage decisions. Here we constructed Baf250aF/F ;Gdf9-cre (Baf250aCKO ) mice to specifically delete BAF250a in oocytes to investigate the role of maternal BAF250a in female germ cells and embryo development. Our results showed that BAF250a deletion did not affect folliculogenesis, ovulation, and fertilization, but it caused late embryonic death. RNA sequencing analysis showed that the expression of genes involved in cell proliferation and differentiation, tissue morphogenesis, histone modification, and nucleosome remodeling were perturbed in Baf250aCKO MII oocytes. We showed that covalent histone modifications such as H3K27me3 and H3K27ac were also significantly affected in oocytes, which may reduce oocyte quality and lead to birth defects. In addition, the DNA methylation level of Igf2r, Snrpn, and Peg3 differentially methylated regions was decreased in Baf250aCKO oocytes. Quantitative real-time polymerase chain reaction analysis showed that the relative messenger RNA (mRNA) expression levels of Igf2r and Snrpn were significantly increased. The mRNA expression level of Dnmt1, Dnmt3a, Dnmt3l, and Uhrf1 was decreased, and the protein expression in these genes was also reduced, which might be the cause for impaired imprinting establishment. In conclusion, our results demonstrate that BAF250a plays an important role in oocyte transcription regulation, epigenetic modifications, and embryo development.

Degradation of Ccnb3 is essential for maintenance of MII arrest in oocyte.

Before fertilization, ovulated mammalian oocytes are arrested at the metaphase of second meiosis (MII), which is maintained by the so-called cytostatic factor (CSF). It is well known that the continuous synthesis and accumulation of cyclin B is critical for maintaining the CSF-mediated MII arrest. Recent studies by us and others have shown that Ccnb3 is required for the metaphase-to-anaphase transition during the first meiosis of mouse oocytes, but whether Ccnb3 plays a role in MII arrest and exit remains unknown. Here, we showed that the protein level of Ccnb3 gradually decreased during oocyte meiotic maturation, and exogenous expression of Ccnb3 led to release of MII arrest, degradation of securin, separation of sister chromatids, extrusion of the second polar body (PB2), and finally entry into interphase. These phenotypes could be rescued by inhibition of Wee1B or CDK2. Our results indicate that Ccnb3 plays a critical regulatory role in MII arrest and exit in mouse oocytes.