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Background: RNA splicing plays significant roles in fundamental biological activities. However, our knowledge about the roles of alternative splicing and underlying mechanisms during spermatogenesis is limited. Results: Here, we report that Serine/arginine-rich splicing factor 2 (SRSF2), also known as SC35, plays critical roles in alternative splicing and male reproduction. Male germ cell-specific deletion of Srsf2 by Stra8-Cre caused complete infertility and defective spermatogenesis. Further analyses revealed that deletion of Srsf2 disrupted differentiation and meiosis initiation of spermatogonia. Mechanistically, by combining RNA-seq data with LACE-seq data, we showed that SRSF2 regulatory networks play critical roles in several major events including reproductive development, spermatogenesis, meiotic cell cycle, synapse organization, DNA recombination, chromosome segregation, and male sex differentiation. Furthermore, SRSF2 affected expression and alternative splicing of Stra8, Stag3 and Atr encoding critical factors for spermatogenesis in a direct manner. Conclusions: Taken together, our results demonstrate that SRSF2 has important functions in spermatogenesis and male fertility by regulating alternative splicing. © 2023, BioMed Central Ltd., part of Springer Nature.

Gefitinib is a first-line anti-cancer drug for the treatment of advanced non-small cell lung cancer (NSCLC). It has been reported that gefitinib can generate several drug-related adverse effects, including nausea, peripheral edema, decreased appetite and rash. However, the reproductive toxicity of gefitinib has not been clearly defined until now. Here we assessed the effects of gefitinib on oocyte quality by examining the critical events and molecular changes of oocyte maturation. Gefitinib at 1, 2, 5 or 10 μM concentration was added to culture medium (M2). We found that gefitinib at its median peak concentration of 1 μM did not affect oocyte maturation, but 5 μM gefitinib severely blocked oocyte meiotic progression as indicated by decreased rates of germinal vesicle breakdown (GVBD) and polar body extrusion (PBE). We further showed that gefitinib treatment increased phosphorylation of CDK1 at the site of Try15, inhibited cyclin B1 entry into the nucleus, and disrupted normal spindle assembly, chromosome alignment and mitochondria dynamics, finally leading to the generation of aneuploidy and early apoptosis of oocytes. Our study reported here provides valuable evidence for reproductive toxicity of gefitinib administration employed for the treatment of cancer patients. © 2021 Elsevier B.V.

Patulin is a secondary metabolite mainly secreted by fungi and is the most common mycotoxin found in apples and apple-based products. For the past few years, numerous studies suggested the wide distribution and toxicity of patulin. In this study, we investigated the toxic effect of patulin on mouse oocytes and its possible mechanisms. The results showed that patulin treatment did not affect meiotic resumption, but inhibited oocyte maturation as indicated by failure of first polar body extrusion. Further mechanistic study showed that patulin treatment disturbed normal spindle assembly, chromosome alignment and morphology. We also found increased oxidative stress by testing the level of ROS and decreased mitochondrial membrane potential, indicating mitochondria dysfunction. In summary, our results suggest that patulin treatment causes oocyte meiotic arrest by disturbing normal spindle assembly and chromosome alignment, which may be caused by dysfunctions of mitochondria.

The quality of oocytes is a vital factor for embryo development. Meiotic progression through metaphase I usually takes a relatively long time to ensure correct chromosome separation, a process that is critical for determining oocyte quality. Here, we report that cell division cycle 5-like (Cdc5L) plays a critical role in regulating metaphase-to-anaphase I transition during mouse oocyte meiotic maturation. Knockdown of Cdc5L by small interfering RNA injection did not affect spindle assembly but caused metaphase I arrest and subsequent reduced first polar body extrusion due to insufficient anaphase-promoting complex/cyclosome activity. We further showed that Cdc5L could also directly interact with securin, and Cdc5L knockdown led to a continuous high expression level of securin, causing severely compromised meiotic progression. The metaphase-to-anaphase I arrest caused by Cdc5L knockdown could be rescued by knockdown of endogenous securin. In summary, we reveal a novel role for Cdc5L in regulating mouse oocyte meiotic progression by interacting with securin. © Copyright © 2021 Zhang, Li, Ouyang, Meng, Zhang, Yue, Sun and Qian.

Patulin is a secondary metabolite mainly secreted by fungi and is the most common mycotoxin found in apples and apple-based products. For the past few years, numerous studies suggested the wide distribution and toxicity of patulin. In this study, we investigated the toxic effect of patulin on mouse oocytes and its possible mechanisms. The results showed that patulin treatment did not affect meiotic resumption, but inhibited oocyte maturation as indicated by failure of first polar body extrusion. Further mechanistic study showed that patulin treatment disturbed normal spindle assembly, chromosome alignment and morphology. We also found increased oxidative stress by testing the level of ROS and decreased mitochondrial membrane potential, indicating mitochondria dysfunction. In summary, our results suggest that patulin treatment causes oocyte meiotic arrest by disturbing normal spindle assembly and chromosome alignment, which may be caused by dysfunctions of mitochondria.

The quality of oocytes is a vital factor for embryo development. Meiotic progression through metaphase I usually takes a relatively long time to ensure correct chromosome separation, a process that is critical for determining oocyte quality. Here, we report that cell division cycle 5-like (Cdc5L) plays a critical role in regulating metaphase-to-anaphase I transition during mouse oocyte meiotic maturation. Knockdown of Cdc5L by small interfering RNA injection did not affect spindle assembly but caused metaphase I arrest and subsequent reduced first polar body extrusion due to insufficient anaphase-promoting complex/cyclosome activity. We further showed that Cdc5L could also directly interact with securin, and Cdc5L knockdown led to a continuous high expression level of securin, causing severely compromised meiotic progression. The metaphase-to-anaphase I arrest caused by Cdc5L knockdown could be rescued by knockdown of endogenous securin. In summary, we reveal a novel role for Cdc5L in regulating mouse oocyte meiotic progression by interacting with securin. © Copyright © 2021 Zhang, Li, Ouyang, Meng, Zhang, Yue, Sun and Qian.

The quality of oocytes is a vital factor for embryo development. Meiotic progression through metaphase I usually takes a relatively long time to ensure correct chromosome separation, a process that is critical for determining oocyte quality. Here, we report that cell division cycle 5-like (Cdc5L) plays a critical role in regulating metaphase-to-anaphase I transition during mouse oocyte meiotic maturation. Knockdown of Cdc5L by small interfering RNA injection did not affect spindle assembly but caused metaphase I arrest and subsequent reduced first polar body extrusion due to insufficient anaphase-promoting complex/cyclosome activity. We further showed that Cdc5L could also directly interact with securin, and Cdc5L knockdown led to a continuous high expression level of securin, causing severely compromised meiotic progression. The metaphase-to-anaphase I arrest caused by Cdc5L knockdown could be rescued by knockdown of endogenous securin. In summary, we reveal a novel role for Cdc5L in regulating mouse oocyte meiotic progression by interacting with securin. © Copyright © 2021 Zhang, Li, Ouyang, Meng, Zhang, Yue, Sun and Qian.

Patulin is a secondary metabolite mainly secreted by fungi and is the most common mycotoxin found in apples and apple-based products. For the past few years, numerous studies suggested the wide distribution and toxicity of patulin. In this study, we investigated the toxic effect of patulin on mouse oocytes and its possible mechanisms. The results showed that patulin treatment did not affect meiotic resumption, but inhibited oocyte maturation as indicated by failure of first polar body extrusion. Further mechanistic study showed that patulin treatment disturbed normal spindle assembly, chromosome alignment and morphology. We also found increased oxidative stress by testing the level of ROS and decreased mitochondrial membrane potential, indicating mitochondria dysfunction. In summary, our results suggest that patulin treatment causes oocyte meiotic arrest by disturbing normal spindle assembly and chromosome alignment, which may be caused by dysfunctions of mitochondria.

PKCβI, a member of the classical protein kinase C family, plays key roles in regulating cell cycle transition. Here, we report the expression, localization and functions of PKCβI in mouse oocyte meiotic maturation. PKCβI and p-PKCβI (phosphor-PKCβI) were expressed from germinal vesicle (GV) stage to metaphase II (MII) stage. Confocal microscopy revealed that PKCβI was localized in the GV and evenly distributed in the cytoplasm after GV breakdown (GVBD), and it was concentrated at the midbody at telophase in meiotic oocytes. While, p-PKCβI was concentrated at the spindle poles at the metaphase stages and associated with midbody at telophase. Depletion of PKCβI by specific siRNA injection resulted in defective spindles, accompanied with spindle assembly checkpoint activation, metaphase I arrest and failure of first polar body (PB1) extrusion. Live cell imaging analysis also revealed that knockdown of PKCβI resulted in abnormal spindles, misaligned chromosomes, and meiotic arrest of oocytes arrest at the Pro-MI/MI stage. PKCβI depletion did not affect the G2/M transition, but its overexpression delayed the G2/M transition through regulating Cyclin B1 level and Cdc2 activity. Our findings reveal that PKCβI is a critical regulator of meiotic cell cycle progression in oocytes. Abbreviations: PKC, protein kinase C; COC, cumulus-oocyte complexes; GV, germinal vesicle; GVBD, germinal vesicle breakdown; Pro-MI, first pro-metaphase; MI, first metaphase; Tel I, telophase I; MII, second metaphase; PB1, first polar body; SAC, spindle assembly checkpoint. © 2019, © 2019 Informa UK Limited, trading as Taylor & Francis Group.

Chromosome segregation is driven by separase, activity of which is inhibited by binding to securin and cyclin B1/CDK1. In meiosis, premature separase activity will induce aneuploidy or abolish chromosome segregation owing to the untimely destruction of cohesin. Recently, we have proved that cyclin B2 can compensate for cyclin B1 in CDK1 activation for the oocyte meiosis G2/M transition. In the present study, we identify an interaction between cyclin B2/CDK1 and separase in mouse oocytes. We find that cyclin B2 degradation is required for separase activation during the metaphase I-anaphase I transition because the presence of stable cyclin B2 leads to failure of homologous chromosome separation and to metaphase I arrest, especially in the simultaneous absence of securin and cyclin B1. Moreover, non-phosphorylatable separase rescues the separation of homologous chromosomes in stable cyclin B2-arrested cyclin B1-null oocytes. Our results indicate that cyclin B2/CDK1 is also responsible for separase inhibition via inhibitory phosphorylation to regulate chromosome separation in oocyte meiosis, which may not occur in other cell types. © 2019. Published by The Company of Biologists Ltd.

Chromosome segregation is driven by separase, activity of which is inhibited by binding to securin and cyclin B1/CDK1. In meiosis, premature separase activity will induce aneuploidy or abolish chromosome segregation owing to the untimely destruction of cohesin. Recently, we have proved that cyclin B2 can compensate for cyclin B1 in CDK1 activation for the oocyte meiosis G2/M transition. In the present study, we identify an interaction between cyclin B2/CDK1 and separase in mouse oocytes. We find that cyclin B2 degradation is required for separase activation during the metaphase I-anaphase I transition because the presence of stable cyclin B2 leads to failure of homologous chromosome separation and to metaphase I arrest, especially in the simultaneous absence of securin and cyclin B1. Moreover, non-phosphorylatable separase rescues the separation of homologous chromosomes in stable cyclin B2-arrested cyclin B1-null oocytes. Our results indicate that cyclin B2/CDK1 is also responsible for separase inhibition via inhibitory phosphorylation to regulate chromosome separation in oocyte meiosis, which may not occur in other cell types. © 2019. Published by The Company of Biologists Ltd.

Chromosome segregation is driven by separase, activity of which is inhibited by binding to securin and cyclin B1/CDK1. In meiosis, premature separase activity will induce aneuploidy or abolish chromosome segregation owing to the untimely destruction of cohesin. Recently, we have proved that cyclin B2 can compensate for cyclin B1 in CDK1 activation for the oocyte meiosis G2/M transition. In the present study, we identify an interaction between cyclin B2/CDK1 and separase in mouse oocytes. We find that cyclin B2 degradation is required for separase activation during the metaphase I-anaphase I transition because the presence of stable cyclin B2 leads to failure of homologous chromosome separation and to metaphase I arrest, especially in the simultaneous absence of securin and cyclin B1. Moreover, non-phosphorylatable separase rescues the separation of homologous chromosomes in stable cyclin B2-arrested cyclin B1-null oocytes. Our results indicate that cyclin B2/CDK1 is also responsible for separase inhibition via inhibitory phosphorylation to regulate chromosome separation in oocyte meiosis, which may not occur in other cell types. © 2019. Published by The Company of Biologists Ltd.

The simulation of a heterogeneous and fractured coal bed is discussed in relation to its application to the analysis of methane recovery. Coal is heterogeneous and contains natural weaknesses at various scales. As methane recovery begins the pre-existing weaknesses may close, open, extend or induce new fractures due to changes in the effective stress, which may in turn alter the structure of the coal and its flow properties. Mechanical response and flow properties are also affected by natural faults and fractures in the coal. A model was developed to simulate the coal heterogeneity based on the Monte Carlo method assuming that the mechanical characteristics such as elastic modulus and strength follow a Weibull distribution. The fracture system geometry of the coal bed was generated based on Monte Carlo simulations using the probability density functions of fracture parameters formulated according to field mapping results. A virtual generating system for natural heterogeneous and fractured coal bed was then developed in Matlab 6.0. A case study is presented for a methane production well in Liaoning, China, which showed that fractures connected with the well have a much greater effect on gas flow than those unconnected with the well., The simulation of a heterogeneous and fractured coal bed is discussed in relation to its application to the analysis of methane recovery. Coal is heterogeneous and contains natural weaknesses at various scales. As methane recovery begins the pre-existing weaknesses may close, open, extend or induce new fractures due to changes in the effective stress, which may in turn alter the structure of the coal and its flow properties. Mechanical response and flow properties are also affected by natural faults and fractures in the coal. A model was developed to simulate the coal heterogeneity based on the Monte Carlo method assuming that the mechanical characteristics such as elastic modulus and strength follow a Weibull distribution. The fracture system geometry of the coal bed was generated based on Monte Carlo simulations using the probability density functions of fracture parameters formulated according to field mapping results. A virtual generating system for natural heterogeneous and fractured coal bed was then developed in Matlab 6.0. A case study is presented for a methane production well in Liaoning, China, which showed that fractures connected with the well have a much greater effect on gas flow than those unconnected with the well.