zona pellucida
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Author(s):  
Takafumi Matsumura ◽  
Taichi Noda ◽  
Yuhkoh Satouh ◽  
Akane Morohoshi ◽  
Shunsuke Yuri ◽  
...  

Fertilization occurs as the culmination of multi-step complex processes. First, mammalian spermatozoa undergo the acrosome reaction to become fusion-competent. Then, the acrosome-reacted spermatozoa penetrate the zona pellucida and adhere to and finally fuse with the egg plasma membrane. IZUMO1 is the first sperm protein proven to be essential for sperm-egg fusion in mammals, as Izumo1 knockout mouse spermatozoa adhere to but fail to fuse with the oolemma. However, the IZUMO1 function in other species remains largely unknown. Here, we generated Izumo1 knockout rats by CRISPR/Cas9 and found the male rats were infertile. Unlike in mice, Izumo1 knockout rat spermatozoa failed to bind to the oolemma. Further investigation revealed that the acrosome-intact sperm binding conceals a decreased number of the acrosome-reacted sperm bound to the oolemma in Izumo1 knockout mice. Of note, we could not see any apparent defects in the binding of the acrosome-reacted sperm to the oolemma in the mice lacking recently found fusion-indispensable genes, Fimp, Sof1, Spaca6, or Tmem95. Collectively, our data suggest that IZUMO1 is required for the sperm-oolemma binding prior to fusion at least in rat.


Author(s):  
Zhaokang Cui ◽  
Yajuan Lu ◽  
Yilong Miao ◽  
Xiaoxin Dai ◽  
Yu Zhang ◽  
...  

2022 ◽  
Vol 78 (03) ◽  
pp. 6626-2022
Author(s):  
JADWIGA JAWORSKA-ADAMU ◽  
ALEKSANDRA KRAWCZYK ◽  
KAROL RYCERZ

In mammals, oocytes, fertilized eggs and pre-implantation embryos are surrounded by an acellular zona pellucida (zona pellucida – ZP). This structure has a fibro-spongy character but it undergoes constant modifications throughout its existence depending on many internal and external factors. ZP consists of glycoproteins marked as ZP1, ZP2, ZP3 and ZP4, the presence of which is species different. ZP1 and probably ZP4 molecules stabilize the fibrillary skeleton of the zona pellucida formed of ZP2 and ZP3 protein polymers which are ligands for specific spermatozoid receptors. The oligosaccharide chains of ZP3 are responsible for the primary attachment of the male gamete which induces the acrosomal reaction. ZP2 enhances this connection by secondary binding to an acrosome-free spermatozoid. Additionally, oviductal specific glycoprotein 1 which plays a role in interspecific oocyte-sperm interactions, appears around the postovulatory oocyte surrounded by ZP. In addition, this protein modifies the resistance of ZP to the action of proteases released as a result of the cortical reaction during polyspermia block. After fertilization, ZP not only protects the egg and then the embryo until implantation, but also has an embryotrophic effect. Understanding the molecular basics of the structure and properties of ZP can significantly improve animal fertility as well as reproductive rates.


2021 ◽  
Author(s):  
◽  
Jevon Upton

<p>Developing transgenic livestock has become popular in recent years after advances in the field of genetic editing. Cattle are one of the main exports in New Zealand and are a prime target for new genetic editing tools. Applications of genetic editing in cattle can extend to increases in production, and elimination of disease genes. Due to its ease of use, CRISPR/Cas9 has become one of the most popular methods of editing genes, hence this was employed in the research. Cattle embryos in culture are very sensitive to environmental changes and for this reason, a delivery vector is necessary to deliver the genetic material as traditional transfection methods cause high rates of embryo death. The zona pellucida, a glycoprotein coat surrounding the embryo, acts as a protective agent against viral vectors, and needed to be considered in the research. This research aimed to create a novel, high titer lentivirus particle capable of transducing bovine embryos, and causing subsequent genetic modification by integration of CRISPR/Cas9 into the genome. Using fluorescent reporters, viral transduction was monitored. The research found that after optimizing transfection protocols, high-titer lentiviral particles can be produced and can infect bovine embryos. Zona pellucida removal experiments revealed over-digestion in early stage embryos, however, this was not observed in compact morulas. Removing the zona allowed for successful transduction of bovine embryos, resulting in transgenic cells expressing eGFP. While CRISPR/Cas9 experiments were in preliminary stages, these indicated eGFP knock-out in certain eGFP-HEK293T cells. Though challenges were encountered throughout the research process, solutions were explored, and it was shown that transgenic bovine embryos using lentiviral gene delivery can be produced. This indicates the high likelihood that CRISPR/Cas9 systems can be delivered this way, inducing targeted genetic modification.</p>


2021 ◽  
Author(s):  
◽  
Jevon Upton

<p>Developing transgenic livestock has become popular in recent years after advances in the field of genetic editing. Cattle are one of the main exports in New Zealand and are a prime target for new genetic editing tools. Applications of genetic editing in cattle can extend to increases in production, and elimination of disease genes. Due to its ease of use, CRISPR/Cas9 has become one of the most popular methods of editing genes, hence this was employed in the research. Cattle embryos in culture are very sensitive to environmental changes and for this reason, a delivery vector is necessary to deliver the genetic material as traditional transfection methods cause high rates of embryo death. The zona pellucida, a glycoprotein coat surrounding the embryo, acts as a protective agent against viral vectors, and needed to be considered in the research. This research aimed to create a novel, high titer lentivirus particle capable of transducing bovine embryos, and causing subsequent genetic modification by integration of CRISPR/Cas9 into the genome. Using fluorescent reporters, viral transduction was monitored. The research found that after optimizing transfection protocols, high-titer lentiviral particles can be produced and can infect bovine embryos. Zona pellucida removal experiments revealed over-digestion in early stage embryos, however, this was not observed in compact morulas. Removing the zona allowed for successful transduction of bovine embryos, resulting in transgenic cells expressing eGFP. While CRISPR/Cas9 experiments were in preliminary stages, these indicated eGFP knock-out in certain eGFP-HEK293T cells. Though challenges were encountered throughout the research process, solutions were explored, and it was shown that transgenic bovine embryos using lentiviral gene delivery can be produced. This indicates the high likelihood that CRISPR/Cas9 systems can be delivered this way, inducing targeted genetic modification.</p>


Author(s):  
Maider Bizkarguenaga ◽  
Laura Gomez‐Santos ◽  
Juan Francisco Madrid ◽  
Francisco José Sáez ◽  
Edurne Alonso

2021 ◽  
Author(s):  
Ingrid Spies ◽  
Daniel P. Drinan ◽  
Eleni L. Petrou ◽  
Rory Spurr ◽  
Carolyn Tarpey ◽  
...  
Keyword(s):  

2021 ◽  
Vol 18 (22) ◽  
pp. 489
Author(s):  
Tappadit Mitparian ◽  
Sinlapachai Senarat ◽  
Jes Kettratad ◽  
Wannee Jiraungkoorskul ◽  
Gen Kaneko ◽  
...  

Information on the reproductive biology of toadfish remains limited. In this study, we examined the structure and development of gonads in the grunting toadfish Allenbatrachus grunniens (Linnaeus, 1758) using morphological, histological and ultrastructural methods. The fish were collected from the Pranburi River Estuary, Thailand, during the annual reproductive period for this species (January to December 2018). The ovary of this species was paired and had elongated sac-like structures parallel to the kidneys and the digestive tract. In females, we identified 4 oocyte differentiation phases in the ovary including oogonia proliferation phase, a primary growth phase that was further classified into 2 steps (perinucleolar and oil droplets-cortical alveolar steps), and a secondary growth phase that also contained 2 steps (secondary growth and full-grown oocyte steps) and post-ovulatory phases, indicating an asynchronous pattern in ovarian development for this species. Transmission electron microscopy showed the 4 layers including the zona pellucida, basement membrane, granulosa cells and theca cells, all of which initially appeared in the oil droplets-cortical alveolar stage. The zona pellucida and the granulosa cells were highly developed during the secondary growth stage. The granulosa cells contained abundant smooth endoplasmic reticulum near the mitochondria. In males, the spermatogenesis was classified into spermatogonium to spermatozoa. Finally, we associated the morphological gonadal developments (stage I - IV) and the gonadosomatic index (GSI value) with the cellular developmental processes in both sexes. These results help integrate various levels of reproductive observations, which will be applied to understanding the reproductive cycle and development for aquaculture.


2021 ◽  
Vol 12 ◽  
Author(s):  
Longda Wang ◽  
Mengxiang Chen ◽  
Gaofeng Yan ◽  
Shuhua Zhao

Children conceived through intracytoplasmic sperm injection (ICSI) have been reported to have a higher risk of many abnormalities and disorders, including autism and intellectual disability, which may be due to bypassing of the natural sperm selection process during ICSI. Zona pellucida (ZP)-bound spermatozoa (ZPBS) have normal morphology and nuclear DNA. Using these spermatozoa for ICSI results in better outcomes compared with conventional ICSI. However, differences besides morphology that exist between sperm selected by ZP and by an embryologist and whether these differences affect the risk of autism in offspring after ICSI are unclear. To explore these questions, we compared genome-wide DNA methylation profiles between ZPBS and manually selected spermatozoa (MSS)using single-cell bisulfite sequencing. Global DNA methylation levels were significantly lower in ZPBS than in MSS. Using gene ontology (GO) analysis, genes overlapping differentially methylated regions (DMRs) were enriched in biological processes involving neurogenesis. Furthermore, we found that 47.8% of autism candidate genes were associated with DMRs, compared with 37.1% of matched background genes (P&lt;0.001). This was mainly because of the high proportion of autism candidate genes with bivalent chromatin structure. In conclusion, bivalent chromatin structure results in large differences in the methylation of autism genes between MSS and ZPBS. ICSI using MSS, which increases the risk of methylation mutations compared with ZPBS, may lead to a higher risk of autism in offspring.


Author(s):  
Kamila Pulawska ◽  
Donata Ponikwicka-Tyszko ◽  
Weronika Lebiedzinska ◽  
Peilan Guo ◽  
Piotr Bernaczyk ◽  
...  

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