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Development (Cambridge, England) Oct 2021In animals, early development is dependent on a pool of maternal factors, both RNA and proteins, which are required for basic cellular processes and cell differentiation...
In animals, early development is dependent on a pool of maternal factors, both RNA and proteins, which are required for basic cellular processes and cell differentiation until zygotic genome activation. The role of the majority of these maternally expressed factors is not fully understood. By exploiting the biallelic editing ability of CRISPR-Cas9, we identify and characterize maternal-effect genes in a single generation, using a maternal crispant technique. We validated the ability to generate biallelic mutations in the germ line by creating maternal crispants that phenocopied previously characterized maternal-effect genes: birc5b, tmi and mid1ip1. Additionally, by targeting maternally expressed genes of unknown function in zebrafish, we identified two maternal-effect zebrafish genes, kpna7 and fhdc3. The genetic identity of these maternal crispants was confirmed by sequencing haploid progeny from F0 females, which allowed the analysis of newly induced lesions in the maternal germ line. Our studies show that maternal crispants allow for the effective identification and primary characterization of maternal-effect genes in a single generation, facilitating the reverse genetics analysis of maternal factors that drive embryonic development.
Topics: Animals; CRISPR-Cas Systems; Gene Editing; Maternal Inheritance; Ovum; Zebrafish; Zebrafish Proteins
PubMed: 34463742
DOI: 10.1242/dev.199536 -
Biochemical and Biophysical Research... Dec 2019
Review
Topics: Animals; Biological Transport; Cell Membrane; Cell Nucleus; Fertilization; Membrane Potentials; Models, Biological; Ovum; Receptors, Cell Surface; Sea Urchins; Starfish; Zygote
PubMed: 31761083
DOI: 10.1016/j.bbrc.2019.10.024 -
Molecular Human Reproduction Dec 2014Proteins expressed on the surface of sperm and egg mediate gametic compatibility and these proteins can be subject to intense positive selection. In this review, we... (Review)
Review
Proteins expressed on the surface of sperm and egg mediate gametic compatibility and these proteins can be subject to intense positive selection. In this review, we discuss what is known about the patterns of adaptive evolution of gamete recognition proteins (GRPs). We focus on species that broadcast eggs and sperm into the environment for external fertilization, as the ease of observing and manipulating gamete interactions has allowed for greater advances in the understanding of GRP evolution, uncomplicated by confounding behavioral and physiological components that offer alternative evolutionary targets in internal fertilizers. We discuss whether interspecific mechanisms, such as selection to avoid fertilization between species (reinforcement selection), or intraspecific mechanisms, such as selection to increase (or decrease) the affinity between eggs and sperm based on the intensity of sperm competition, may be responsible for the pattern of GRP evolution observed. Variation in these proteins appears to influence gametic compatibility; GRP divergence among species is a better predictor of hybrid fertilization than neutral genetic markers and GRP variation within species predicts reproductive success among individuals within a population. Evidence suggests that sperm competition may play a large role in the evolution of gametic compatibility.
Topics: Animals; Biological Evolution; Evolution, Molecular; Female; Fertilization; Male; Ovum; Reproduction; Selection, Genetic; Sexual Behavior, Animal; Sperm-Ovum Interactions; Spermatozoa
PubMed: 25323969
DOI: 10.1093/molehr/gau069 -
Philosophical Transactions of the Royal... Oct 2020Dynamic rearrangements of epithelial cells play central roles in shaping tissues and organs during development. There are also scenarios, however, in which epithelial... (Review)
Review
Dynamic rearrangements of epithelial cells play central roles in shaping tissues and organs during development. There are also scenarios, however, in which epithelial cell movements synergize with the secretion of extracellular matrix to build rigid, acellular structures that persist long after the cells are gone. The formation of the micropyle provides an elegant example of this epithelial craftsmanship. The micropyle is a cone-shaped projection of the eggshell through which the sperm will enter to fertilize the oocyte. Though simple on the surface, both the inner structure and construction of the micropyle are remarkably complex. In this review, I first provide an overview of egg development, focusing on the key events required to understand micropyle formation. I then describe the structure of the micropyle, the cellular contributions to its morphogenesis and some interesting open questions about this process. There is a brief discussion of micropyle formation in other insects and fish to highlight the potential for comparative studies. Finally, I discuss how new studies of micropyle formation could reveal general mechanisms that epithelia use to build complex extracellular structures. This article is part of a discussion meeting issue 'Contemporary morphogenesis'.
Topics: Animals; Drosophila melanogaster; Embryo, Nonmammalian; Epithelium; Fertilization; Morphogenesis; Ovum
PubMed: 32829690
DOI: 10.1098/rstb.2019.0561 -
Cold Spring Harbor Protocols Aug 2018Most drug screening methods use purified proteins, cultured cells, and/or small model organisms such as , zebrafish, flies, or nematodes. These systems have proven...
Most drug screening methods use purified proteins, cultured cells, and/or small model organisms such as , zebrafish, flies, or nematodes. These systems have proven successes in drug discovery, but they also have weaknesses. Although purified cellular components allow for identification of compounds with activity against specific targets, such systems lack the complex biological interactions present in cellular and organismal screens. In vivo systems overcome these weaknesses, but the lack of cellular permeability, efflux by cellular pumps, and/or toxicity can be major limitations. egg extract, a concentrated and biologically active cytosol, can potentially overcome these weaknesses. Drug interactions occur in a near-physiological milieu, thereby functioning in a "truer" endogenous manner than purified components. Also, egg extract is a cell-free system that lacks intact plasma membranes that could restrict drug access to potential targets. Finally, egg extract is readily manipulated at the protein level: Proteins are easily depleted or added to the system, an important feature for analyzing drug effects in disease states. Thus, egg extract offers an attractive media for screening drugs that merges strengths of both in vitro and in vivo systems.
Topics: Animals; Cell-Free System; Drug Evaluation, Preclinical; Ovum; Xenopus laevis
PubMed: 29475996
DOI: 10.1101/pdb.prot098277 -
Astrobiology Jan 2017Tardigrades are microscopic organisms renowned for their ability to survive extreme environmental conditions. Tardigrade extreme-tolerance research has centered on the...
Tardigrades are microscopic organisms renowned for their ability to survive extreme environmental conditions. Tardigrade extreme-tolerance research has centered on the ability to withstand desiccation, low and high temperatures, and high hydrostatic pressure and radiation levels. Tardigrade tolerance to hypergravity, however, has yet to be described. We used the eutardigrade species Hypsibius dujardini to investigate short-term tolerance to g-equivalent accelerations (i.e., mimicking g-forces). Data obtained from specimens centrifuged between 3421g and 16,060g for 1 min inclusively reveal tolerance in an acceleration-dependent relation, with lower survivorship and egg production at higher accelerations. This is the first study to demonstrate tardigrade potential for tolerance to hypergravity and describe expected effects on tardigrade survival and reproduction. These findings will prove to be useful in lithopanspermia research (i.e., viable spread in meteoritic rocks). Key Words: Astrobiology-Extreme tolerance-Hypergravity-Tardigrade. Astrobiology 17, 55-60.
Topics: Acceleration; Animals; Centrifugation; Gravitation; Ovum; Tardigrada
PubMed: 28051326
DOI: 10.1089/ast.2015.1439 -
Gene Feb 2021Balbaini body (Bb) plays a vital role in germ plasm (GP) assembly and dorsoventral pattern, which is of critical important in germline specification and development....
Balbaini body (Bb) plays a vital role in germ plasm (GP) assembly and dorsoventral pattern, which is of critical important in germline specification and development. Bucky ball (buc) is reported to be essential for boosting primordial germ cell (PGC) through Bb in previous research. In the present study, a buc homolog (Olbuc) was identified in medaka (Oryzias latipes), and the roles of Olbuc on PGC development were further elucidated. The full length of Olbuc was 2148 bp, which contains a 1724 bp CDS (Coding sequence), a 167 bp 5' UTR (Untranslated region), and a 257 bp 3' UTR. By RT-PCR, the Olbuc RNA expression was maternally provided during embryogenesis and was restricted in the ovary of adult tissues. By in situ hybridization, Olbuc RNA was abundant in oocyte of meiotic stage, but gradually decreased as the oogenesis proceeded. Surprisingly, Olbuc was not co-localized with dazl, the marker gene of Bb. Interestingly, GFP can be specifically and stably expressed through the induction of Olbuc 3'UTR in PGCs. Furthermore, overexpression of Olbuc mRNA could increase PGC number and generate ectopic PGC in medaka and zebrafish embryos. In summary, our results showed that Olbuc performs a conserved function in PGC development in medaka.
Topics: 3' Untranslated Regions; Animals; Body Patterning; Embryo, Nonmammalian; Female; Fish Proteins; Gene Expression Regulation, Developmental; Maternal Inheritance; Oryzias; Ovum
PubMed: 33221537
DOI: 10.1016/j.gene.2020.145317 -
PloS One 2019Intestinal mucosa is the interface between the microbial content of the gut and the host's milieu. The goal of this study was to modulate chicken intestinal microflora...
Intestinal mucosa is the interface between the microbial content of the gut and the host's milieu. The goal of this study was to modulate chicken intestinal microflora by in ovo stimulation with galactooligosaccharides (GOS) prebiotic and to demonstrate the molecular responses of the host. The animal trial was performed on meat-type chickens (Ross 308). GOS was delivered by in ovo injection performed into the air cell on day 12 of egg incubation. Analysis of microbial communities and mucosal gene expression was performed at slaughter (day 42 post-hatching). Chyme (for DNA isolation) and intestinal mucosa (for RNA isolation) from four distinct intestinal segments (duodenum, jejunum, ileum, and caecum) was sampled. The relative abundance of Bifidobacterium spp. and Lactobacillus spp. in DNA isolated from chyme samples was determined using qPCR. On the host side, the mRNA expression of 13 genes grouped into two panels was analysed with RT-qPCR. Panel (1) included genes related to intestinal innate immune responses (IL-1β, IL-10 and IL-12p40, AvBD1 and CATHL2). Panel (2) contained genes involved in intestinal barrier function (MUC6, CLDN1 and TJAP1) and nutrients sensing (FFAR2 and FFAR4, GLUT1, GLUT2 and GLUT5). GOS increased the relative abundance of Bifidobacterium in caecum (from 1.3% to 3.9%). Distinct effects of GOS on gene expression were manifested in jejunum and caecum. Cytokine genes (IL-1β, IL-10 and IL-12p40) were up-regulated in the jejunum and caecum of the GOS-treated group. Host defence peptides (AvBD1 and CATHL2) were up-regulated in the caecum of the GOS-treated group. Free fatty acid receptors (FFAR2 and FFAR4) were up-regulated in all three compartments of the intestine (except the duodenum). Glucose transporters were down-regulated in duodenum (GLUT2 and GLUT5) but up-regulated in the hindgut (GLUT1 and GLUT2). In conclusion, GOS delivered in ovo had a bifidogenic effect in adult chickens. It also modulated gene expression related to intestinal immune responses, gut barrier function, and nutrient sensing.
Topics: Animals; Chickens; Female; Galactose; Gastrointestinal Microbiome; Gene Expression Regulation; Intestinal Mucosa; Intestines; Oligosaccharides; Ovum; Probiotics
PubMed: 30811518
DOI: 10.1371/journal.pone.0212318 -
Frontiers in Immunology 2020Gastropod Molluscs rely exclusively on the innate immune system to protect from pathogens, defending their embryos through maternally transferred effectors. In this...
Gastropod Molluscs rely exclusively on the innate immune system to protect from pathogens, defending their embryos through maternally transferred effectors. In this regard, snail eggs, in addition to immune defenses, have evolved the perivitellin-2 or PV2 combining two immune proteins into a neurotoxin: a lectin and a pore-forming protein from the Membrane Attack Complex/Perforin (MACPF) family. This binary structure resembles AB-toxins, a group of toxins otherwise restricted to bacteria and plants. Many of these are enterotoxins, leading us to explore this activity in PV2. Enterotoxins found in bacteria and plants act mainly as pore-forming toxins and toxic lectins, respectively. In animals, although both pore-forming proteins and lectins are ubiquitous, no enterotoxins have been reported. Considering that snail eggs ingestion induce morpho-physiological changes in the intestinal mucosa of rodents and is cytotoxic to intestinal cells in culture, we seek for the factor causing these effects and identified PmPV2 from eggs. We characterized the enterotoxic activity of PmPV2 through and assays. We determined that it withstands the gastrointestinal environment and resisted a wide pH range and enzymatic proteolysis. After binding to Caco-2 cells it promoted changes in surface morphology and an increase in membrane roughness. It was also cytotoxic to both epithelial and immune cells from the digestive system of mammals. It induced enterocyte death by a lytic mechanism and disrupted enterocyte monolayers in a dose-dependent manner. Further, after oral administration to mice PmPV2 attached to enterocytes and induced large dose-dependent morphological changes on their small intestine mucosa, reducing the absorptive surface. Additionally, PmPV2 was detected in the Peyer's patches where it activated lymphoid follicles and triggered apoptosis. We also provide evidence that the toxin can traverse the intestinal barrier and induce oral adaptive immunity with evidence of circulating antibody response. As a whole, these results indicate that PmPV2 is a true enterotoxin, a role that has never been reported to lectins or perforin in animals. This extends by convergent evolution the presence of plant- and bacteria-like enterotoxins to animals, thus expanding the diversity of functions of MACPF proteins in nature.
Topics: Animals; Complement Membrane Attack Complex; Enterotoxins; Immunity, Innate; Intestinal Mucosa; Mice; Mollusk Venoms; Ovum; Perforin; Snails
PubMed: 32231667
DOI: 10.3389/fimmu.2020.00428 -
Poultry Science Jul 2023The aim of the study was to assess the influence of shell defects on the quality of eggs after storage. The study material consisted of 1,800 brown-shelled eggs from...
The aim of the study was to assess the influence of shell defects on the quality of eggs after storage. The study material consisted of 1,800 brown-shelled eggs from cage rearing system which were candled on the day of laying to determine the shell quality. Eggs with the 6 most common shell defects (external crack, severe stripe marks, points, wrinkled, pimples, sandy) and eggs without defects (control group) were then stored for 35 days at 14°C and 70% humidity. The weight loss of eggs was monitored every 7 days, and the quality characteristics of whole eggs (weight, specific gravity, shape), shell (defects, strength, color, weight, thickness, density), albumen (weight, height, pH) and yolk (weight, color, pH) of 30 eggs from each group were analysed at the beginning (0 days) and after 28 and 35 days of storage. The changes resulting from water loss (air cell depth, weight loss, shell permeability) were also evaluated. The study showed that all investigated shell defects significantly influenced the characteristics of the whole egg during the storage, modifying traits such as specific gravity, water loss, shell permeability, albumen height and pH, as well as proportion, index and pH of the yolk. Furthermore, an interaction between time and the shell defects presence was found.
Topics: Animals; Egg Shell; Chickens; Ovum; Albumins; Weight Loss; Eggs; Egg Yolk
PubMed: 37220686
DOI: 10.1016/j.psj.2023.102749