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Autophagy Apr 2024Despite the well-described discrepancy between (macroautophagy/autophagy-related) genes in the regulation of hematopoiesis, varying essentiality of core ATG proteins in...
Despite the well-described discrepancy between (macroautophagy/autophagy-related) genes in the regulation of hematopoiesis, varying essentiality of core ATG proteins in vertebrate definitive hematopoiesis remains largely unclear. Here, we employed zebrafish () to compare the functions of six core genes, including , (beclin1), , , , and , in vertebrate definitive hematopoiesis via clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 ribonucleoprotein and morpholino targeting. Zebrafish with various mutations showed autophagic deficiency and presented partially consistent hematopoietic abnormalities during early development. All six mutations led to a declined number of (Spi-1 proto-oncogene b) myeloid progenitor cells. However, only mutation resulted in the expansion of (v-myb avian myeloblastosis viral oncogene homolog) hematopoietic stem and progenitor cells (HSPCs) and transiently increased (coronin, actin binding protein, 1A) leukocytes, whereas mutation decreased the number of HSPCs and leukocytes. Proteomic analysis of caudal hematopoietic tissue identified (SIN3 transcription regulator family member Aa) as a potential modulator of - and -regulated definitive hematopoiesis. Disruption of rescued the expansion of HSPCs and leukocytes in mutants and exacerbated the decrease of HSPCs in mutants. Double mutations were also performed to examine alternative functions of various genes in definitive hematopoiesis. Notably, mutation failed to induce HSPCs expansion with one of the other five mutations. These findings demonstrated the distinct roles of genes and their interplays in zebrafish definitive hematopoiesis, thereby suggesting that the vertebrate definitive hematopoiesis is regulated in an gene-dependent manner.: AGM: aorta-gonad-mesonephros; AO: acridine orange; : autophagy related; : beclin 1, autophagy related; CHT: caudal hematopoietic tissue; CKO: conditional knockout; : coronin, actin binding protein, 1A; CQ: chloroquine; CRISPR: clustered regularly interspaced short palindromic repeats; dpf: days post fertilization; FACS: fluorescence-activated cell sorting; : hemoglobin, alpha embryonic 1.1; HSCs: hematopoietic stem cells; HSPCs: hematopoietic stem and progenitor cells; KD: knockdown; KO: knockout; : microtubule-associated protein 1 light chain 3; MO: morpholino; : macrophage expressed 1, tandem duplicate 1; : myeloid-specific peroxidase; : v-myb avian myeloblastosis viral oncogene homolog; PE: phosphatidylethanolamine; : phospho-H3 histone; PtdIns3K: class 3 phosphatidylinositol 3-kinase; : recombination activating 1; : RB1-inducible coiled-coil 1; RFLP: restriction fragment length polymorphism; RNP: ribonucleoprotein; : SIN3 transcription regulator family member Aa; : Spi-1 proto-oncogene b; : unc-51 like autophagy activating kinase; : vitellogenin 1; WISH: whole-mount in situ hybridization.
Topics: Animals; Zebrafish; Hematopoiesis; Autophagy; Zebrafish Proteins; Mutation; Hematopoietic Stem Cells; Autophagy-Related Proteins; Proto-Oncogene Mas
PubMed: 37921505
DOI: 10.1080/15548627.2023.2274251 -
International Archives of Allergy and... 2024German cockroach (GCr) aeroallergens are associated with allergic rhinitis and asthma. Vitellogenin (Vg) and vitellin (Vn) are abundant proteins in GCr blood and eggs...
INTRODUCTION
German cockroach (GCr) aeroallergens are associated with allergic rhinitis and asthma. Vitellogenin (Vg) and vitellin (Vn) are abundant proteins in GCr blood and eggs (including egg cases), respectively, and are possible high molecular mass allergens. Prior efforts to purify Vg/Vn yielded amounts too small for subsequent studies. In this study, we report the affinity purification of Vg/Vn from whole-body defatted GCr powder and determination of the binding of Vg/Vn to anti-GCr IgE.
METHOD
New Zealand white rabbits were immunized with pure Vg/Vn in Freund's adjuvant, and IgG was purified from the rabbit sera and conjugated to cyanogen bromide (CNBr)-activated Sepharose. Aqueous extracts from GCr powder were passed over the column. After extensive washing, putative Vg/Vn was eluted in low-pH buffer, neutralized, and analyzed by SDS-PAGE and liquid chromatography high-resolution mass spectrometry (LC-HRMS). IgE binding of Vg/Vn was evaluated by inhibition of IgE binding to GCr-ImmunoCAP(I6) in sera from 10 GCr-allergic individuals. In addition, Vg/Vn was biotinylated and bound to ImmunoCAP-streptavidin, and direct IgE antibody binding to the immobilized Vg/Vn was determined in sera from 26 GCr-allergic individuals.
RESULTS
Vg/Vn isolated by affinity chromatography was 91% pure by LC-HRMS; contaminants included Bla g 3 (0.9%), human keratin (6%), and rabbit IgG. Vg/Vn inhibited IgE binding to GCr-ImmunoCAP(I6) in 8 of 10 sera. In direct-binding experiments, 21/26 (80%) sera had anti-Vg/Vn IgE at >0.10 kUA/L, while 11/26 (42%) sera were >0.35 kUA/L.
CONCLUSIONS
We affinity-purified Vg/Vn and demonstrated that Vg/Vn-specific IgE antibody is a major component of GCr-specific IgE.
Topics: Animals; Allergens; Immunoglobulin E; Rabbits; Humans; Vitellogenins; Blattellidae; Male; Female; Adult; Immunoglobulin G; Child
PubMed: 38253039
DOI: 10.1159/000535935 -
Biomedicine & Pharmacotherapy =... Oct 2023Fructus psoraleae (FP) is a commonly used herb with potential reproductive toxicity. Bavachin (BV), one of essential active ingredients of FP, was found to exhibit...
Fructus psoraleae (FP) is a commonly used herb with potential reproductive toxicity. Bavachin (BV), one of essential active ingredients of FP, was found to exhibit estrogenic activity, but its effect on female reproductive system remains unknown. In this study, the impact of BV on the female zebrafish reproductive system and underlying molecular mechanism were determined in vivo and ex vivo. The results showed that BV could accumulate in zebrafish ovary, leading to obvious follicular atresia and increase in gonadal index and vitellogenin content. Endoplasmic reticulum (ER) swelling and hypertrophy were observed in the BV-treated zebrafish ovary, accompanied by an increase in the expressions of ER stress and unfolded protein response (UPR) related genes, namely atf6, ire-1α and xbp1s. In the ex vivo study, BV was found to decrease the survival rate and maturation rate of oocytes, while increasing the expression of Ca. Additionally, BV led to an elevation in the level of estrogen receptor ESR1 and the expressions of genes involved in ER stress and UPR, including atf6, ire-1α, xbp1s, chop and perk. Moreover, molecular docking revealed that BV could directly bind to immunoglobulin heavy chain binding protein (BiP) and estrogen receptor 1 (ESR1). Besides, the alterations induced by BV could be partially reversed by fulvestrant (FULV) and 4-phenylbutyric acid (4-PBA), respectively. Thus, long-termed BV-containing medicine treatment could generate reproductive toxicity in female zebrafish by causing follicular atresia through BiP- and ESR-mediated ER stress and UPR, providing a potential target for the prevention of reproductive toxicity caused by BV.
Topics: Female; Animals; Zebrafish; Ovary; Follicular Atresia; Molecular Docking Simulation; Signal Transduction; Endoplasmic Reticulum Stress; Unfolded Protein Response; Apoptosis
PubMed: 37586115
DOI: 10.1016/j.biopha.2023.115322 -
Frontiers in Endocrinology 2023Within insects, corticotropin-releasing factor/diuretic hormones (CRF/DHs) are responsible for the modulation of a range of physiological and behavioural processes such...
Within insects, corticotropin-releasing factor/diuretic hormones (CRF/DHs) are responsible for the modulation of a range of physiological and behavioural processes such as feeding, diuresis, and reproduction. Rhopr-CRF/DH plays a key role in feeding and diuresis in , a blood-gorging insect and a vector for human Chagas disease. Here, we extend our understanding on the role of this neurohormone in reproduction in adult female . Double-label immunohistochemistry displays co-localized staining of CRF-like and the glycoprotein hormone (GPA2/GPB5) subunit GPB5-like immunoreactivity in the same neurosecretory cells (NSCs) in the mesothoracic ganglionic mass (MTGM) and in their neurohemal sites in adult female , suggesting these peptides could work together to regulate physiological processes. qPCR analysis reveals that the transcript for Rhopr-CRF/DH receptor 2 () is expressed in reproductive tissues and fat body (FB) in adult female , and its expression increases post blood meal (PBM), a stimulus that triggers diuresis and reproduction. Using RNA interference, transcript expression of was knocked down, and egg production monitored by examining the major yolk protein, vitellogenin (Vg), the number and quality of eggs laid, and their hatching ratio. Injection of dsCRFR2 into adult females reduces transcript expression, accelerates oogenesis, increases the number of eggs produced, and reduces hatching rate in female . Downregulation of leads to an increase in the transcript expression of in the fat body and ovaries, and increases the transcript level for the Vg receptor, , in the ovaries. A significant increase in Vg content in the fat body and in the hemolymph is also observed. Incubation of isolated tissues with Rhopr-CRF/DH leads to a significant decrease in transcript expression of in the fat body and in the ovaries. In addition, Rhopr-CRF/DH reduces transcript expression of the ecdysteroid biosynthetic enzymes and reduces ecdysteroid titer in the culture medium containing isolated ovaries. These results suggest the involvement of the CRF-signaling pathway in reproduction, and that Rhopr-CRF/DH acts as a gonad-inhibiting hormone in the adult female , as previously shown for the colocalized glycoprotein, GPA2/GPB5.
Topics: Animals; Female; Adult; Humans; Corticotropin-Releasing Hormone; Diuretics; Rhodnius; Ecdysteroids; Gonadal Hormones; Glycoproteins; Gonads
PubMed: 37842303
DOI: 10.3389/fendo.2023.1279929 -
Journal of Insect Physiology Dec 2023Effects of dietary protein quality on insect development (not just growth) are unclear. Dietary amino acid blends matching yolk proteins support reproduction and...
Effects of dietary protein quality on insect development (not just growth) are unclear. Dietary amino acid blends matching yolk proteins support reproduction and juvenile development in Drosophila melanogaster. We matched amino acids to vitellogenin and tested development of juvenile male lubber grasshoppers, which do not produce vitellogenin. Last instars were fed classic dry diets with amino acids substituted for proteins. Matching amino acids to vitellogenin allowed molting to adulthood, while an unmatched isonitrogenous diet did not. Health on dry diets was poor, so we developed wet diets with agar, horse feed, and amino acids. Juveniles fed these diets matched to vitellogenin developed comparably to juveniles fed lettuce. However, wet diets with amino acids dissimilar to vitellogenin (low-quality) slowed development but maintained size at adulthood. We observed no compensatory feeding on low-quality diets. Theory suggests accumulation of proteins permits development. To detect a threshold, we started last juvenile instars on high-quality diets, then abruptly switched them to low-qualities diets. When switched to the poor-quality diet at 6d, grasshoppers molted at a similar age (∼17d) to grasshoppers continuously on the high-quality diet. Total hemolymph proteins levels were unaffected by the timing of diet switches. Last, methionine is essential but can be noxious at high levels. Diets with low-quality protein except for methionine slowed growth early but did not alter the time or size at molt. Overall, the feeding threshold is solely due to essential amino acids, and low-quality protein diets slowed development but did not affect adult size.
Topics: Male; Animals; Horses; Vitellogenins; Drosophila melanogaster; Grasshoppers; Amino Acids; Methionine; Diet; Embryonic Development; Animal Feed; Dietary Proteins
PubMed: 37989476
DOI: 10.1016/j.jinsphys.2023.104586 -
General and Comparative Endocrinology Feb 2024The Pacific halibut (Hippoglossus stenolepis) is a large migratory demersal flatfish species that occupies a top trophic role in the North Pacific Ocean and Bering Sea...
The Pacific halibut (Hippoglossus stenolepis) is a large migratory demersal flatfish species that occupies a top trophic role in the North Pacific Ocean and Bering Sea ecosystems, where it also supports various fisheries. As a first attempt to characterize the endocrine mechanisms driving sexual maturation in this important species, we collected pituitary, ovarian and blood samples from Pacific halibut females captured in the wild that were classified histologically into various female developmental stages. We conducted gene expression analyses of gonadotropin beta subunits in the pituitary and observed that mRNA expression levels of fshb gradually increased throughout vitellogenesis, remained elevated until before ovulation and declined after spawning. In contrast, the mRNA expression levels of lhb markedly increased during oocyte maturation and remained elevated until after spawning. Ovarian mRNA expression levels of the gonadotropin receptor genes fshr and lhr peaked during oocyte maturation and before spawning, respectively, immediately following the developmental stage at which pituitary fshb and lhb mRNA expression first reached maximum levels. The ovarian gene expression patterns of steroidogenic enzyme genes cyp19a1 and hsd20b2 paralleled those of fshr and lhr, respectively. Testosterone and 17β-estradiol (E) plasma levels increased concomitantly with fshr and cyp19a1 mRNA expression levels, and vitellogenin plasma levels increased throughout vitellogenesis and reached maximum levels prior to spawning. These results are consistent with the notion that in female Pacific halibut, as in other teleosts, vitellogenesis and oocyte maturation and ovulation are likely under the control of pituitary gonadotropic hormones Fsh and Lh, respectively.
Topics: Animals; Female; Flounder; Ecosystem; Gonadotropins, Pituitary; Gonadotropins; RNA, Messenger
PubMed: 38101488
DOI: 10.1016/j.ygcen.2023.114425 -
Veterinary Parasitology Jul 2024Tick infestations transmit various infectious agents and result in significant socioeconomic consequences. Currently, the primary focus of tick control efforts is...
Tick infestations transmit various infectious agents and result in significant socioeconomic consequences. Currently, the primary focus of tick control efforts is identifying potential targets for immune intervention. In a previous study, we identified a highly conserved protein abundant in tick haemolymph extracellular vesicles (EVs) known as translationally controlled tumour protein (TCTP). We have found that native TCTP is present in various tissues of the Rhipicephalus haemaphysaloides tick, including salivary glands, midgut, ovary, and fat body. Notably, TCTP is particularly abundant in the tick ovary and its levels increase progressively from the blood-feeding stage to engorgement. When the TCTP gene was knocked down by RNAi, there was a noticeable delay in ovarian development, and the reproductive performance, in terms of egg quantity and survival, was also hindered. Our investigations have revealed that the observed effects in ovary and eggs in dsRNA-treated ticks are not attributable to cell death mechanisms like apoptosis and autophagy but rather to the reduction in the expression of vitellogenin (Vg1, Vg2, and Vg3) and ferritin (ferritin 1 and ferritin 2) proteins crucial for ovarian development and embryo survival in ticks. Additionally, phylogenetic analysis and structural comparisons of RhTCTP and its orthologues across various tick species, vertebrate hosts, and humans have shown that TCTP is conserved in ticks but differs significantly between ticks and their hosts, particularly in the TCTP_1 and TCTP_2 domains. Overall, TCTP plays a vital role in tick reproductive development and presents itself as a potential target for tick control in both humans and animals.
Topics: Animals; Female; Rhipicephalus; Tumor Protein, Translationally-Controlled 1; Ovary; Oviposition; Phylogeny; Vitellogenins; Biomarkers, Tumor
PubMed: 38781831
DOI: 10.1016/j.vetpar.2024.110212 -
Integrative Organismal Biology (Oxford,... 2024For many fishes, reproductive function is thermally constrained such that exposure to temperatures above some upper threshold has detrimental effects on gametic...
For many fishes, reproductive function is thermally constrained such that exposure to temperatures above some upper threshold has detrimental effects on gametic development and maturation, spawning frequency, and mating behavior. Such impairment of reproductive performance at elevated temperatures involves changes to hypothalamic-pituitary-gonadal (HPG) axis signaling and diminished gonadal steroidogenesis. However, how HPG pathways respond to consistently high versus temporally elevated temperatures is not clear. Here, sexually mature Amargosa River Pupfish () were maintained under thermal regimes of either stable ∼25°C (low temperature), diurnal cycling temperatures between ∼27 and 35°C (fluctuating temperature), or stable ∼35°C (high temperature) conditions for 50 days to examine effects of these conditions on HPG endocrine signaling components in the pituitary gland and gonad, ovarian and testicular gametogenesis status, and liver gene expression relating to oogenesis. Female pupfish maintained under stable high and fluctuating temperature treatments showed reduced gonadosomatic index values as well as a lower proportion of oocytes in the lipid droplet and vitellogenic stages. Females in both fluctuating and stable 35°C conditions exhibited reduced ovarian mRNAs for steroid acute regulatory protein (), cholesterol side chain-cleavage enzyme, P450scc (), and 3β-hydroxysteroid dehydrogenase (), while ovarian transcripts encoding 11β-hydroxysteroid dehydrogenase () and sex hormone-binding globulin () were elevated in females at constant 35°C only. Ovarian aromatase () mRNA levels were unaffected, but circulating 17β-estradiol (E) was lower in females at 35°C compared to the fluctuating temperature condition. In the liver, mRNA levels for choriogenins and vitellogenin were downregulated in both the fluctuating and 35°C conditions, while hepatic estrogen receptor 2a () and mRNAs were elevated in 35°C females. Taken together, these results demonstrate the potential for elevated temperatures to impair ovarian steroidogenesis and reduce egg envelope and vitellogenin protein production in female pupfish, while also shedding light on how thermal regimes that only intermittently reach the upper thermal range for reproduction have differential impacts on reproductive endocrine pathways than constantly warm conditions.
PubMed: 38464886
DOI: 10.1093/iob/obae003 -
International Journal of Molecular... Dec 2023The red imported fire ant ( Buren) is a social pest species with a robust reproductive ability that causes extensive damage. Identification of the genes involved in...
The red imported fire ant ( Buren) is a social pest species with a robust reproductive ability that causes extensive damage. Identification of the genes involved in queen fertility is critical in order to better understand the reproductive biology and screening for the potential molecular targets in . Here, we used the mRNA deep sequencing (RNA-seq) approach to identify differentially expressed genes (DEGs) in the transcriptomes of three reproductive caste types of , including queen (QA) and winged female (FA) and male (MA) ants. The genes that were specific to and highly expressed in the queens were then screened, and the and genes were chosen as targets to explore their functions in oogenesis and fertility. A minimum of 6.08 giga bases (Gb) of clean reads was obtained from all samples, with a mapping rate > 89.78%. There were 7524, 7133, and 977 DEGs identified in the MA vs. QA, MA vs. FA, and FA vs. QA comparisons, respectively. qRT-PCR was used to validate 10 randomly selected DEGs, including () and (), and their expression patterns were mostly consistent with the RNA-seq data. The Vgs included conserved domains and motifs that are commonly found in most insect Vgs. and were highly expressed in queens and winged females and were most highly expressed in the thorax, followed by the fat body, head, and epidermis. Evaluation based on a loss-of-function-based knockdown analysis showed that the downregulation of either or both of these genes resulted in smaller ovaries, less oogenesis, and less egg production. The results of transcriptional sequencing provide a foundation for clarifying the regulators of queen fertility in . The functions of and as regulators of oogenesis highlight their importance in queen fecundity and their potential as targets of reproductive disruption in control.
Topics: Animals; Female; Male; Vitellogenins; Fire Ants; Reproduction; Fertility; Ants
PubMed: 38138959
DOI: 10.3390/ijms242417130 -
The Science of the Total Environment Dec 2023Microplastics could act as vectors for the transport of harmful bacteria, such as pathogens and antibiotic resistance bacteria (ARB), but their combined effects have not...
Microplastics could act as vectors for the transport of harmful bacteria, such as pathogens and antibiotic resistance bacteria (ARB), but their combined effects have not been reported yet. Here, ARB Shigella flexneri with sulfonamides resistance and micro-polystyrene (micro-PS) were used to investigate their possible combined effects on the growth and expression of functional genes in Daphnia magna. Results showed that micro-PS colonized with S. flexneri were ingested by D. magna and blocked in their intestine after 24 h exposure. Changes were observed in the life history and morphology of D. magna, as well as the expression of functional genes in all treatments, but with no difference in the survival rate. We also determined the expression of six functional genes involved in energy and metabolism (arginine kinase, AK) and oxidative stress response (thioredoxin reductase, TRxR, catalase, CAT, and glutathione S-transferases, GSTs), as well as in growth, development and reproduction (vitellogenin, Vtg1 and ecdysone receptor, EcR). AK and Vtg1 did not show significant differences, however, EcR was down-regulated and the other three genes (TRxR, CAT, GSTs) were up-regulated in the combined-treated group. Antibiotic resistance gene (ARGs) sul1 was detected when exposed to micro-PS colonized with S. flexneri., suggesting that D. magna could acquire resistance genes through microplastic biofilms. These results indicated that MPs could act as a carrier of ARB to transfer ARGs into D. magna, and affect the life history, morphology, and the expression of related functional genes of D. magna, to adapt to the stress caused by MPs and ARB.
Topics: Animals; Microplastics; Plastics; Anti-Bacterial Agents; Daphnia; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Polystyrenes; Bacteria; Water Pollutants, Chemical
PubMed: 37709097
DOI: 10.1016/j.scitotenv.2023.166880