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Developmental Dynamics : An Official... Oct 2022MLLT3 (AF9) is a nuclear transcription factor crucial for hematopoietic stem cell and progenitor cell maintenance, but its role during embryonic hematopoiesis remains...
BACKGROUND
MLLT3 (AF9) is a nuclear transcription factor crucial for hematopoietic stem cell and progenitor cell maintenance, but its role during embryonic hematopoiesis remains uncertain. Here, we examine the role of mllt3 in developmental hematopoiesis during embryogenesis using zebrafish.
RESULTS
Cloning, sequencing, phylogenetic, and synteny analyses showed high evolutionary conservation between important functional domains of the zebrafish orthologue of mllt3 and MLLT3 in humans. Quantitative reverse transcription-PCR and in situ hybridization analyses revealed that mllt3 is maternally supplied and zygotically expressed throughout embryonic development, and that expression is highest between 10 and 24 hours post-fertilization (hpf) coincident with enrichment in the intermediate cell mass (ICM) and posterior blood island, which are the sites of the primitive and transient definitive hematopoiesis in zebrafish, respectively. Further, we found co-expression of mllt3 with the early hematopoietic progenitor markers tal1, gata2, and gata1a in the posterior ICM. By investigating zebrafish hematopoietic mutants, we discovered that mllt3 is involved in erythroid precursor formation. By 48-72 hpf, mllt3 expression proved to be restricted to non-hematopoietic tissues including head structures, pronephric tubules, and liver primordium.
CONCLUSIONS
These findings establish a link between mllt3 and primitive erythropoiesis and provide the basis for future functional investigations.
Topics: Animals; Embryonic Development; Gene Expression Regulation, Developmental; Leukemia; Nuclear Proteins; Phylogeny; Transcription Factors; Zebrafish
PubMed: 35429189
DOI: 10.1002/dvdy.477 -
Communications Biology Oct 2021The enpp ectonucleotidases regulate lipidic and purinergic signalling pathways by controlling the extracellular concentrations of purines and bioactive lipids. Although...
The enpp ectonucleotidases regulate lipidic and purinergic signalling pathways by controlling the extracellular concentrations of purines and bioactive lipids. Although both pathways are key regulators of kidney physiology and linked to human renal pathologies, their roles during nephrogenesis remain poorly understood. We previously showed that the pronephros was a major site of enpp expression and now demonstrate an unsuspected role for the conserved vertebrate enpp4 protein during kidney formation in Xenopus. Enpp4 over-expression results in ectopic renal tissues and, on rare occasion, complete mini-duplication of the entire kidney. Enpp4 is required and sufficient for pronephric markers expression and regulates the expression of RA, Notch and Wnt pathway members. Enpp4 is a membrane protein that binds, without hydrolyzing, phosphatidylserine and its effects are mediated by the receptor s1pr5, although not via the generation of S1P. Finally, we propose a novel and non-catalytic mechanism by which lipidic signalling regulates nephrogenesis.
Topics: Animals; Body Patterning; Embryo, Nonmammalian; Embryonic Development; Gene Regulatory Networks; Kidney; Phosphoric Diester Hydrolases; Signal Transduction; Xenopus Proteins; Xenopus laevis
PubMed: 34620987
DOI: 10.1038/s42003-021-02688-9 -
Cell Discovery Mar 2022Primary cilia are antenna-like subcellular structures to act as signaling platforms to regulate many cellular processes and embryonic development. mA RNA modification...
Primary cilia are antenna-like subcellular structures to act as signaling platforms to regulate many cellular processes and embryonic development. mA RNA modification plays key roles in RNA metabolism and gene expression; however, the physiological function of mA modification remains largely unknown. Here we find that the mA demethylase ALKBH3 significantly inhibits ciliogenesis in mammalian cells by its demethylation activity. Mechanistically, ALKBH3 removes mA sites on mRNA of Aurora A, a master suppressor of ciliogenesis. Depletion of ALKBH3 enhances Aurora A mRNA decay and inhibits its translation. Moreover, alkbh3 morphants exhibit ciliary defects, including curved body, pericardial edema, abnormal otoliths, and dilation in pronephric ducts in zebrafish embryos, which are significantly rescued by wild-type alkbh3, but not by its catalytically inactive mutant. The ciliary defects caused by ALKBH3 depletion in both vertebrate cells and embryos are also significantly reversed by ectopic expression of Aurora A mRNA. Together, our data indicate that ALKBH3-dependent mA demethylation has a crucial role in the regulation of Aurora A mRNA, which is essential for ciliogenesis and cilia-associated developmental events in vertebrates.
PubMed: 35277482
DOI: 10.1038/s41421-022-00385-3 -
Frontiers in Genetics 2022The (Solute carrier family 12 member 3) gene encodes a sodium-chloride cotransporter and mediates Na and Cl reabsorption in the distal convoluted tubule of kidneys. An... (Review)
Review
The (Solute carrier family 12 member 3) gene encodes a sodium-chloride cotransporter and mediates Na and Cl reabsorption in the distal convoluted tubule of kidneys. An experimental study has previously showed that with knockdown of zebrafish ortholog, slc12a3 led to structural abnormality of kidney pronephric distal duct at 1-cell stage, suggesting that may have genetic effects in renal disorders. Many clinical reports have demonstrated that the function-loss mutations in the gene, mainly including Thr60Met, Asp486Asn, Gly741Arg, Leu859Pro, Arg861Cys, Arg913Gln, Arg928Cys and Cys994Tyr, play the pathogenic effects in Gitelman syndrome. This kidney disease is inherited as an autosomal recessive trait. In addition, several population genetic association studies have indicated that the single nucleotide variant Arg913Gln in the gene is associated with diabetic kidney disease in type 2 diabetes subjects. In this review, we first summarized bioinformatics of the gene and its genetic variation. We then described the different genetic and biological effects of in Gitelman syndrome and diabetic kidney disease. We also discussed about further genetic and biological analyses of as pharmacokinetic targets of diuretics.
PubMed: 35591852
DOI: 10.3389/fgene.2022.799224 -
Ecotoxicology and Environmental Safety Jul 2023Gold nanoparticles (AuNPs) are widely used in biomedicine and their specific properties including, size, geometrics, and surface coating, will affect their fate and...
Gold nanoparticles (AuNPs) are widely used in biomedicine and their specific properties including, size, geometrics, and surface coating, will affect their fate and behaviour in biological systems. These properties are well studied for their intended biological targets, but there is a lack of understanding on the mechanisms by which AuNPs interact in non-target organisms when they enter the environment. We investigated the effects of size and surface chemistry of AuNPs on their bioavailability, tissue distribution and potential toxicity using zebrafish (Danio rerio) as an experimental model. Larval zebrafish were exposed to fluorescently tagged AuNPs of different sizes (10-100 nm) and surface modifications (TNFα, NHS/PAMAM and PEG), and uptake, tissue distribution and depuration rates were measured using selective-plane illumination microscopy (SPIM). The gut and pronephric tubules were found to contain detectable levels of AuNPs, and the concentration-dependent accumulation was related to the particle size. Surface addition of PEG and TNFα appeared to enhance particle accumulation in the pronephric tubules compared to uncoated particles. Depuration studies showed a gradual removal of particles from the gut and pronephric tubules, although fluorescence indicating the presence of the AuNPs remained in the pronephros 96 h after exposure. Toxicity assessment using two transgenic zebrafish reporter lines, however, revealed no AuNP-related renal injury or cellular oxidative stress. Collectively, our data show that AuNPs used in medical applications across the size range 40-80 nm, are bioavailable to larval zebrafish and some may persist in renal tissue, although their presence did not result in measurable toxicity with respect to pronephric organ function or cellular oxidative stress for short term exposures.
Topics: Animals; Zebrafish; Gold; Metal Nanoparticles; Tumor Necrosis Factor-alpha; Tissue Distribution; Biological Availability; Particle Size
PubMed: 37269610
DOI: 10.1016/j.ecoenv.2023.115019 -
Aging Sep 2020Cellular senescence is considered a stress response imposing a stable cell cycle arrest to restrict the growth of damaged cells. More recently however, cellular...
Cellular senescence is considered a stress response imposing a stable cell cycle arrest to restrict the growth of damaged cells. More recently however, cellular senescence was identified during mouse embryo development at particular structures during specific periods of time. This programmed cell senescence has been proposed to serve developmental and morphogenetic functions and to potentially represent an evolutionary origin of senescence. Cellular senescence has also been described to take place during bird (chick and quail) and amphibian (xenopus and axoltl) development. Fish however, have been described to show a very narrow and restricted pattern of developmental cell senescence. Here we carried out a detailed characterization of senescence during zebrafish development and found it to be conserved and widespread. Apart from yolk and cloaca, previously described structures, we also identified senescence in the developing central nervous system, intestine, liver, pronephric ducts, and crystalline. Interestingly, senescence at these developing structures disappeared upon treatment with senolytic compound ABT-263, supporting their senescent identity and opening the possibility of studying the contribution of this process to development. In summary, our findings extend the description of developmentally-programmed cell senescence to lower vertebrates contributing to the notion of the relevance of this process for embryo development.
PubMed: 32991320
DOI: 10.18632/aging.103968 -
Computational and Structural... 2023Diabetic nephropathy (DN) is one of the most established microvascular complications of diabetes and a key cause of end-stage renal disease. It is well established that...
Diabetic nephropathy (DN) is one of the most established microvascular complications of diabetes and a key cause of end-stage renal disease. It is well established that gene susceptibility to DN plays a critical role in disease pathophysiology. Therefore, many genetic studies have been performed to categorize candidate genes in prominent diabetic cohorts, aiming to investigate DN pathogenesis and etiology. In this study, we performed a meta-analysis on the expression profiles of GSE1009, GSE30122, GSE96804, GSE99340, GSE104948, GSE104954, and GSE111154 to identify critical transcriptional factors associated with DN progression. The analysis was conducted for all individual datasets for each kidney tissue (glomerulus, tubules, and kidney cortex). We identified distinct clusters of susceptibility genes that were dysregulated in a renal compartment-specific pattern. Further, we recognized a small but a closely connected set of these susceptibility genes enriched for podocyte differentiation, several of which were characterized as genes encoding critical transcriptional factors (TFs) involved in DN development and podocyte function. To validate the role of identified TFs in DN progression, we functionally validated the three main TFs (DACH1, LMX1B, and WT1) identified through differential gene expression and network analysis using the hyperglycemic zebrafish model. We report that hyperglycemia-induced altered gene expression of the key TF genes leads to morphological abnormalities in zebrafish glomeruli, pronephric tubules, proximal and distal ducts. This study demonstrated that altered expression of these TF genes could be associated with hyperglycemia-induced nephropathy and, thus, aids in understanding the molecular drivers, essential genes, and pathways that trigger DN initiation and development.
PubMed: 36659918
DOI: 10.1016/j.csbj.2022.12.054 -
Cells Apr 2022The anterior-posterior (AP) axis in chordates is regulated by a conserved set of genes and signaling pathways, including genes and retinoic acid (RA), which play...
The anterior-posterior (AP) axis in chordates is regulated by a conserved set of genes and signaling pathways, including genes and retinoic acid (RA), which play well-characterized roles in the organization of the chordate body plan. The intermediate mesoderm (IM), which gives rise to all vertebrate kidneys, is an example of a tissue that differentiates sequentially along this axis. Yet, the conservation of the spatiotemporal regulation of the IM across vertebrates remains poorly understood. In this study, we used a comparative developmental approach focusing on non-conventional model organisms, a chondrichthyan (catshark), a cyclostome (lamprey), and a cephalochordate (amphioxus), to assess the involvement of RA in the regulation of chordate and vertebrate pronephros formation. We report that the anterior expression boundary of early pronephric markers ( and ), positioned at the level of somite 6 in amniotes, is conserved in the catshark and the lamprey. Furthermore, RA, driving the expression of genes like in amniotes, regulates the anterior pronephros boundary in the catshark. We find no evidence for the involvement of this regulatory hierarchy in the AP positioning of the lamprey pronephros and the amphioxus pronephros homolog, Hatschek's nephridium. This suggests that despite the conservation of and expressions in chordate pronephros homologs, the responsiveness of the IM, and hence of pronephric genes, to RA- and -dependent regulation is a gnathostome novelty.
Topics: Animals; Chordata; Genes, Homeobox; Lampreys; Pronephros; Tretinoin; Vertebrates
PubMed: 35455988
DOI: 10.3390/cells11081304 -
Cells May 2020Automated high-throughput workflows allow for chemical toxicity testing and drug discovery in zebrafish disease models. Due to its conserved structural and functional...
Automated high-throughput workflows allow for chemical toxicity testing and drug discovery in zebrafish disease models. Due to its conserved structural and functional properties, the zebrafish pronephros offers a unique model to study renal development and disease at larger scale. Ideally, scoring of pronephric phenotypes includes morphological and functional assessments within the same larva. However, to efficiently upscale such assays, refinement of existing methods is required. Here, we describe the development of a multiparametric in vivo screening pipeline for parallel assessment of pronephric morphology, kidney function and heart rate within the same larva on a single imaging platform. To this end, we developed a novel 3D-printed orientation tool enabling multiple consistent orientations of larvae in agarose-filled microplates. Dorsal pronephros imaging was followed by assessing renal clearance and heart rates upon fluorescein isothiocyanate (FITC)-inulin microinjection using automated time-lapse imaging of laterally positioned larvae. The pipeline was benchmarked using a set of drugs known to induce developmental nephrotoxicity in humans and zebrafish. Drug-induced reductions in renal clearance and heart rate alterations were detected even in larvae exhibiting minor pronephric phenotypes. In conclusion, the developed workflow enables rapid and semi-automated in vivo assessment of multiple morphological and functional parameters.
Topics: Animals; Biological Assay; Embryo, Nonmammalian; Fluorescein-5-isothiocyanate; Heart Function Tests; Heart Rate; Kidney; Larva; Pronephros; Zebrafish
PubMed: 32443839
DOI: 10.3390/cells9051269 -
Biology Open Jun 2022Endocytosis mediates the cellular uptake of numerous molecules from the extracellular space and is a fundamentally important process. In the renal proximal tubule, the...
Endocytosis mediates the cellular uptake of numerous molecules from the extracellular space and is a fundamentally important process. In the renal proximal tubule, the scavenger receptor megalin and its co-receptor cubilin mediate endocytosis of low molecular weight proteins from the renal filtrate. However, the extent to which megalin endocytosis relies on different components of the trafficking machinery remains relatively poorly defined in vivo. In this study, we identify a functional requirement for the F-BAR protein pacsin2 in endocytosis in the renal proximal tubule of zebrafish larvae. Pacsin2 is expressed throughout development and in all zebrafish tissues, similar to the mammalian orthologue. Within renal tubular epithelial cells, pacsin2 is enriched at the apical pole where it is localised to endocytic structures. Loss of pacsin2 results in reduced endocytosis within the proximal tubule, which is accompanied by a reduction in the abundance of megalin and endocytic organelles. Our results indicate that pacsin2 is required for efficient endocytosis in the proximal tubule, where it likely cooperates with other trafficking machinery to maintain endocytic uptake and recycling of megalin.
Topics: Animals; Biological Transport; Endocytosis; Kidney Tubules, Proximal; Low Density Lipoprotein Receptor-Related Protein-2; Mammals; Zebrafish
PubMed: 35616009
DOI: 10.1242/bio.059150