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Human Molecular Genetics Apr 2022Endocytosis is a fundamentally important process through which material is internalized into cells from the extracellular environment. In the renal proximal tubule,...
Endocytosis is a fundamentally important process through which material is internalized into cells from the extracellular environment. In the renal proximal tubule, endocytosis of the abundant scavenger receptor megalin and its co-receptor cubilin play a vital role in retrieving low molecular weight proteins from the renal filtrate. Although we know much about megalin and its ligands, the machinery and mechanisms by which the receptor is trafficked through the endosomal system remain poorly defined. In this study, we show that inositol phosphatase interacting protein of 27 kDa (Ipip27A), an interacting partner of the Lowe syndrome protein oculocerebrorenal syndrome of Lowe (OCRL), is required for endocytic traffic of megalin within the proximal renal tubule of zebrafish larvae. Knockout of Ipip27A phenocopies the endocytic phenotype seen upon loss of OCRL, with a deficit in uptake of both fluid-phase and protein cargo, which is accompanied by a reduction in megalin abundance and altered endosome morphology. Rescue and co-depletion experiments indicate that Ipip27A functions together with OCRL to support proximal tubule endocytosis. The results therefore identify Ipip27A as a new player in endocytic traffic in the proximal tubule in vivo and support the view that defective endocytosis underlies the renal tubulopathy in Lowe syndrome and Dent-2 disease.
Topics: Animals; Endocytosis; Endosomes; Female; Humans; Inositol Phosphates; Kidney Tubules, Proximal; Low Density Lipoprotein Receptor-Related Protein-2; Male; Oculocerebrorenal Syndrome; Phosphoric Monoester Hydrolases; Proteins; Zebrafish; Zebrafish Proteins
PubMed: 34673953
DOI: 10.1093/hmg/ddab307 -
Results and Problems in Cell... 2017The pronephros is the first kidney type to form in vertebrate embryos. The first step of pronephrogenesis in the zebrafish is the formation of the intermediate mesoderm... (Review)
Review
The pronephros is the first kidney type to form in vertebrate embryos. The first step of pronephrogenesis in the zebrafish is the formation of the intermediate mesoderm during gastrulation, which occurs in response to secreted morphogens such as BMPs and Nodals. Patterning of the intermediate mesoderm into proximal and distal cell fates is induced by retinoic acid signaling with downstream transcription factors including wt1a, pax2a, pax8, hnf1b, sim1a, mecom, and irx3b. In the anterior intermediate mesoderm, progenitors of the glomerular blood filter migrate and fuse at the midline and recruit a blood supply. More posteriorly localized tubule progenitors undergo epithelialization and fuse with the cloaca. The Notch signaling pathway regulates the formation of multi-ciliated cells in the tubules and these cells help propel the filtrate to the cloaca. The lumenal sheer stress caused by flow down the tubule activates anterior collective migration of the proximal tubules and induces stretching and proliferation of the more distal segments. Ultimately these processes create a simple two-nephron kidney that is capable of reabsorbing and secreting solutes and expelling excess water-processes that are critical to the homeostasis of the body fluids. The zebrafish pronephric kidney provides a simple, yet powerful, model system to better understand the conserved molecular and cellular progresses that drive nephron formation, structure, and function.
Topics: Animals; Models, Animal; Organogenesis; Pronephros; Zebrafish; Zebrafish Proteins
PubMed: 28409341
DOI: 10.1007/978-3-319-51436-9_2 -
Methods in Cell Biology 2016The kidney of the zebrafish shares many features with other vertebrate kidneys including the human kidney. Similar cell types and shared developmental and patterning...
The kidney of the zebrafish shares many features with other vertebrate kidneys including the human kidney. Similar cell types and shared developmental and patterning mechanisms make the zebrafish pronephros a valuable model for kidney organogenesis. Here we review recent advances in studies of zebrafish pronephric development and provide experimental protocols to analyze kidney cell types and structures, measure nephron function, live image kidney cells in vivo, and probe mechanisms of kidney regeneration after injury.
Topics: Animals; Gene Expression Regulation, Developmental; Humans; Kidney; Nephrons; Organogenesis; Pronephros; Regeneration; Zebrafish
PubMed: 27312500
DOI: 10.1016/bs.mcb.2016.03.041 -
Kidney International Jan 2023The kidney is an essential organ that ensures bodily fluid homeostasis and removes soluble waste products from the organism. Nephrons, the functional units of the...
The kidney is an essential organ that ensures bodily fluid homeostasis and removes soluble waste products from the organism. Nephrons, the functional units of the kidney, comprise a blood filter, the glomerulus or glomus, and an epithelial tubule that processes the filtrate from the blood or coelom and selectively reabsorbs solutes, such as sugars, proteins, ions, and water, leaving waste products to be eliminated in the urine. Genes coding for transporters are segmentally expressed, enabling the nephron to sequentially process the filtrate. The Xenopus embryonic kidney, the pronephros, which consists of a single large nephron, has served as a valuable model to identify genes involved in nephron formation and patterning. Therefore, the developmental patterning program that generates these segments is of great interest. Prior work has defined the gene expression profiles of Xenopus nephron segments via in situ hybridization strategies, but a comprehensive understanding of the cellular makeup of the pronephric kidney remains incomplete. Here, we carried out single-cell mRNA sequencing of the functional Xenopus pronephric nephron and evaluated its cellular composition through comparative analyses with previous Xenopus studies and single-cell mRNA sequencing of the adult mouse kidney. This study reconstructs the cellular makeup of the pronephric kidney and identifies conserved cells, segments, and associated gene expression profiles. Thus, our data highlight significant conservation in podocytes, proximal and distal tubule cells, and divergence in cellular composition underlying the capacity of each nephron to remove wastes in the form of urine, while emphasizing the Xenopus pronephros as a model for physiology and disease.
Topics: Animals; Mice; Gene Expression Regulation, Developmental; Kidney; Kidney Glomerulus; Nephrons; RNA, Messenger; Xenopus laevis
PubMed: 36055600
DOI: 10.1016/j.kint.2022.07.027 -
Methods in Cell Biology 2010The zebrafish pronephric kidney provides a useful and relevant model of kidney development and function. It is composed of cell types common to all vertebrate kidneys...
The zebrafish pronephric kidney provides a useful and relevant model of kidney development and function. It is composed of cell types common to all vertebrate kidneys and pronephric organogenesis is regulated by transcription factors that have highly conserved functions in mammalian kidney development. Pronephric nephrons are a good model of tubule segmentation and differentiation of epithelial cell types. The pronephric glomerulus provides a simple model to assay gene function in regulating cell structure and cell interactions that form the blood filtration apparatus. The relative simplicity of the pronephric kidney combined with the ease of genetic manipulation in zebrafish makes it well suited for mutation analysis and gene discovery, in vivo imaging, functional screens of candidate genes from other species, and cell isolation by FACS . In addition, the larval and adult zebrafish kidneys have emerged as systems to study kidney regeneration after injury. This chapter provides a review of pronephric structure and development as well as current methods to study the pronephros.
Topics: Animals; Embryo, Nonmammalian; Kidney; Zebrafish
PubMed: 21111220
DOI: 10.1016/B978-0-12-384892-5.00009-8 -
Development (Cambridge, England) Jan 1989On the basis of its distribution pattern in embryos of the axolotl (Ambystoma mexicanum), we recently identified alkaline phosphatase as a molecule potentially involved...
On the basis of its distribution pattern in embryos of the axolotl (Ambystoma mexicanum), we recently identified alkaline phosphatase as a molecule potentially involved in guiding the migration of the pronephric duct. Alkaline phosphatase is a cell surface protein anchored to cell membranes via a covalent linkage to a phosphatidylinositol glycan (PI-G). The enzyme phosphatidylinositol-specific phospholipase C (PIPLC) specifically releases from cell surfaces molecules anchored by the PI-G linkage. In order to test the possibility that a PI-G anchored protein is involved in directing pronephric duct cell migration, PIPLC was applied to axolotl embryos. The enzyme was introduced into embryos through the use of a novel slow-release bead material, hydrolysed polyacrylamide. PIPLC blocked pronephric duct cell migration without interfering with somite fissure formation, a concurrent, neighbouring morphogenetic cell rearrangement which occurs with little if any alkaline phosphatase present. In addition, alkaline phosphatase activity was markedly diminished in the vicinity of the implanted beads. These observations suggest that at least one protein anchored to the cell membrane by a PI-G linkage, possibly alkaline phosphatase, is involved in guiding or promoting pronephric duct cell migration.
Topics: Alkaline Phosphatase; Ambystoma; Animals; Cell Movement; Kidney; Microscopy, Electron, Scanning; Phosphatidylinositol Diacylglycerol-Lyase; Phosphoinositide Phospholipase C; Phosphoric Diester Hydrolases
PubMed: 2553384
DOI: 10.1242/dev.105.1.1 -
Developmental Biology Oct 1995The pronephros serves as the embryonic kidney of the lower vertebrates. In this report we describe the development of the pronephric system of Xenopus laevis utilizing...
The pronephros serves as the embryonic kidney of the lower vertebrates. In this report we describe the development of the pronephric system of Xenopus laevis utilizing scanning electron microscopy and novel monoclonal antibodies that specifically recognize different parts of the pronephros. Antibody 3G8 recognizes the tubules and nephrostomes of the pronephroi only and does not react with the duct. Antibody 4A6 stains only the duct and the nephrostomes. These antibodies thus allow the positive identification of these two intermediate mesoderm derivatives. Both reagents detect antigens expressed some time after the pronephric structures first form and probably represent markers of terminal differentiation. When the tubules and duct first form they are separate structures that can easily be distinguished; the connective tubules have a distinctive organization, the collecting (or common) tubule is broader than other tubules, and the narrow pronephric duct has a specific shape and position. In later stages the collecting tubule and the rostral portion of the duct undergo a considerable amount of convolution, and both contribute to the final coiled tubular body of the pronephros. The ability of 3G8 and 4A6 to distinguish these two elements of the nephric system was used to reexplore classical experiments on the interaction between these two structures during development of the pronephric system. The use of whole-mount analysis has allowed us to examine large numbers of embryos from different stages and dissected in a variety of planes. These experiments demonstrate the dynamic nature of the intermediate mesoderm and indicate that although the pronephros may be specified by mid-neurula stages, patterning is not complete until tailbud stages.
Topics: Animals; Antibodies, Monoclonal; Biomarkers; Female; Kidney; Xenopus laevis
PubMed: 7556934
DOI: 10.1006/dbio.1995.1302 -
Scientific Reports Feb 2017The human ubiquitous protein cystinosin is responsible for transporting the disulphide amino acid cystine from the lysosomal compartment into the cytosol. In humans,...
The human ubiquitous protein cystinosin is responsible for transporting the disulphide amino acid cystine from the lysosomal compartment into the cytosol. In humans, Pathogenic mutations of CTNS lead to defective cystinosin function, intralysosomal cystine accumulation and the development of cystinosis. Kidneys are initially affected with generalized proximal tubular dysfunction (renal Fanconi syndrome), then the disease rapidly affects glomeruli and progresses towards end stage renal failure and multiple organ dysfunction. Animal models of cystinosis are limited, with only a Ctns knockout mouse reported, showing cystine accumulation and late signs of tubular dysfunction but lacking the glomerular phenotype. We established and characterized a mutant zebrafish model with a homozygous nonsense mutation (c.706āCā>āT; p.Q236X) in exon 8 of ctns. Cystinotic mutant larvae showed cystine accumulation, delayed development, and signs of pronephric glomerular and tubular dysfunction mimicking the early phenotype of human cystinotic patients. Furthermore, cystinotic larvae showed a significantly increased rate of apoptosis that could be ameliorated with cysteamine, the human cystine depleting therapy. Our data demonstrate that, ctns gene is essential for zebrafish pronephric podocyte and proximal tubular function and that the ctns-mutant can be used for studying the disease pathogenic mechanisms and for testing novel therapies for cystinosis.
Topics: Amino Acid Sequence; Amino Acid Transport Systems, Neutral; Animals; Apoptosis; Cystine; Cystinosis; Disease Models, Animal; Gene Knockout Techniques; Glomerular Filtration Rate; Humans; Kidney Glomerulus; Kidney Tubules, Proximal; Locomotion; Lysosomes; Mutation; Phenotype; Podocytes; Zebrafish
PubMed: 28198397
DOI: 10.1038/srep42583 -
Mechanisms of Development 2009The pronephric kidney controls water and electrolyte balance during early fish and amphibian embryogenesis. Many Wnt signaling components have been implicated in kidney...
The pronephric kidney controls water and electrolyte balance during early fish and amphibian embryogenesis. Many Wnt signaling components have been implicated in kidney development. Specifically, in Xenopus pronephric development as well as the murine metanephroi, the secreted glycoprotein Wnt-4 has been shown to be essential for renal tubule formation. Despite the importance of Wnt signals in kidney organogenesis, little is known of the definitive downstream signaling pathway(s) that mediate their effects. Here we report that inhibition of Wnt/beta-catenin signaling within the pronephric field of Xenopus results in significant losses to kidney epithelial tubulogenesis with little or no effect on adjoining axis or somite development. We find that the requirement for Wnt/beta-catenin signaling extends throughout the pronephric primordium and is essential for the development of proximal and distal tubules of the pronephros as well as for the development of the duct and glomus. Although less pronounced than effects upon later pronephric tubule differentiation, inhibition of the Wnt/beta-catenin pathway decreased expression of early pronephric mesenchymal markers indicating it is also needed in early pronephric patterning. We find that upstream inhibition of Wnt/beta-catenin signals in zebrafish likewise reduces pronephric epithelial tubulogenesis. We also find that exogenous activation of Wnt/beta-catenin signaling within the Xenopus pronephric field results in significant tubulogenic losses. Together, we propose Wnt/beta-catenin signaling is required for pronephric tubule, duct and glomus formation in Xenopus laevis, and this requirement is conserved in zebrafish pronephric tubule formation.
Topics: Animals; Biomarkers; Embryo, Nonmammalian; Gene Expression Regulation, Developmental; Homeodomain Proteins; Intercellular Signaling Peptides and Proteins; Kidney; Kidney Tubules, Collecting; Larva; Membrane Proteins; Mesoderm; Mutation; Organogenesis; Phenotype; Recombinant Fusion Proteins; Signal Transduction; Somites; Wnt Proteins; Xenopus; Xenopus Proteins; Zebrafish; beta Catenin
PubMed: 19100832
DOI: 10.1016/j.mod.2008.11.007 -
Development, Growth & Differentiation Apr 2002The earliest form of embryonic kidney, the pronephros, consists of three components: glomus, tubule and duct. Treatment of the undifferentiated animal pole ectoderm of...
The earliest form of embryonic kidney, the pronephros, consists of three components: glomus, tubule and duct. Treatment of the undifferentiated animal pole ectoderm of Xenopus laevis with activin A and retinoic acid (RA) induces formation of the pronephric tubule and glomus. In this study, the rate of induction of the pronephric duct, the third component of the pronephros, was investigated in animal caps treated with activin A and RA. Immunohistochemistry using pronephric duct-specific antibody 4A6 revealed that a high proportion of the treated explants contained 4A6-positive tubular structures. Electron microscopy showed that the tubules in the explants were similar to the pronephric ducts of normal larvae, and they also expressed Gremlin and c-ret, molecular markers for pronephric ducts. These results suggest that the treatment of Xenopus ectoderm with activin A and RA induces a high rate of differentiation of pronephric ducts, in addition to the differentiation of the pronephric tubule and glomus, and that this in vitro system can serve as a simple and effective model for analysis of the mechanism of pronephros differentiation.
Topics: Activins; Animals; Cell Differentiation; Cytokines; Drosophila Proteins; Ectoderm; Embryonic Induction; Genetic Markers; Inhibin-beta Subunits; Intercellular Signaling Peptides and Proteins; Kidney; Proteins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-ret; Receptor Protein-Tyrosine Kinases; Tretinoin; Xenopus Proteins; Xenopus laevis
PubMed: 11940102
DOI: 10.1046/j.1440-169x.2002.00631.x