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1.
Kidney Int ; 105(4): 731-743, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38158181

RESUMO

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a leading cause of kidney failure and is associated with substantial morbidity and mortality. Interstitial inflammation is attributed to the action of infiltrating macrophages and is a feature thought to aggravate disease progression. Here, we investigated the therapeutic potential of the anti-inflammatory IL37b cytokine as a treatment for ADPKD using genetic mouse models, demonstrating that transgenic expression of human IL37b reduced collecting duct cyst burden in both early and adult-onset ADPKD rodent models. Moreover, injection of recombinant human IL37b could also reduce cyst burden in early onset ADPKD mice, an observation not associated with increased macrophage number at early stages of cyst formation. Interestingly, transgenic IL37b expression also did not alter macrophage numbers in advanced disease. Whole kidney RNA-seq highlighted an IL37b-mediated upregulation of the interferon signaling pathway and single-cell RNA-seq established that these changes originate at least partly from kidney resident macrophages. We further found that blocking type I interferon signaling in mice expressing IL37b resulted in increased cyst number, confirming this as an important pathway by which IL37b exerts its beneficial effects. Thus, our studies show that IL37b promotes interferon signaling in kidney resident macrophages which suppresses cyst initiation, identifying this protein as a potential therapy for ADPKD.


Assuntos
Cistos , Rim Policístico Autossômico Dominante , Camundongos , Humanos , Animais , Rim Policístico Autossômico Dominante/tratamento farmacológico , Rim Policístico Autossômico Dominante/genética , Inflamação/genética , Inflamação/complicações , Rim/metabolismo , Cistos/complicações , Interleucinas , Interferons
2.
Development ; 147(21)2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32439764

RESUMO

Laminin alpha 5 (LAMA5) is a member of a large family of proteins that trimerise and then polymerise to form a central component of all basement membranes. Consequently, the protein plays an instrumental role in shaping the normal development of the kidney, skin, neural tube, lung and limb, and many other organs and tissues. Pathogenic mutations in some laminins have been shown to cause a range of largely syndromic conditions affecting the competency of the basement membranes to which they contribute. We report the identification of a mutation in the polymerisation domain of LAMA5 in a patient with a complex syndromic disease characterised by defects in kidney, craniofacial and limb development, and by a range of other congenital defects. Using CRISPR-generated mouse models and biochemical assays, we demonstrate the pathogenicity of this variant, showing that the change results in a failure of the polymerisation of α/ß/γ laminin trimers. Comparing these in vivo phenotypes with those apparent upon gene deletion in mice provides insights into the specific functional importance of laminin polymerisation during development and tissue homeostasis.


Assuntos
Deficiências do Desenvolvimento/genética , Desenvolvimento Fetal , Laminina/genética , Mutação/genética , Polimerização , Sequência de Aminoácidos , Animais , Animais Recém-Nascidos , Pré-Escolar , Deficiências do Desenvolvimento/patologia , Feto/embriologia , Humanos , Hidronefrose/patologia , Recém-Nascido , Rim/anormalidades , Rim/embriologia , Rim/patologia , Laminina/química , Pulmão/anormalidades , Pulmão/embriologia , Pulmão/patologia , Masculino , Camundongos , Domínios Proteicos , Síndrome
3.
Dev Biol ; 454(2): 156-169, 2019 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-31242448

RESUMO

Adamts18 encodes a secreted metalloprotease restricted to branch-tip progenitor pools directing the morphogenesis of multiple mammalian organs. Adamts18 was targeted to explore a potential role in branching morphogenesis. In the kidney, an arborized collecting system develops through extensive branching morphogenesis of an initial epithelial outgrowth of the mesonephric duct, the ureteric bud. Adamts18 mutants displayed a weakly penetrant phenotype: duplicated ureteric outgrowths forming enlarged, bi-lobed kidneys with an increased nephron endowment. In contrast, Adamts18 mutants showed a fully penetrant lung phenotype: epithelial growth was markedly reduced and early secondary branching scaled to the reduced length of the primary airways. Furthermore, there was a pronounced delay in the appearance of differentiated cell types in both proximal and distally positions of the developing airways. Adamts18 is closely related to Adamts16. In the kidney but not the lung, broad epithelial Adamts16 expression overlaps Adamts18 in branch tips. However, compound Adamts16/18 mutants displayed a comparable low penetrance duplicated ureteric phenotype, ruling out a possible role for Adamts16 as a functional modifier of the Adamts18 kidney phenotype. Given the predicted action of secreted Adamts18 metalloprotease, and broad expression of Adamts18 in branching organ systems, these findings suggest distinct requirements for matrix modelling in the morphogenesis of epithelial networks.


Assuntos
Proteínas ADAMTS/metabolismo , Organogênese/fisiologia , Proteínas ADAMTS/fisiologia , Animais , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Rim/citologia , Rim/embriologia , Rim/metabolismo , Pulmão/embriologia , Pulmão/metabolismo , Masculino , Metaloproteases/genética , Metaloproteases/metabolismo , Camundongos , Camundongos Knockout , Morfogênese , Néfrons/metabolismo , Técnicas de Cultura de Órgãos/métodos , Ureter/metabolismo
4.
Development ; 144(23): 4377-4385, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29038307

RESUMO

Metanephric kidney development is orchestrated by the iterative branching morphogenesis of the ureteric bud. We describe an underlying patterning associated with the ramification of this structure and show that this pattern is conserved between developing kidneys, in different parts of the organ and across developmental time. This regularity is associated with a highly reproducible branching asymmetry that is consistent with locally operative growth mechanisms. We then develop a class of tip state models to represent elaboration of the ureteric tree and describe rules for 'half-delay' branching morphogenesis that describe almost perfectly the patterning of this structure. Spatial analysis suggests that the observed asymmetry may arise from mutual suppression of bifurcation, but not extension, between the growing ureteric tips, and demonstrates that disruption of patterning occurs in mouse mutants in which the distribution of tips on the surface of the kidney is altered. These findings demonstrate that kidney development occurs by way of a highly conserved reiterative pattern of asymmetric bifurcation that is governed by intrinsic and locally operative mechanisms.


Assuntos
Rim/embriologia , Morfogênese/fisiologia , Ureter/embriologia , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Padronização Corporal/genética , Padronização Corporal/fisiologia , Proteína Morfogenética Óssea 7/deficiência , Proteína Morfogenética Óssea 7/genética , Proteína Morfogenética Óssea 7/fisiologia , Imageamento Tridimensional , Conceitos Matemáticos , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , Modelos Biológicos , Morfogênese/genética , Mutação , Fenótipo , Fosfoproteínas/deficiência , Fosfoproteínas/genética , Fosfoproteínas/fisiologia , Fator de Crescimento Transformador beta2/deficiência , Fator de Crescimento Transformador beta2/genética , Fator de Crescimento Transformador beta2/fisiologia
5.
Development ; 142(8): 1458-69, 2015 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-25790853

RESUMO

Epigenetic mechanisms involved in the establishment of lung epithelial cell lineage identities during development are largely unknown. Here, we explored the role of the histone methyltransferase Ezh2 during lung lineage determination. Loss of Ezh2 in the lung epithelium leads to defective lung formation and perinatal mortality. We show that Ezh2 is crucial for airway lineage specification and alveolarization. Using optical projection tomography imaging, we found that branching morphogenesis is affected in Ezh2 conditional knockout mice and the remaining bronchioles are abnormal, lacking terminally differentiated secretory club cells. Remarkably, RNA-seq analysis revealed the upregulation of basal genes in Ezh2-deficient epithelium. Three-dimensional imaging for keratin 5 further showed the unexpected presence of a layer of basal cells from the proximal airways to the distal bronchioles in E16.5 embryos. ChIP-seq analysis indicated the presence of Ezh2-mediated repressive marks on the genomic loci of some but not all basal genes, suggesting an indirect mechanism of action of Ezh2. We found that loss of Ezh2 de-represses insulin-like growth factor 1 (Igf1) expression and that modulation of IGF1 signaling ex vivo in wild-type lungs could induce basal cell differentiation. Altogether, our work reveals an unexpected role for Ezh2 in controlling basal cell fate determination in the embryonic lung endoderm, mediated in part by repression of Igf1 expression.


Assuntos
Diferenciação Celular/fisiologia , Fator de Crescimento Insulin-Like I/metabolismo , Pulmão/citologia , Pulmão/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Animais , Diferenciação Celular/genética , Imunoprecipitação da Cromatina , Proteína Potenciadora do Homólogo 2 de Zeste , Citometria de Fluxo , Fator de Crescimento Insulin-Like I/genética , Queratina-5/genética , Queratina-5/metabolismo , Pulmão/embriologia , Camundongos , Complexo Repressor Polycomb 2/genética , Reação em Cadeia da Polimerase
6.
J Am Soc Nephrol ; 27(10): 3093-3104, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26975438

RESUMO

Podocyte depletion is sufficient for the development of numerous glomerular diseases and can be absolute (loss of podocytes) or relative (reduced number of podocytes per volume of glomerulus). Commonly used methods to quantify podocyte depletion introduce bias, whereas gold standard stereologic methodologies are time consuming and impractical. We developed a novel approach for assessing podocyte depletion in whole glomeruli that combines immunofluorescence, optical clearing, confocal microscopy, and three-dimensional analysis. We validated this method in a transgenic mouse model of selective podocyte depletion, in which we determined dose-dependent alterations in several quantitative indices of podocyte depletion. This new approach provides a quantitative tool for the comprehensive and time-efficient analysis of podocyte depletion in whole glomeruli.


Assuntos
Contagem de Células/métodos , Tamanho Celular , Glomérulos Renais/citologia , Podócitos/citologia , Animais , Imageamento Tridimensional , Camundongos
7.
Mamm Genome ; 26(1-2): 57-79, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25552398

RESUMO

Genetic background plays a dominant role in mammary gland development and breast cancer (BrCa). Despite this, the role of genetics is only partially understood. This study used strain-dependent variation in an inbred mouse mapping panel, to identify quantitative trait loci (QTL) underlying structural variation in mammary ductal development, and determined if these QTL correlated with genomic intervals conferring BrCa susceptibility in humans. For about half of the traits, developmental variation among the complete set of strains in this study was greater (P < 0.05) than that of previously studied strains, or strains in current common use for mammary gland biology. Correlations were also detected with previously reported variation in mammary tumor latency and metastasis. In-silico genome-wide association identified 20 mammary development QTL (Mdq). Of these, five were syntenic with previously reported human BrCa loci. The most significant (P = 1 × 10(-11)) association of the study was on MMU6 and contained the genes Plxna4, Plxna4os1, and Chchd3. On MMU5, a QTL was detected (P = 8 × 10(-7)) that was syntenic to a human BrCa locus on h12q24.5 containing the genes Tbx3 and Tbx5. Intersection of linked SNP (r(2) > 0.8) with genomic and epigenomic features, and intersection of candidate genes with gene expression and survival data from human BrCa highlighted several for further study. These results support the conclusion that mammary tumorigenesis and normal ductal development are influenced by common genetic factors and that further studies of genetically diverse mice can improve our understanding of BrCa in humans.


Assuntos
Neoplasias da Mama/genética , Glândulas Mamárias Animais/crescimento & desenvolvimento , Camundongos Endogâmicos/genética , Locos de Características Quantitativas/genética , Animais , Neoplasias da Mama/fisiopatologia , Mapeamento Cromossômico , Simulação por Computador , Feminino , Estudo de Associação Genômica Ampla , Técnicas Histológicas , Humanos , Camundongos , Polimorfismo de Nucleotídeo Único/genética , Especificidade da Espécie , Sintenia/genética , Tomografia Óptica
8.
J Theor Biol ; 365: 226-37, 2015 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-25308508

RESUMO

Bifurcating developmental branching morphogenesis gives rise to complex organs such as the lung and the ureteric tree of the kidney. However, a few quantitative methods or tools exist to compare and distinguish, at a structural level, the critical features of these important biological systems. Here we develop novel graph alignment techniques to quantify the structural differences of rooted bifurcating trees and demonstrate their application in the analysis of developing kidneys from in normal and mutant mice. We have developed two graph based metrics: graph discordance, which measures how well the graphs representing the branching structures of distinct trees graphs can be aligned or overlayed; and graph inclusion, which measures the degree of containment of a tree graph within another. To demonstrate the application of these approaches we first benchmark the discordance metric on a data set of 32 normal and 28Tgfß(+/-) mutant mouse ureteric trees. We find that the discordance metric better distinguishes control and mutant mouse kidneys than alternative metrics based on graph size and fingerprints - the distribution of tip depths. Using this metric we then show that the structure of the mutant trees follows the same pattern as the normal kidneys, but undergo a major delay in elaboration at later stages. Analysis of both controls and mutants using the inclusion metric gives strong support to the hypothesis that ureteric tree growth is stereotypic. Additionally, we present a new generalised multi-tree alignment algorithm that minimises the sum of pairwise graph discordance and which can be used to generate maximum consensus trees that represent the archetype for fixed developmental stages. These tools represent an advance in the analysis and quantification of branching patterns and will be invaluable in gaining a deeper understanding of the mechanisms that drive development. All code is being made available with documentation and example data with this publication.


Assuntos
Morfogênese , Ureter/crescimento & desenvolvimento , Animais , Rim/crescimento & desenvolvimento , Rim/metabolismo , Camundongos , Mutação/genética , Fator de Crescimento Transformador beta2/metabolismo , Ureter/metabolismo
9.
PLoS Genet ; 7(9): e1002278, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21931569

RESUMO

The premature fusion of the paired frontal bones results in metopic craniosynostosis (MC) and gives rise to the clinical phenotype of trigonocephaly. Deletions of chromosome 9p22.3 are well described as a cause of MC with variably penetrant midface hypoplasia. In order to identify the gene responsible for the trigonocephaly component of the 9p22.3 syndrome, a cohort of 109 patients were assessed by high-resolution arrays and MLPA for copy number variations (CNVs) involving 9p22. Five CNVs involving FREM1, all of which were de novo variants, were identified by array-based analyses. The remaining 104 patients with MC were then subjected to targeted FREM1 gene re-sequencing, which identified 3 further mutant alleles, one of which was de novo. Consistent with a pathogenic role, mouse Frem1 mRNA and protein expression was demonstrated in the metopic suture as well as in the pericranium and dura mater. Micro-computed tomography based analyses of the mouse posterior frontal (PF) suture, the human metopic suture equivalent, revealed advanced fusion in all mice homozygous for either of two different Frem1 mutant alleles, while heterozygotes exhibited variably penetrant PF suture anomalies. Gene dosage-related penetrance of midfacial hypoplasia was also evident in the Frem1 mutants. These data suggest that CNVs and mutations involving FREM1 can be identified in a significant percentage of people with MC with or without midface hypoplasia. Furthermore, we present Frem1 mutant mice as the first bona fide mouse model of human metopic craniosynostosis and a new model for midfacial hypoplasia.


Assuntos
Cromossomos Humanos Par 9/genética , Craniossinostoses/genética , Variações do Número de Cópias de DNA , Proteínas da Matriz Extracelular/genética , Receptores de Interleucina/genética , Animais , Suturas Cranianas/anormalidades , Suturas Cranianas/patologia , Citocinas/genética , Heterozigoto , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Fenótipo , Deleção de Sequência
10.
Nat Commun ; 15(1): 371, 2024 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-38191531

RESUMO

Aurora Kinase A (AURKA) promotes cell proliferation and is overexpressed in different types of polycystic kidney disease (PKD). To understand AURKA's role in regulating renal cyst development we conditionally deleted the gene in mouse models of Autosomal Dominant PKD (ADPKD) and Joubert Syndrome, caused by Polycystin 1 (Pkd1) and Inositol polyphosphate-5-phosphatase E (Inpp5e) mutations respectively. We show that while Aurka is dispensable for collecting duct development and homeostasis, its deletion prevents cyst formation in both disease models. Cross-comparison of transcriptional changes implicated AKT signaling in cyst prevention and we show that (i) AURKA and AKT physically interact, (ii) AURKA regulates AKT activity in a kinase-independent manner and (iii) inhibition of AKT can reduce disease severity. AKT activation also regulates Aurka expression, creating a feed-forward loop driving renal cystogenesis. We find that the AURKA kinase inhibitor Alisertib stabilises the AURKA protein, agonizing its cystogenic functions. These studies identify AURKA as a master regulator of renal cyst development in different types of PKD, functioning in-part via AKT.


Assuntos
Aurora Quinase A , Cistos , Doenças Renais Policísticas , Rim Policístico Autossômico Dominante , Animais , Camundongos , Aurora Quinase A/genética , Monoéster Fosfórico Hidrolases , Doenças Renais Policísticas/genética , Doenças Renais Policísticas/prevenção & controle , Proteínas Proto-Oncogênicas c-akt/genética
11.
bioRxiv ; 2024 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-38106143

RESUMO

Background: Low nephron number has a direct impact on the development of hypertension and chronic kidney disease later in life. While intrauterine growth restriction caused by maternal low protein diet (LPD) is thought to be a significant cause of reduced nephron endowment in impoverished communities, its influence on the cellular and molecular processes which drive nephron formation are poorly understood. Methods: We conducted a comprehensive characterization of the impact of LPD on kidney development using tomographic and confocal imaging to quantify changes in branching morphogenesis and the cellular and morphological features of nephrogenic niches across development. These analyses were paired with single-cell RNA sequencing to dissect the transcriptional changes that LPD imposes during renal development to affect nephron number. Results: Single cell analysis at E14.5 and P0 revealed differences in the expression of genes and pathways involved in metabolism, cell cycle, epigenetic regulators and reciprocal inductive signals in most cell types analyzed, yielding imbalances and shifts in cellular energy production and cellular trajectories. In the nephron progenitor cells, LPD impeded cellular commitment and differentiation towards pre-tubular and renal vesicle structures. Confocal microscopy revealed a reduction in the number of pre-tubular aggregates and proliferation in nephron progenitor cells. We also found changes in branching morphogenesis, with a reduction in cell proliferation in the ureteric tips as well as reduced tip and tip parent lengths by optical projection tomography which causes patterning defects. Conclusions: This unique profiling demonstrates how a fetal programming defect leads to low nephron endowment which is intricately linked to changes in both branching morphogenesis and the commitment of nephron progenitor cells. The commitment of progenitor cells is pivotal for nephron formation and is significantly influenced by nutritional factors, with a low protein diet driving alterations in this program which directly results in a reduced nephron endowment. Significance Statement: While a mother's diet can negatively impact the number of nephrons in the kidneys of her offspring, the root cellular and molecular drivers of these deficits have not been rigorously explored. In this study we use advanced imaging and gene expression analysis in mouse models to define how a maternal low protein diet, analogous to that of impoverished communities, results in reduced nephron endowment. We find that low protein diet has pleiotropic effects on metabolism and the normal developmental programs of gene expression. These profoundly impact the process of branching morphogenesis necessary to establish niches for nephron generation and change cell behaviors which regulate how and when nephron progenitor cells commit to differentiation.

12.
J Med Genet ; 48(6): 375-82, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21507892

RESUMO

BACKGROUND: Manitoba-oculo-tricho-anal (MOTA) syndrome is a rare condition defined by eyelid colobomas, cryptophthalmos and anophthalmia/microphthalmia, an aberrant hairline, a bifid or broad nasal tip, and gastrointestinal anomalies such as omphalocele and anal stenosis. Autosomal recessive inheritance had been assumed because of consanguinity in the Oji-Cre population of Manitoba and reports of affected siblings, but no locus or cytogenetic aberration had previously been described. METHODS AND RESULTS: This study shows that MOTA syndrome is caused by mutations in FREM1, a gene previously mutated in bifid nose, renal agenesis, and anorectal malformations (BNAR) syndrome. MOTA syndrome and BNAR syndrome can therefore be considered as part of a phenotypic spectrum that is similar to, but distinct from and less severe than, Fraser syndrome. Re-examination of Frem1(bat/bat) mutant mice found new evidence that Frem1 is involved in anal and craniofacial development, with anal prolapse, eyelid colobomas, telecanthus, a shortened snout and reduced philtral height present in the mutant mice, similar to the human phenotype in MOTA syndrome. CONCLUSIONS: The milder phenotypes associated with FREM1 deficiency in humans (MOTA syndrome and BNAR syndrome) compared to that resulting from FRAS1 and FREM2 loss of function (Fraser syndrome) are also consistent with the less severe phenotypes resulting from Frem1 loss of function in mice. Together, Fraser, BNAR and MOTA syndromes constitute a clinically overlapping group of FRAS-FREM complex diseases.


Assuntos
Anormalidades Múltiplas/genética , Anus Imperfurado/genética , Coloboma/genética , Proteínas da Matriz Extracelular/genética , Síndrome de Fraser/genética , Hipertelorismo/genética , Receptores de Interleucina/genética , Anormalidades Múltiplas/patologia , Adolescente , Adulto , Canal Anal/anormalidades , Canal Anal/patologia , Animais , Malformações Anorretais , Anus Imperfurado/patologia , Sequência de Bases , Criança , Pré-Escolar , Coloboma/patologia , Pálpebras/anormalidades , Feminino , Síndrome de Fraser/patologia , Dosagem de Genes , Hérnia Umbilical/genética , Hérnia Umbilical/patologia , Humanos , Hipertelorismo/patologia , Masculino , Camundongos , Dados de Sequência Molecular , Mutação , Nariz/anormalidades , Doenças Nasais/genética , Análise de Sequência com Séries de Oligonucleotídeos , Linhagem , Fenótipo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Síndrome
13.
Kidney Int ; 77(12): 1132-9, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20200502

RESUMO

Branching morphogenesis is a central process in renal development, but imaging and quantifying this process beyond early organogenesis presents challenges due to growth of the kidney preventing ready imaging of the complex structures. Current analysis of renal development relies heavily on explant organ culture and visualization by confocal microscopy, as a more developmentally advanced native tissue is too thick for conventional microscopic imaging. Cultured renal primordia lack vascularization and a supportive matrix for normal growth, resulting in tissue compression and distortion of ureteric branching. To overcome this, we used optical projection tomography to image and reconstruct the branching ureter epithelium of ex vivo embryonic kidneys and developed software to quantify these three-dimensional (3D) data. Ureteric branching was assessed by measuring tree and terminal branch length, tip number, branching iterations, branch angles, and inter-tip distances in 3D space. To validate this approach for analyzing genetic influences on renal development, we assessed branching and organ morphology in Tgfbeta2(+/-) embryos from E12.5 through E15.5. We found decreased branching, contrary to previous findings using organ culture, and quantified a primary defect in renal pelvic formation. Our approach offers many advantages from improved throughput, analysis, and observation of in vivo branching states, and has demonstrated its utility in studying the basis of renal developmental disease.


Assuntos
Rim/embriologia , Morfogênese , Tomografia/métodos , Animais , Epitélio , Camundongos , Fator de Crescimento Transformador beta/genética , Ureter/embriologia , Ureter/crescimento & desenvolvimento
14.
Anat Rec (Hoboken) ; 303(10): 2578-2587, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32790143

RESUMO

Branching morphogenesis is an integral developmental mechanism central to the formation of a range of organs including the kidney, lung, pancreas and mammary gland. The ramified networks of epithelial tubules it establishes are critical for the processes of secretion, excretion and exchange mediated by these tissues. In the kidney, branching serves to establish the collecting duct system that transports urine from the nephrons into the renal pelvis, ureter and finally the bladder. Generally speaking, the formation of these networks in different organs begins with the specification and differentiation of simple bud-like organ anlage, which then undergo a process of elaboration, typically by bifurcation. This process is often governed by the interaction of progenitor cells at the tips of the epithelia with neighboring mesenchymal cell populations which direct the branching process and which often themselves differentiate to form part of the adult organ. In the kidney, the tips of ureteric bud elaborate through a dynamic cell signaling relationship with overlying nephron progenitor cell populations. These cells sequentially commit to differentiation and the resulting nephrons reintegrate with the ureteric epithelium as development progresses. This review will describe recent advances in understanding the how the elaboration of the ureteric bud is patterned and consider the extent to which this process is shared with other organs.


Assuntos
Diferenciação Celular/fisiologia , Rim/embriologia , Organogênese/fisiologia , Humanos , Células-Tronco/fisiologia
15.
J Mol Biol ; 369(1): 1-10, 2007 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-17428496

RESUMO

The B-box type 2 domain is a prominent feature of a large and growing family of RING, B-box, coiled-coil (RBCC) domain-containing proteins and is also present in more than 1500 additional proteins. Most proteins usually contain a single B-box2 domain, although some proteins contain tandem domains consisting of both type 1 and type 2 B-boxes, which actually share little sequence similarity. Recently, we determined the solution structure of B-box1 from MID1, a putative E3 ubiquitin ligase that is mutated in X-linked Opitz G/BBB syndrome, and showed that it adopted a betabetaalpha RING-like fold. Here, we report the tertiary structure of the B-box2 (CHC(D/C)C(2)H(2)) domain from MID1 using multidimensional NMR spectroscopy. This MID1 B-box2 domain consists of a short alpha-helix and a structured loop with two short anti-parallel beta-strands and adopts a tertiary structure similar to the B-box1 and RING structures, even though there is minimal primary sequence similarity between these domains. By mutagenesis, ESI-FTICR and ICP mass spectrometry, we show that the B-box2 domain coordinates two zinc atoms with a 'cross-brace' pattern: one by Cys175, His178, Cys195 and Cys198 and the other by Cys187, Asp190, His204, and His207. Interestingly, this is the first case that an aspartic acid is involved in zinc atom coordination in a zinc-finger domain, although aspartic acid has been shown to coordinate non-catalytic zinc in matrix metalloproteinases. In addition, the finding of a Cys195Phe substitution identified in a patient with X-linked Opitz GBBB syndrome supports the importance of proper zinc coordination for the function of the MID1 B-box2 domain. Notably, however, our structure differs from the only other published B-box2 structure, that from XNF7, which was shown to coordinate one zinc atom. Finally, the similarity in tertiary structures of the B-box2, B-box1 and RING domains suggests these domains have evolved from a common ancestor.


Assuntos
Sequência Conservada , Evolução Molecular , Proteínas dos Microtúbulos/química , Proteínas dos Microtúbulos/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Dobramento de Proteína , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Dedos de Zinco , Zinco/metabolismo , Sequência de Aminoácidos , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Fosfoproteínas/química , Estrutura Terciária de Proteína , Soluções , Ubiquitina-Proteína Ligases
16.
Elife ; 72018 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-30063208

RESUMO

Branching morphogenesis of the ureteric bud is integral to kidney development; establishing the collecting ducts of the adult organ and driving organ expansion via peripheral interactions with nephron progenitor cells. A recent study suggested that termination of tip branching within the developing kidney involved stochastic exhaustion in response to nephron formation, with such a termination event representing a unifying developmental process evident in many organs. To examine this possibility, we have profiled the impact of nephron formation and maturation on elaboration of the ureteric bud during mouse kidney development. We find a distinct absence of random branch termination events within the kidney or evidence that nephrogenesis impacts the branching program or cell proliferation in either tip or progenitor cell niches. Instead, organogenesis proceeds in a manner indifferent to the development of these structures. Hence, stochastic cessation of branching is not a unifying developmental feature in all branching organs.


Assuntos
Néfrons/embriologia , Organogênese , Animais , Proliferação de Células , Embrião de Mamíferos/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Camundongos , Néfrons/citologia , Ureter/embriologia
17.
Elife ; 72018 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-30516471

RESUMO

A normal endowment of nephrons in the mammalian kidney requires a balance of nephron progenitor self-renewal and differentiation throughout development. Here, we provide evidence for a novel action of ureteric branch tip-derived Wnt11 in progenitor cell organization and interactions within the nephrogenic niche, ultimately determining nephron endowment. In Wnt11 mutants, nephron progenitors dispersed from their restricted niche, intermixing with interstitial progenitors. Nephron progenitor differentiation was accelerated, kidneys were significantly smaller, and the nephron progenitor pool was prematurely exhausted, halving the final nephron count. Interestingly, RNA-seq revealed no significant differences in gene expression. Live imaging of nephron progenitors showed that in the absence of Wnt11 they lose stable attachments to the ureteric branch tips, continuously detaching and reattaching. Further, the polarized distribution of several markers within nephron progenitors is disrupted. Together these data highlight the importance of Wnt11 signaling in directing nephron progenitor behavior which determines a normal nephrogenic program.


Assuntos
Polaridade Celular/genética , Regulação da Expressão Gênica no Desenvolvimento , Néfrons/metabolismo , Organogênese/genética , Células-Tronco/metabolismo , Proteínas Wnt/genética , Animais , Diferenciação Celular , Movimento Celular , Embrião de Mamíferos , Feminino , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Queratina-8/genética , Queratina-8/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Néfrons/citologia , Néfrons/crescimento & desenvolvimento , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transdução de Sinais , Células-Tronco/citologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Wnt/metabolismo
18.
J Mol Biol ; 358(2): 532-45, 2006 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-16529770

RESUMO

B-box domains are a defining feature of the tripartite RBCC (RING, B-box, coiled-coil) or TRIM proteins, many of which are E3 ubiquitin ligases. However, little is known about the biological function of B-boxes. In some RBCC/TRIM proteins there is only a single B-box (type 2) domain, while others have both type 1 and type 2 B-box domains in tandem adjacent to their RING domain. These two types of B-boxes share little sequence similarity, except the presence of cysteine and histidine residues: eight in most B-box1 domains and seven in B-box2 domains. We report here the high-resolution solution structure of the first B-box1 domain (from the human RBCC protein, MID1) based on 670 nuclear Overhauser effect (NOE)-derived distance restraints, 12 hydrogen bonds, and 44 dihedral angles. The domain consists of a three-turn alpha-helix, two short beta-strands, and three beta-turns, encompassing Val117 to Pro164, which binds two zinc atoms. One zinc atom is coordinated by cysteine residues 119, 122, 142, 145, while cysteine 134, 137 and histidine 150, 159 coordinate the other. This topology is markedly different from the only other B-box structure reported; that of a type 2 B-box from Xenopus XNF7, which binds a single zinc atom. Of note, the B-box1 structure closely resembles the folds of the RING, ZZ and U-box domains of E3 and E4 ubiquitin enzymes, raising the possibility that the B-box1 domain either has E3 activity itself or enhances the activity of RING type E3 ligases (i.e. confers E4 enzyme activity). The structure of the MID1 B-box1 also reveals two potential protein interaction surfaces. One of these is likely to provide the binding interface for Alpha 4 that is required for the localized turnover of the catalytic subunit of PP2A, the major Ser/Thr phosphatase.


Assuntos
Proteínas dos Microtúbulos/química , Proteínas Nucleares/química , Fatores de Transcrição/química , Zinco/metabolismo , Sequência de Aminoácidos , Humanos , Proteínas dos Microtúbulos/genética , Proteínas dos Microtúbulos/metabolismo , Dados de Sequência Molecular , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Estrutura Terciária de Proteína , Homologia de Sequência de Aminoácidos , Soluções , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Dedos de Zinco
19.
Results Probl Cell Differ ; 60: 233-256, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28409348

RESUMO

The kidney develops as an outgrowth of the epithelial nephric duct known as the ureteric bud, in a position specified by a range of rostral and caudal factors which serve to ensure two kidneys form in the appropriate positions in the body. At its simplest level, kidney development can be viewed as the process by which this single bud then undergoes a process of arborisation to form a complex connected network of ducts which will serve to drain urine from the nephrons in the adult organ. The process of bud elaboration is dictated by factors expressed by both the bud itself and by surrounding cells of the metanephric mesenchyme which control cell division and bifurcation. These cells play two critical roles. Firstly, they potentiate the ongoing elaboration of the ureteric tree: remove them and branching ceases. Secondly, they harbour progenitor cells which are fated to undergo their own process of tubulogenesis to form the nephrons of the adult organ. In this chapter, we will discuss how the ureteric bud arises in the developing embryo, how it undergoes branching, how we can measure and study this process and finally the likely relevance that this process has for our understanding of congenital and acquired kidney disease.


Assuntos
Rim/embriologia , Morfogênese/fisiologia , Animais , Humanos , Processamento de Imagem Assistida por Computador
20.
Nat Rev Nephrol ; 12(12): 754-767, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27818506

RESUMO

The mammalian kidney develops from a simple epithelial bud to an arborized network of tubules, which are fated to form the ureter, renal pelvis and collecting ducts. This process of ductal elaboration is achieved through an ancient developmental mechanism known as branching morphogenesis that is widely employed in glandular organs, the vasculature and lungs. It breaks up large solid tissues facilitating secretion, excretion and gas exchange, depending on the tissue. In the kidney, growth of the ureteric bud is driven by interactions between progenitor cells in the tips of the epithelial tree and their mesenchymal 'caps'. The cells of the cap mesenchyme give rise to nephrons; therefore, the interaction between these two cell populations is likely to be a critical driver of nephron number, which is determined during gestation. These cellular interactions are potentially affected by genetic mutations (congenital kidney diseases) and by changes in the fetal environment. Understanding the aetiology of congenital and acquired kidney diseases therefore requires a full appreciation of the processes involved in establishing the cellular architecture of the kidney and of the factors that affect the commitment of progenitor cells to form nephrons.


Assuntos
Nefropatias/embriologia , Rim/embriologia , Morfogênese , Organogênese , Animais , Humanos , Camundongos , Morfogênese/genética , Morfogênese/fisiologia , Néfrons/embriologia , Organogênese/genética , Organogênese/fisiologia , Células-Tronco , Ureter/embriologia
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