RESUMEN
Vitamin D plays an important pleiotropic role in maintaining global homeostasis of the human body. Its functions go far beyond skeletal health, playing a crucial role in a plethora of cellular functions, as well as in extraskeletal health, ensuring the proper functioning of multiple human organs, including the skin. Genes from the Grainyhead-like (GRHL) family code for transcription factors necessary for the development and maintenance of various epithelia. Even though they are involved in many processes regulated by vitamin D, a direct link between vitamin D-mediated cellular pathways and GRHL genes has never been described. We employed various bioinformatic methods, quantitative real-time PCR, chromatin immunoprecipitation, reporter gene assays, and calcitriol treatments to investigate this issue. We report that the vitamin D receptor (VDR) binds to a regulatory region of the Grainyhead-like 1 (GRHL1) gene and regulates its expression. Ectopic expression of VDR and treatment with calcitriol alters the expression of the GRHL1 gene. The evidence presented here indicates a role of VDR in the regulation of expression of GRHL1 and correspondingly a role of GRHL1 in mediating the actions of vitamin D.
Asunto(s)
Regulación de la Expresión Génica , Receptores de Calcitriol , Factores de Transcripción , Receptores de Calcitriol/genética , Receptores de Calcitriol/metabolismo , Humanos , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Regulación de la Expresión Génica/efectos de los fármacos , Calcitriol/farmacología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Unión Proteica , Regiones Promotoras Genéticas , Proteínas RepresorasRESUMEN
Malignant melanoma is a dangerous skin cancer, accounting for the majority of skin cancer-related deaths. Many patients with this cancer have the V600E mutation in the BRAF gene. This mutation causes constitutive activation of the MAPK/ERK signaling pathway, significantly contributing to the process of carcinogenesis. We discuss the drug design process on the example of a specific BRAF V600E inhibitor, vemurafenib. We begin with the most commonly used drug design methods. The second part of the article focuses on vemurafenib. We analyze the invention of this BRAF V600E inhibitor and its analogue as well as the course of three stages of clinical trials. Then we provide information about other popular drugs for malignant melanoma, i.e. dacarbazine, ipilimumab and dabrafenib, and about the advantages of therapy with the simultaneous use of two inhibitors. Finally, we briefly discuss the role of artificial intelligence in the future of drug design.
Asunto(s)
Antineoplásicos , Melanoma , Neoplasias Cutáneas , Humanos , Vemurafenib/uso terapéutico , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/uso terapéutico , Inteligencia Artificial , Indoles , Sulfonamidas/farmacología , Sulfonamidas/uso terapéutico , Melanoma/tratamiento farmacológico , Melanoma/genética , Melanoma/patología , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Inhibidores de Proteínas Quinasas/farmacología , Mutación , Resistencia a Antineoplásicos , Melanoma Cutáneo MalignoRESUMEN
Known for a long time, well-tested substances are still finding new applications in science, medicine and industry. This is a popular and cost-effective strategy because, when searching for new applications, effective methods of their large-scale production and pharmacological activity, and the results of pharmacokinetic and toxicological studies are usually already known. Tamoxifen is known mainly as a drug used in the treatment of estrogen receptor-dependent breast cancer. Despite the discovery of this effective and profitable property many years ago and the constant expansion of related applications and patents, completely new ways of using tamoxifen and its derivatives in various fields continue to appear, and the number of patents for novel applications unrelated to breast cancer remains high. The aim of this article is to illustrate drug repositioning on the example of tamoxifen and to bring the ever-developing story of discoveries related to it to a wider audience.
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Neoplasias de la Mama , Tamoxifeno , Humanos , Femenino , Tamoxifeno/farmacología , Tamoxifeno/uso terapéutico , Reposicionamiento de Medicamentos , Neoplasias de la Mama/tratamiento farmacológico , Receptores de Estrógenos/uso terapéuticoRESUMEN
This Special Issue comprises three original studies and five review articles [...].
Asunto(s)
Neoplasias , Factores de Transcripción , Humanos , Neoplasias/genética , Factores de Transcripción/genéticaRESUMEN
Grainyhead-like (GRHL) factors are essential, highly conserved transcription factors (TFs) that regulate processes common to both natural cellular behaviours during embryogenesis, and de-regulation of growth and survival pathways in cancer. Serving to drive the transcription, and therefore activation of multiple co-ordinating pathways, the three GRHL family members (GRHL1-3) are a critical conduit for modulating the molecular landscape that guides cellular decision-making processes during proliferation, epithelial-mesenchymal transition (EMT) and migration. Animal models and in vitro approaches harbouring GRHL loss or gain-of-function are key research tools to understanding gene function, which gives confidence that resultant phenotypes and cellular behaviours may be translatable to humans. Critically, identifying and characterising the target genes to which these factors bind is also essential, as they allow us to discover and understand novel genetic pathways that could ultimately be used as targets for disease diagnosis, drug discovery and therapeutic strategies. GRHL1-3 and their transcriptional targets have been shown to drive comparable cellular processes in Drosophila, C. elegans, zebrafish and mice, and have recently also been implicated in the aetiology and/or progression of a number of human congenital disorders and cancers of epithelial origin. In this review, we will summarise the state of knowledge pertaining to the role of the GRHL family target genes in both development and cancer, primarily through understanding the genetic pathways transcriptionally regulated by these factors across disparate disease contexts.
Asunto(s)
Proteínas de Unión al ADN , Neoplasias , Proteínas Represoras/metabolismo , Animales , Caenorhabditis elegans/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Drosophila/metabolismo , Transición Epitelial-Mesenquimal/genética , Ratones , Neoplasias/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Pez Cebra/genética , Pez Cebra/metabolismoRESUMEN
Krüppel-like factor 4 (KLF4) is a transcription factor highly conserved in evolution. It is particularly well known for its role in inducing pluripotent stem cells. In addition, KLF4 plays many roles in cancer. The results of most studies suggest that KLF4 is a tumor suppressor. However, the functioning of KLF4 is regulated at many levels. These include regulation of transcription, alternative splicing, miRNA, post-translational modifications, subcellular localization, protein stability and interactions with other molecules. Simple experiments aimed at assaying transcript levels or protein levels fail to address this complexity and thus may deliver misleading results. Tumor subtypes are also important; for example, in prostate cancer KLF4 is highly expressed in indolent tumors where it impedes tumor progression, while it is absent from aggressive prostate tumors. KLF4 is important in regulating response to many known drugs, and it also plays a role in tumor microenvironment. More and more information is available about upstream regulators, downstream targets and signaling pathways associated with the involvement of KLF4 in cancer. Furthermore, KLF4 performs critical function in the overall regulation of tissue homeostasis, cellular integrity, and progression towards malignancy. Here we summarize and analyze the latest findings concerning this fascinating transcription factor.
Asunto(s)
Factores de Transcripción de Tipo Kruppel/metabolismo , Neoplasias/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Factor 4 Similar a Kruppel , Masculino , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genéticaRESUMEN
The mammalian Grainyhead-like 3 (GRHL3) transcription factor is essential for epithelial development and plays a protective role against squamous cell carcinoma of the skin and of the oral cavity. A single nucleotide polymorphism (SNP) in GRHL3, rs141193530 (p.P455A), is associated with non-melanoma skin cancer in human patients. Moreover, it is known that this SNP, as well as another variant, rs41268753 (p.T454M), are associated with nonsyndromic cleft palate and that rs41268753 negatively affects GRHL3 transcriptional activity. These SNPs are located in adjacent codons of the GRHL3 gene, and the occurrence of either SNP abolishes a putative threonine-proline phosphorylation motif at T454 in the encoded protein. The role of phosphorylation in regulating mammalian GRHL function is currently unknown. In this work we show that GRHL3 is phosphorylated at several residues in a human keratinocyte cell line, among them at T454. This site is essential for the full transcriptional activity of GRHL3. The T454 residue is phosphorylated by p38 MAPK in vitro and activation of p38 signaling in cells causes an increase in GRHL3 activity. The regulation of GRHL3 function by this pathway is dependent on T454, as the substitution of T454 with methionine inhibits the activation of GRHL3. Taken together, our results show that T454 is one of the phosphorylated residues in GRHL3 in keratinocytes and this residue is important for the upregulation of GRHL3 transcriptional activity by the p38 pathway.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Transducción de Señal , Treonina/metabolismo , Factores de Transcripción/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Secuencia de Aminoácidos , Animales , Línea Celular , Fisura del Paladar/genética , Fisura del Paladar/metabolismo , Proteínas de Unión al ADN/análisis , Proteínas de Unión al ADN/genética , Humanos , Queratinocitos/metabolismo , Fosforilación , Mutación Puntual , Polimorfismo de Nucleótido Simple , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Treonina/análisis , Treonina/genética , Factores de Transcripción/análisis , Factores de Transcripción/genéticaRESUMEN
Epidermal growth factor receptor (EGFR) is a central transmitter of mitogenic signals in epithelial cells; enhanced EGFR activity is observed in many tumors of epithelial origin. S100A6 is a small calcium-binding protein, characteristic mainly of epithelial cells and fibroblasts, strongly implicated in cell proliferation and upregulated in tumors. In this study, using biochemical assays along with immunohistochemical and immunocytochemical analysis of organotypic and standard cultures of HaCaT keratinocytes with S100A6 overexpression or knock-down, we have examined the effect of S100A6 on EGFR activity and downstream signaling. We found that HaCaT cells overexpressing S100A6 had enhanced EGFR, phospho EGFR, and phospho extracellular signal-regulated kinase 1/2 (pERK1/2) staining intensity and level coupled to higher signal transducer and activator of transcription 3 (STAT3) activity. Conversely, S100A6 knockdown cells had impaired EGFR signaling that could be enhanced by addition of recombinant S100A6 to the culture media. Altogether the results show that S100A6 may exert its proproliferative effects through activating EGFR.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Queratinocitos/metabolismo , Proteína A6 de Unión a Calcio de la Familia S100/metabolismo , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/genética , Línea Celular , Proliferación Celular/fisiología , Receptores ErbB/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Queratinocitos/citología , Sistema de Señalización de MAP Quinasas , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteína A6 de Unión a Calcio de la Familia S100/antagonistas & inhibidores , Proteína A6 de Unión a Calcio de la Familia S100/genética , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador alfa/metabolismoRESUMEN
BACKGROUND: The Grainyhead-like (GRHL) transcription factors have been linked to many different types of cancer. However, no previous study has attempted to investigate potential correlations in expression of different GRHL genes in this context. Furthermore, there is very little information concerning damaging mutations and/or single nucleotide polymorphisms in GRHL genes that may be linked to cancer. METHODS: DNA and RNA were extracted from human non-melanoma skin cancers (NMSC) and adjacent normal tissues (n = 33 pairs of samples). The expression of GRHL genes was measured by quantitative real time PCR. Regulation of GRHL expression by miRNA was studied using cell transfection methods and dual-luciferase reporter system. Targeted deep sequencing of GRHL genes in tumor samples and control tissues were employed to search for mutations and single nucleotide polymorphisms. Single marker rs141193530 was genotyped with pyrosequencing in additional NMSC replication cohort (n = 176). Appropriate statistical and bioinformatic methods were used to analyze and interpret results. RESULTS: We discovered that the expression of two genes - GRHL1 and GRHL3 - is reduced in a coordinated manner in tumor samples, in comparison to the control healthy skin samples obtained from the same individuals. It is possible that both GRHL1 and GRHL3 are regulated, at least to some extent, by different strands of the same oncogenic microRNA - miR-21, what would at least partially explain observed correlation. No de novo mutations in the GRHL genes were detected in the examined tumor samples. However, some single nucleotide polymorphisms in the GRHL genes occur at significantly altered frequencies in the examined group of NMSC patients. CONCLUSIONS: Non-melanoma skin cancer growth is accompanied by coordinated reduced expression of epidermal differentiation genes: GRHL1 and GRHL3, which may be regulated by miR-21-3p and -5p, respectively. Some potentially damaging single nucleotide polymorphisms in GRHL genes occur with altered frequencies in NMSC patients, and they may in particular impair the expression of GRHL3 gene or functioning of encoded protein. The presence of these polymorphisms may indicate an increased risk of NMSC development in affected people.
Asunto(s)
Proteínas de Unión al ADN/genética , MicroARNs/genética , Proteínas Represoras/genética , Neoplasias Cutáneas/genética , Factores de Transcripción/genética , Diferenciación Celular/genética , Epidermis/crecimiento & desarrollo , Epidermis/patología , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Mutación , Polimorfismo de Nucleótido Simple/genética , Neoplasias Cutáneas/patologíaRESUMEN
In recent years, the TFCP2 (transcription factor cellular promoter 2)/TFCP2L1 (TFCP2-like 1)/UBP1 (upstream binding protein 1) subfamily of transcription factors has been attracting increasing attention in the scientific community. These factors are very important in cancer, Alzheimer's disease, and other human conditions, and they can be attractive targets for drug development. However, the interpretation of experimental results is complicated, as in principle, any of these factors could substitute for the lack of another. Thus, studying their hitherto little known functions should enhance our understanding of mechanisms of their functioning, and analogous mechanisms might govern their functioning in medically relevant contexts. For example, there are numerous parallels between placental development and cancer growth; therefore, investigating the roles of TFCP2, TFCP2L1, and UBP1 in the placenta may help us better understand their functioning in cancer, as is evidenced by the studies of various other proteins and pathways. Our review article aims to call the attention of the scientific community to these neglected functions, and encourage further research in this field. Here, we present a systematic review of current knowledge of the TFCP2/TFCP2L1/UBP1 subfamily in reproduction, embryonic development, renal function, blood-pressure regulation, brain function, and other processes, where their involvement has not been studied much until now.
Asunto(s)
Proteínas de Unión al ADN/fisiología , Regulación de la Expresión Génica , Factores de Transcripción/fisiología , Presión Sanguínea/genética , Encéfalo/metabolismo , Encéfalo/fisiología , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Desarrollo Embrionario/genética , Humanos , Riñón/metabolismo , Riñón/fisiología , Reproducción/genética , Factores de Transcripción/química , Factores de Transcripción/metabolismoRESUMEN
The Ca2+-binding protein, S100A6, belongs to the S100 family. Binding of Ca2+ induces a conformational change, which causes an increase in the overall S100A6 hydrophobicity and allows it to interact with many targets. S100A6 is expressed in different normal tissues and in many tumors. Up to now it has been shown that S100A6 is involved in cell proliferation, cytoskeletal dynamics and tumorigenesis, and that it might have some extracellular functions. In this review, we summarize novel discoveries concerning S100A6 targets, its involvement in cellular signaling pathways, and presence in stem/progenitor cells, extracellular matrix and body fluids of diseased patients.
Asunto(s)
Proteínas S100/metabolismo , Animales , Líquidos Corporales/metabolismo , Calcio/metabolismo , Matriz Extracelular/metabolismo , Humanos , Transducción de Señal , Células Madre/metabolismoRESUMEN
The involvement of Grainyhead-like (GRHL) transcription factors in various cancers is well documented. However, little is known about their role in clear cell renal cell carcinoma (ccRCC). We discovered that the expression of two of these factors-GRHL1 and GRHL2-are downregulated in ccRCC samples, and their expression is correlated with the expression of VHL gene. This suggests a functional link between the GRHL transcription factors and one of the best known tumor suppressors. Although the GRHL genes are not mutated in ccRCC, some of the single nucleotide polymorphisms in these genes may indicate an increased risk of ccRCC development and/or may allow to assess patients' prognoses and predict their responses to various forms of therapy. Silencing of GRHL2 expression in non-tumorigenic kidney cell line results in increased cell proliferation, increased resistance to apoptosis, as well as changes in the levels of selected proteins involved in the pathogenesis of ccRCC. These changes support the potential role for GRHL2 as a suppressor of ccRCC.
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Carcinoma de Células Renales/genética , Proteínas de Unión al ADN/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/genética , Riñón/patología , Factores de Transcripción/genética , Carcinoma de Células Renales/patología , Línea Celular Tumoral , Femenino , Silenciador del Gen , Humanos , Riñón/metabolismo , Neoplasias Renales/patología , Masculino , Polimorfismo de Nucleótido Simple , Proteínas Represoras/genéticaRESUMEN
Phosphorylation and dephosphorylation play a fundamental role in most signaling pathways, as these processes can directly regulate various aspects of protein function. It is estimated that there are about 100,000 potential phosphorylation sites in proteins encoded by the human genome and about 30-50% of all proteins in the cell can be phosphorylated, which is directly related to the functions they perform. To determine whether a given protein is phosphorylated, any changes in its mobility caused by this modification are examined during PAGE electrophoresis. Concurrently, tandem mass spectrometry (MS/MS) allows to identify specific phosphorylation sites. The next step involves the prediction (using in silico analysis) which kinases can phosphorylate a specific site in the given protein. Then, in order to verify the information obtained from databases, in vitro and/or in vivo experiments are carried out.
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Proteínas/metabolismo , Transducción de Señal , Secuencia de Aminoácidos , Electroforesis en Gel de Poliacrilamida , Humanos , Fosforilación , Espectrometría de Masas en TándemRESUMEN
The isthmic organiser located at the midbrain-hindbrain boundary (MHB) is the crucial developmental signalling centre responsible for patterning mesencephalic and metencephalic regions of the vertebrate brain. Formation and maintenance of the MHB is characterised by a hierarchical program of gene expression initiated by fibroblast growth factor 8 (Fgf8), coupled with cellular morphogenesis, culminating in the formation of the tectal-isthmo-cerebellar structures. Here, we show in zebrafish that one orthologue of the transcription factor grainy head-like 2 (Grhl2), zebrafish grhl2b plays a central role in both MHB maintenance and folding by regulating two distinct, non-linear pathways. Loss of grhl2b expression induces neural apoptosis and extinction of MHB markers, which are rescued by re-expression of engrailed 2a (eng2a), an evolutionarily conserved target of the Grhl family. Co-injection of sub-phenotypic doses of grhl2b and eng2a morpholinos reproduces the apoptosis and MHB marker loss, but fails to substantially disrupt formation of the isthmic constriction. By contrast, a novel direct grhl2b target, spec1, identified by phylogenetic analysis and confirmed by ChIP, functionally cooperates with grhl2b to induce MHB morphogenesis, but plays no role in apoptosis or maintenance of MHB markers. Collectively, these data show that MHB maintenance and morphogenesis are dissociable events regulated by grhl2b through diverse transcriptional targets.
Asunto(s)
Proteínas Portadoras/metabolismo , Mesencéfalo/crecimiento & desarrollo , Morfogénesis , Rombencéfalo/crecimiento & desarrollo , Factores de Transcripción/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/crecimiento & desarrollo , Animales , Apoptosis , Proteínas Portadoras/genética , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas de Homeodominio/biosíntesis , Proteínas de Homeodominio/genética , Mesencéfalo/metabolismo , Morfolinos/genética , Proteínas del Tejido Nervioso/biosíntesis , Proteínas del Tejido Nervioso/genética , Filogenia , Rombencéfalo/metabolismo , Transducción de Señal , Factores de Transcripción/genética , Pez Cebra/genética , Proteínas de Pez Cebra/genéticaRESUMEN
The Grainy head-like 3 (Grhl3) gene encodes a transcription factor that plays essential roles in epidermal morphogenesis during embryonic development, with deficient mice exhibiting failed skin barrier formation, defective wound repair, and loss of eyelid fusion. Despite sharing significant sequence homology, overlapping expression patterns, and an identical core consensus DNA binding site, the other members of the Grhl family (Grhl1 and -2) fail to compensate for the loss of Grhl3 in these processes. Here, we have employed diverse genetic models, coupled with biochemical studies, to define the inter-relationships of the Grhl factors in epidermal development. We show that Grhl1 and Grhl3 have evolved complete functional independence, as evidenced by a lack of genetic interactions in embryos carrying combinations of targeted alleles of these genes. In contrast, compound heterozygous Grhl2/Grhl3 embryos displayed failed wound repair, and loss of a single Grhl2 allele in Grhl3-null embryos results in fully penetrant eyes open at birth. Expression of Grhl2 from the Grhl3 locus in homozygous knock-in mice corrects the wound repair defect, but these embryos still display a complete failure of skin barrier formation. This functional dissociation is due to unexpected differences in target gene specificity, as both GRHL2 and GRHL3 bind to and regulate expression of the wound repair gene Rho GEF 19, but regulation of the barrier forming gene, Transglutaminase 1 (TGase1), is unique to GRHL3. Our findings define the mechanisms underpinning the unique and cooperative roles of the Grhl genes in epidermal development.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Epidermis/embriología , Morfogénesis/fisiología , Fenotipo , Proteínas Represoras/metabolismo , Factores de Transcripción/metabolismo , Animales , Cartilla de ADN/genética , Proteínas de Unión al ADN/genética , Ensayo de Cambio de Movilidad Electroforética , Epidermis/ultraestructura , Técnicas de Sustitución del Gen , Inmunohistoquímica , Hibridación in Situ , Ratones , Microscopía Electrónica de Rastreo , Proteínas Represoras/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Transcripción/genética , Transglutaminasas/metabolismo , Cicatrización de Heridas/fisiologíaRESUMEN
In Drosophila, the grainy head (grh) gene plays a range of key developmental roles through the regulation of members of the cadherin gene family. We now report that mice lacking the grh homologue grainy head-like 1 (Grhl1) exhibit hair and skin phenotypes consistent with a reduction in expression of the genes encoding the desmosomal cadherin, desmoglein 1 (Dsg1). Grhl1-null mice show an initial delay in coat growth, and older mice exhibit hair loss as a result of poor anchoring of the hair shaft in the follicle. The mice also develop palmoplantar keratoderma, analogous to humans with DSG1 mutations. Sequence analysis, DNA binding, and chromatin immunoprecipitation experiments demonstrate that the human and mouse Dsg1 promoters are direct targets of GRHL1. Ultrastructural analysis reveals reduced numbers of abnormal desmosomes in the interfollicular epidermis. These findings establish GRHL1 as an important regulator of the Dsg1 genes in the context of hair anchorage and epidermal differentiation, and suggest that cadherin family genes are key targets of the grainy head-like genes across 700 million years of evolution.
Asunto(s)
Cadherinas Desmosómicas/genética , Desmosomas/genética , Regulación de la Expresión Génica/fisiología , Proteínas Represoras/genética , Animales , Diferenciación Celular/genética , Desmogleína 1/biosíntesis , Desmogleína 1/genética , Cadherinas Desmosómicas/antagonistas & inhibidores , Cadherinas Desmosómicas/biosíntesis , Desmosomas/metabolismo , Cabello/anomalías , Folículo Piloso/embriología , Folículo Piloso/metabolismo , Ratones , Ratones Noqueados , Proteínas Represoras/antagonistas & inhibidores , Proteínas Represoras/biosíntesisRESUMEN
The LSF/Grainyhead family of transcription factors consists of proteins whose structure and functions have been preserved in the course of eukaryotic evolution--from primitive unicellular life forms to complex multicellular organisms. In the latter, these factors display tissue specificity and are active mainly in the covering epithelium. The roles of GRH factors are associated with regulation of expression of genes essential for correct differentiation and functioning of the epithelia of ectodermal origin. The Grh gene expression profiles are diverse and variable, especially during embryonic development. Research on the role of GRHL transcription factors is carried out on cellular and organismal level. In experimental animals, aberrant Grh gene expression leads to many diseases, including failure of epidermal wound healing and neural tube defects. Changes of these genes' expression levels are also linked to carcinogenesis. GRHL transcription factors participate in signaling pathways involved in cellular proliferation and apoptosis.
Asunto(s)
Epidermis/metabolismo , Factores de Transcripción/metabolismo , Animales , Apoptosis/genética , Proliferación Celular , Proteínas de Unión al ADN/metabolismo , Modelos Animales de Enfermedad , Epidermis/embriología , Expresión Génica , Humanos , Neoplasias/genética , Defectos del Tubo Neural/genética , Factores de Transcripción/genética , Cicatrización de Heridas/genéticaRESUMEN
Primary neurulation in mammals has been defined by distinct anatomical closure sites, at the hindbrain/cervical spine (closure 1), forebrain/midbrain boundary (closure 2), and rostral end of the forebrain (closure 3). Zones of neurulation have also been characterized by morphologic differences in neural fold elevation, with non-neural ectoderm-induced formation of paired dorso-lateral hinge points (DLHP) essential for neural tube closure in the cranial and lower spinal cord regions, and notochord-induced bending at the median hinge point (MHP) sufficient for closure in the upper spinal region. Here we identify a unifying molecular basis for these observations based on the function of the non-neural ectoderm-specific Grainy head-like genes in mice. Using a gene-targeting approach we show that deletion of Grhl2 results in failed closure 3, with mutants exhibiting a split-face malformation and exencephaly, associated with failure of neuro-epithelial folding at the DLHP. Loss of Grhl3 alone defines a distinct lower spinal closure defect, also with defective DLHP formation. The two genes contribute equally to closure 2, where only Grhl gene dosage is limiting. Combined deletion of Grhl2 and Grhl3 induces severe rostral and caudal neural tube defects, but DLHP-independent closure 1 proceeds normally in the upper spinal region. These findings provide a molecular basis for non-neural ectoderm mediated formation of the DLHP that is critical for complete neuraxis closure.
Asunto(s)
Proteínas de Unión al ADN/genética , Tubo Neural/embriología , Factores de Transcripción/genética , Animales , Proteínas de Unión al ADN/metabolismo , Embrión de Mamíferos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Transgénicos , Tubo Neural/crecimiento & desarrollo , Defectos del Tubo Neural/genética , Factores de Transcripción/metabolismoRESUMEN
The neural tube defects (NTDs) spina bifida and anencephaly are widely prevalent severe birth defects. The mouse mutant curly tail (ct/ct) has served as a model of NTDs for 50 years, even though the responsible genetic defect remained unrecognized. Here we show by gene targeting, mapping and genetic complementation studies that a mouse homolog of the Drosophila grainyhead (grh) gene, grainyhead-like-3 (Grhl3), is a compelling candidate for the gene underlying the curly tail phenotype. The NTDs in Grhl3-null mice are more severe than those in the curly tail strain, as the Grhl3 alleles in ct/ct mice are hypomorphic. Spina bifida in ct/ct mice is folate resistant, but its incidence can be markedly reduced by maternal inositol supplementation periconceptually. The NTDs in Grhl3-/- embryos are also folate resistant, but unlike those in ct/ct mice, they are resistant to inositol. These findings suggest that residual Grhl3 expression in ct/ct mice may be required for inositol rescue of folate-resistant NTDs.
Asunto(s)
Proteínas de Unión al ADN/deficiencia , Ácido Fólico/farmacología , Inositol/farmacología , Defectos del Tubo Neural/etiología , Defectos del Tubo Neural/prevención & control , Factores de Transcripción/deficiencia , Animales , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Resistencia a Medicamentos , Epitelio/patología , Femenino , Humanos , Ratones , Ratones Noqueados , Ratones Mutantes Neurológicos , Defectos del Tubo Neural/genética , Defectos del Tubo Neural/patología , Proteínas Nucleares , Fenotipo , Embarazo , Factores de Transcripción/genética , Factores de Transcripción/fisiologíaRESUMEN
Genes from the Grainyhead-like (GRHL) family code for transcription factors necessary for the development and maintenance of various epithelia. These genes are also very important in the development of many types of cancer. However, little is known about the regulation of expression of GRHL genes. Previously, there were no systematic analyses of the promoters of GRHL genes or transcription factors that bind to these promoters. Here we report that the Krüppel-like factor 4 (KLF4) and the paired box 5 factor (PAX5) bind to the regulatory regions of the GRHL genes and regulate their expression. Ectopic expression of KLF4 or PAX5 alters the expression of GRHL genes. In KLF4-overexpressing HEK293 cells, the expression of GRHL1 and GRHL3 genes was upregulated by 32% and 60%, respectively, whereas the mRNA level of GRHL2 gene was lowered by 28% when compared to the respective controls. The levels of GRHL1 and GRHL3 expression were decreased by 30% or 33% in PAX5-overexpressing HEK293 cells. The presence of minor frequency allele of single nucleotide polymorphism rs115898376 in the promoter of the GRHL1 gene affected the binding of KLF4 to this site. The evidence presented here suggests an important role of KLF4 and PAX5 in the regulation of expression of GRHL1-3 genes.