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1.
DNA Cell Biol ; 38(11): 1323-1337, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31536386

RESUMO

Our previous study has indicated that the parathyroid hormone type 1 receptor (PTHR1) may play important roles in development and progression of osteosarcoma (OS) by regulating Wnt, angiogenesis, and inflammation pathway genes. The goal of this study was to further illuminate the roles of PTHR1 in OS by investigating upstream regulation mechanisms (including microRNA [miRNA] and transcription factors [TFs]) of crucial genes. The microarray dataset GSE46861 was downloaded from the Gene Expression Omnibus database, in which six tumors with short hairpin RNA (shRNA) PTHR1 knockdown (PTHR1.358) and six tumors with shRNA control knockdown (Ren.1309) were collected from mice. Differentially expressed genes (DEGs) between PTHR1.358 and Ren.1309 were identified using the linear models for microarray data (LIMMA) method, and then the miRNA-TF-mRNA regulatory network was constructed using data from corresponding databases, followed by module analysis, to screen crucial regulatory relationships. OS-related human miRNAs were extracted from the curated Osteosarcoma Database. Gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched using the Database for Annotation, Visualization, and Integrated Discovery (DAVID) tool. As a result, the miRNA-TF-mRNA regulatory network, including 1049 nodes (516 miRNA, 25 TFs, and 508 DEGs) and 15942 edges (interaction relationships, such as Pparg-Abca1 and miR-590-3p-AXIN2), was constructed, from which three significant modules were extracted and modules 2 and 3 contained interactions between miRNAs/TFs and DEGs such as miR-103-3p-AXIN2, miR-124-3p-AR-Tgfb1i1, and miR-27a-3p-PPARG-Abca1. miR-27a-3p was a known miRNA associated with OS. Abca1, AR, and miR-124-3p were hub genes in the miRNA-TF-mRNA network. Tgfb1i1 was involved in cell proliferation, Abca1 participated in the cholesterol metabolic process, and AXIN2 was associated with the canonical Wnt signaling pathway. Furthermore, we also confirmed upregulation of miR-590-3p and downregulation of AXIN2 in the mouse OS cell line K7M2-WT transfected with PTHR1 shRNA. In conclusion, PTHR1 may play important roles in progression of OS by activating miR-124-3p-AR-Tgfb1i1, miR-27a-3p-PPARG-Abca1, and miR-103/590-3p-AXIN2 axes.


Assuntos
Neoplasias Ósseas , Osteossarcoma , Receptor Tipo 1 de Hormônio Paratireóideo/fisiologia , Transportador 1 de Cassete de Ligação de ATP/genética , Transportador 1 de Cassete de Ligação de ATP/fisiologia , Animais , Proteína Axina/genética , Proteína Axina/fisiologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/fisiologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/fisiologia , Progressão da Doença , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Células HEK293 , Humanos , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/fisiologia , Osteossarcoma/genética , Osteossarcoma/patologia , PPAR gama/genética , PPAR gama/fisiologia , Receptor Tipo 1 de Hormônio Paratireóideo/genética , Receptores Androgênicos/genética , Receptores Androgênicos/fisiologia , Transdução de Sinais/genética , Células Tumorais Cultivadas
2.
Nature ; 574(7777): 259-263, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31554973

RESUMO

Chikungunya virus (CHIKV) is a re-emerging alphavirus that is transmitted to humans by mosquito bites and causes musculoskeletal and joint pain1,2. Despite intensive investigations, the human cellular factors that are critical for CHIKV infection remain unknown, hampering the understanding of viral pathogenesis and the development of anti-CHIKV therapies. Here we identified the four-and-a-half LIM domain protein 1 (FHL1)3 as a host factor that is required for CHIKV permissiveness and pathogenesis in humans and mice. Ablation of FHL1 expression results in the inhibition of infection by several CHIKV strains and o'nyong-nyong virus, but not by other alphaviruses and flaviviruses. Conversely, expression of FHL1 promotes CHIKV infection in cells that do not normally express it. FHL1 interacts directly with the hypervariable domain of the nsP3 protein of CHIKV and is essential for the replication of viral RNA. FHL1 is highly expressed in CHIKV-target cells and is particularly abundant in muscles3,4. Dermal fibroblasts and muscle cells derived from patients with Emery-Dreifuss muscular dystrophy that lack functional FHL15 are resistant to CHIKV infection. Furthermore,  CHIKV infection  is undetectable in Fhl1-knockout mice. Overall, this study shows that FHL1 is a key factor expressed by the host that enables CHIKV infection and identifies the interaction between nsP3 and FHL1 as a promising target for the development of anti-CHIKV therapies.


Assuntos
Febre de Chikungunya/virologia , Vírus Chikungunya/patogenicidade , Fatores Celulares Derivados do Hospedeiro/metabolismo , Interações Hospedeiro-Patógeno , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/metabolismo , Proteínas Musculares/metabolismo , Animais , Células Cultivadas , Febre de Chikungunya/tratamento farmacológico , Vírus Chikungunya/efeitos dos fármacos , Vírus Chikungunya/genética , Vírus Chikungunya/crescimento & desenvolvimento , Feminino , Fibroblastos/virologia , Células HEK293 , Fatores Celulares Derivados do Hospedeiro/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Domínio LIM/deficiência , Proteínas com Domínio LIM/genética , Masculino , Camundongos , Proteínas Musculares/deficiência , Proteínas Musculares/genética , Mioblastos/virologia , Vírus O'nyong-nyong/crescimento & desenvolvimento , Vírus O'nyong-nyong/patogenicidade , Ligação Proteica , RNA Viral/biossíntese , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/metabolismo , Replicação Viral
3.
Life Sci ; 233: 116715, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31376371

RESUMO

AIMS: PDZ and LIM domain protein 4 (PDLIM4) is frequently repressed in cancer tissues. However, the expression and role of PDLIM4 in ovarian cancer has not been addressed. MAIN METHODS: In this study, we examined the expression and prognostic significance of PDLIM4 in ovarian cancer. The function of PDLIM4 in ovarian cancer cell growth, invasion, and tumorigenesis was further explored. KEY FINDINGS: PDLIM4 is downregulated in ovarian cancer compared to adjacent normal ovarian tissues. Downregulation of PDLIM4 is correlated with advanced tumor stage and lymph node metastasis. Low PDLIM4 expression is significantly associated with shorter overall survival in patients with ovarian cancer (P = 0.0136). Biologically, PDLIM4 overexpression suppresses the proliferation, colony formation, migration, and invasion of both CAOV3 and SKOV3 ovarian cancer cells, compared to empty vector-transfected cells. Consistently, in vivo data show that PDLIM4 overexpression inhibits the growth of SKOV3 xenograft tumors. Mechanistic investigation reveals that overexpression of PDLIM4 blocks the phosphorylation of STAT3 and represses STAT3-dependent transcriptional activation. Moreover, ectopic expression of PDLIM4 downregulates the expression of CCND1 and MMP9 in ovarian cancer cells. Rescue experiments demonstrate that overexpression of constitutively active STAT3 reverses PDLIM4-induced anticancer effects on ovarian cancer cells. SIGNIFICANCE: Overall, PDLIM4 downregulation is associated with aggressive tumor features and poor prognosis in ovarian cancer patients. PDLIM4 suppresses ovarian cancer cell growth and invasion by inhibiting STAT3 signaling. This study provides a potential therapeutic target for ovarian cancer.


Assuntos
Movimento Celular , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas com Domínio LIM/metabolismo , Neoplasias Ovarianas/patologia , Fator de Transcrição STAT3/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Estudos de Casos e Controles , Proteínas de Ligação a DNA/genética , Feminino , Humanos , Proteínas com Domínio LIM/genética , Metástase Linfática , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Fosforilação , Prognóstico , Fator de Transcrição STAT3/genética , Transdução de Sinais , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Forensic Sci Int ; 302: 109906, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31419596

RESUMO

The aim of this study is to determine the molecular mechanism of sudden death in a previously healthy patient. Clinical exome sequencing revealed I536T-RBM20 variant, which alters RNA splicing of TTN and is causative for dilated cardiomyopathy. Comprehensive RNA sequencing (RNA-seq) was also performed in the patient samples and the control samples. Splicing abnormality was compared in cardiac muscle and skeletal muscle. RNA-seq analysis of the cardiac and skeletal muscle showed abnormal splicing of LDB3, not of TTN. Exon 11 of LDB3 was abnormally included in the patient samples compared with the control samples. This abnormal LDB3 splicing pattern in skeletal muscle has been reported in myotonic dystrophy type 1 (DM1) patients. We, thus, confirmed that the patient had expanded CTG repeat in DMPK and the diagnosis was genetically DM1. This finding suggest that one of the molecular mechanisms of sudden cardiac death in this asymptomatic subclinical DM1 patient might be LDB3 abnormal splicing due to the CTG repeat in DMPK, rather than RBM20 variant. RNA-seq analysis is useful to determine the exact molecular diagnosis for sudden cardiac death.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Morte Súbita Cardíaca/etiologia , Proteínas com Domínio LIM/genética , Distrofia Miotônica/diagnóstico , Processamento de RNA , Doenças Assintomáticas , Éxons , Humanos , Masculino , Miotonina Proteína Quinase/genética , Análise de Sequência de RNA , Expansão das Repetições de Trinucleotídeos , Adulto Jovem
5.
Gene ; 712: 143963, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31279706

RESUMO

BACKGROUND: The aim of this study was to identify the expression of LIM and calponin-homology domains 1 (LIMCH1) in lung cancer and normal tissues, to determine the interaction between LIMCH1 and HUWE1 in regulating p53 stability. METHODS: The expression of LIMCH1 was detected by the Oncomine and Cancer Genome Atlas databases. Expression of LIMCH1 mRNA was identified using qRT-PCR. In transfected human lung cancer cells, co-immunoprecipitation experiments were performed. The mechanism that HUWE1 sustained lung cancer malignancy was verified by western blotting. The proliferation of tranfected cells was assessed by CCK-8 assay and colony formation. RESULTS: Bioinformatic data and e TCGA database suggested LIMCH1 mRNA levels in tumor tissues were down-regulated compared to tumor adjacent tissues. We found low expression of LIMCH1 mRNA in tumor sites and tumor cell line. Exogenous expression of LIMCH1 interacts with HUWE1 promotes expression of p53. Use of siRNA or shRNA against LIMCH1 resulted in decreased p53 protein levels. LIMCH1 deletion lead to enhance of p53 ubiquitination and protein expression of p53 and substrate p21, puma. Growth curve showed that LIMCH1 deletion significantly promoted the proliferation of A549 cells. CONCLUSIONS: LIMCH1 was a negative regulator and indicated a new molecular mechanism for the pathogenesis of lung cancer via modulating HUWE1 and p53.


Assuntos
Regulação Neoplásica da Expressão Gênica , Proteínas com Domínio LIM/metabolismo , Neoplasias Pulmonares/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Células A549 , Idoso , Linhagem Celular Tumoral , Proliferação de Células , Biologia Computacional , Regulação para Baixo , Feminino , Perfilação da Expressão Gênica , Humanos , Proteínas com Domínio LIM/genética , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Processamento de Proteína Pós-Traducional , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteína Supressora de Tumor p53/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
6.
N Engl J Med ; 380(20): 1918-1928, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31091373

RESUMO

BACKGROUND: In the context of kidney transplantation, genomic incompatibilities between donor and recipient may lead to allosensitization against new antigens. We hypothesized that recessive inheritance of gene-disrupting variants may represent a risk factor for allograft rejection. METHODS: We performed a two-stage genetic association study of kidney allograft rejection. In the first stage, we performed a recessive association screen of 50 common gene-intersecting deletion polymorphisms in a cohort of kidney transplant recipients. In the second stage, we replicated our findings in three independent cohorts of donor-recipient pairs. We defined genomic collision as a specific donor-recipient genotype combination in which a recipient who was homozygous for a gene-intersecting deletion received a transplant from a nonhomozygous donor. Identification of alloantibodies was performed with the use of protein arrays, enzyme-linked immunosorbent assays, and Western blot analyses. RESULTS: In the discovery cohort, which included 705 recipients, we found a significant association with allograft rejection at the LIMS1 locus represented by rs893403 (hazard ratio with the risk genotype vs. nonrisk genotypes, 1.84; 95% confidence interval [CI], 1.35 to 2.50; P = 9.8×10-5). This effect was replicated under the genomic-collision model in three independent cohorts involving a total of 2004 donor-recipient pairs (hazard ratio, 1.55; 95% CI, 1.25 to 1.93; P = 6.5×10-5). In the combined analysis (discovery cohort plus replication cohorts), the risk genotype was associated with a higher risk of rejection than the nonrisk genotype (hazard ratio, 1.63; 95% CI, 1.37 to 1.95; P = 4.7×10-8). We identified a specific antibody response against LIMS1, a kidney-expressed protein encoded within the collision locus. The response involved predominantly IgG2 and IgG3 antibody subclasses. CONCLUSIONS: We found that the LIMS1 locus appeared to encode a minor histocompatibility antigen. Genomic collision at this locus was associated with rejection of the kidney allograft and with production of anti-LIMS1 IgG2 and IgG3. (Funded by the Columbia University Transplant Center and others.).


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Variações do Número de Cópias de DNA , Rejeição de Enxerto/genética , Transplante de Rim , Proteínas com Domínio LIM/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Estudos de Coortes , Estudos de Associação Genética , Genótipo , Antígenos HLA/genética , Teste de Histocompatibilidade , Humanos , Imunoglobulina G/sangue , Proteínas com Domínio LIM/imunologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Polimorfismo de Nucleotídeo Único , Doadores de Tecidos
8.
Nat Commun ; 10(1): 2316, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31127120

RESUMO

Chromatin looping allows enhancer-bound regulatory factors to influence transcription. Large domains, referred to as topologically associated domains, participate in genome organization. However, the mechanisms underlining interactions within these domains, which control gene expression, are not fully understood. Here we report that activation of embryonic myogenesis is associated with establishment of long-range chromatin interactions centered on Pax3-bound loci. Using mass spectrometry and genomic studies, we identify the ubiquitously expressed LIM-domain binding protein 1 (Ldb1) as the mediator of looping interactions at a subset of Pax3 binding sites. Ldb1 is recruited to Pax3-bound elements independently of CTCF-Cohesin, and is necessary for efficient deposition of H3K4me1 at these sites and chromatin looping. When Ldb1 is deleted in Pax3-expressing cells in vivo, specification of migratory myogenic progenitors is severely impaired. These results highlight Ldb1 requirement for Pax3 myogenic activity and demonstrate how transcription factors can promote formation of sub-topologically associated domain interactions involved in lineage specification.


Assuntos
Cromossomos de Mamíferos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas com Domínio LIM/metabolismo , Desenvolvimento Muscular , Fator de Transcrição PAX3/metabolismo , Animais , Diferenciação Celular , Linhagem Celular , Cromatina/metabolismo , Cromossomos de Mamíferos/química , Proteínas de Ligação a DNA/genética , Embrião de Mamíferos , Feminino , Fibroblastos , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HEK293 , Histonas/metabolismo , Humanos , Proteínas com Domínio LIM/genética , Masculino , Camundongos , Camundongos Transgênicos , Células-Tronco Embrionárias Murinas
9.
Gene ; 707: 36-43, 2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-30930226

RESUMO

Muscle LIM protein (MLP/CSRP3/CRP3) is a microtubule-associated protein preferentially expressed in cardiac and skeletal muscle and has a central role during muscle development and for architectural maintenance of muscle cells. LIM-domain proteins act as both modulators and downstream targets of TGF-ß signaling, which is well documented to negatively regulate differentiation of myogenic precursor cells or myoblasts. Herein, we determined whether CSRP3 regulates chicken satellite cell proliferation and differentiation in vitro, and examined its mechanism of action by focusing on the TGF-ß signaling pathway. Interference of CSRP3 mRNA expression had no effect on the proliferation of satellite cells, but significantly inhibited satellite cell differentiation into myotubes at 24, 48, and 72 h after initiation of differentiation. However, CSRP3 overexpression did not affect the proliferation or differentiation of satellite cells. Moreover, knockdown of CSRP3 caused up-regulation of TGF-ß and Smad3 mRNA and protein levels. The phosphorylation of Smad3 in CSRP3-knockdown cells was greater than that in wild-type cells at 24, 48, and 72 h after initiation of differentiation. Collectively, knockdown of CSRP3 suppressed chicken satellite cell differentiation by regulating Smad3 phosphorylation in the TGF-ß signaling pathway. Our results indicate that CSRP3 might play an important role in promoting satellite cell differentiation in chicken.


Assuntos
Proteínas com Domínio LIM/genética , Proteínas Musculares/genética , Células Satélites de Músculo Esquelético/citologia , Proteína Smad3/genética , Proteína Smad3/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Galinhas , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Fosforilação , Células Satélites de Músculo Esquelético/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo
10.
Biochim Biophys Acta Gene Regul Mech ; 1862(6): 625-633, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31022553

RESUMO

The eukaryotic genome is organized at varying levels into chromosome territories, transcriptional compartments and topologically associating domains (TADs), which are architectural features largely shared between different cell types and across species. In contrast, within TADs, chromatin loops connect enhancers and their target genes to establish unique transcriptomes that distinguish cells and tissues from each other and underlie development and differentiation. How these tissue-specific and temporal stage-specific long-range contacts are formed and maintained is a fundamental question in biology. The widely expressed Lim domain binding 1 protein, LDB1, plays a critical role in connecting enhancers and genes by forming complexes with cell-type specificity across diverse developmental pathways including neurogenesis, cardiogenesis, retinogenesis and hematopoiesis. Here we review the multiple roles of LDB1 in cell fate determination and in chromatin loop formation, with an emphasis on mammalian systems, to illuminate how LDB1 functions in normal cells and in diseases such as cancer.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ligação a DNA/metabolismo , Elementos Facilitadores Genéticos , Proteínas com Domínio LIM/metabolismo , Fatores de Transcrição/metabolismo , Animais , Caenorhabditis elegans , Diferenciação Celular , Cromatina , Proteínas de Ligação a DNA/genética , Expressão Gênica , Humanos , Proteínas com Domínio LIM/genética , Modelos Moleculares , Neurogênese , Proteínas Nucleares/metabolismo , Organogênese , Regiões Promotoras Genéticas , Domínios Proteicos/fisiologia , Domínios e Motivos de Interação entre Proteínas , Fatores de Transcrição/genética , Transcriptoma
11.
Cancer Biomark ; 25(1): 53-65, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31006665

RESUMO

BACKGROUND: Thyroid carcinoma is the most common endocrine malignancy worldwide. Changes in DNA methylation can cause silencing of normally active genes, especially tumour suppressor genes (TSG) or activation of normally silent genes. OBJECTIVE: The aim of this study is to evaluate the degree of promoter methylation for a panel of markers for thyroid neoplasms and to establish their relationship with thyroid oncogenesis. METHODS: To generate a comprehensive DNA methylation signature of TSGs involved in thyroid neoplasia, we use Human TSG EpiTect Methyl II Signature PCR Array-Qiagen for 24 samples (follicular adenomas and papillary thyroid carcinomas) compared with normal thyroid tissue. We extended the evaluation for three TSGs (TP73, WIF1, PDLIM4) using qMS-PCR. Statistical analysis was performed with GraphPad Prism. RESULTS: We noted four important genes NEUROG1, ESR1, RUNX3, MLH1, which presented methylated promoter in tumour samples compared to normal. We found new characteristic of thyroid tumours: methylation of TP73, WIF1 and PDLIM4 TSGs, which can contribute to thyroid neoplasia. A significant correlation between BRAF V600E mutation and RET/PTC rearrangements with TIMP3 and CDH13, RARB methylation, respectively was observed. CONCLUSIONS: TSGs promoter hypermethylation is a hallmark of cancer and a test that uses methylation quantification method is suitable for diagnosis and prognosis of thyroid cancer.


Assuntos
Metilação de DNA , Genes Supressores de Tumor/fisiologia , Neoplasias da Glândula Tireoide/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Adolescente , Adulto , Idoso , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Feminino , Expressão Gênica , Humanos , Proteínas com Domínio LIM/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras Genéticas , Proteínas Repressoras/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , Proteína Tumoral p73/genética , Adulto Jovem
12.
eNeuro ; 6(1)2019.
Artigo em Inglês | MEDLINE | ID: mdl-30873428

RESUMO

LIM domain binding protein 1 (LDB1) is a protein cofactor that participates in several multiprotein complexes with transcription factors that regulate mouse forebrain development. Since Ldb1 null mutants display early embryonic lethality, we used a conditional knockout strategy to examine the role of LDB1 in early forebrain development using multiple Cre lines. Loss of Ldb1 from E8.75 using Foxg1Cre caused a disruption of midline boundary structures in the dorsal telencephalon. While this Cre line gave the expected pattern of recombination of the floxed Ldb1 locus, unexpectedly, standard Cre lines that act from embryonic day (E)10.5 (Emx1Cre) and E11.5 (NesCre) did not show efficient or complete recombination in the dorsal telencephalon by E12.5. Intriguingly, this effect was specific to the Ldb1 floxed allele, since three other lines including floxed Ai9 and mTmG reporters, and a floxed Lhx2 line, each displayed the expected spatial patterns of recombination. Furthermore, the incomplete recombination of the floxed Ldb1 locus using NesCre was limited to the dorsal telencephalon, while the ventral telencephalon and the diencephalon displayed the expected loss of Ldb1. This permitted us to examine the requirement for LDB1 in the development of the thalamus in a context wherein the cortex continued to express Ldb1. We report that the somatosensory VB nucleus is profoundly shrunken upon loss of LDB1. Our findings highlight the unusual nature of the Ldb1 locus in terms of recombination efficiency, and also report a novel role for LDB1 during the development of the thalamus.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas com Domínio LIM/metabolismo , Telencéfalo/embriologia , Telencéfalo/metabolismo , Tálamo/embriologia , Tálamo/metabolismo , Animais , Animais Recém-Nascidos , Proteínas de Ligação a DNA/genética , Feminino , Proteínas com Domínio LIM/genética , Masculino , Camundongos Transgênicos
13.
Nat Commun ; 10(1): 1015, 2019 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-30833559

RESUMO

Candida albicans can switch from commensal to pathogenic mode, causing mucosal or disseminated candidiasis. The host relies on pattern-recognition receptors including Toll-like receptors (TLRs) and C-type lectin receptors (CLRs) to sense invading fungal pathogens and launch immune defense mechanisms. However, the complex interplay between fungus and host innate immunity remains incompletely understood. Here we report that C. albicans upregulates expression of a small secreted cysteine-rich protein Sel1 upon encountering limited nitrogen and abundant serum. Sel1 activates NF-κB and MAPK signaling pathways, leading to expression of proinflammatory cytokines and chemokines. Comprehensive genetic and biochemical analyses reveal both TLR2 and TLR4 are required for the recognition of Sel1. Further, SEL1-deficient C. albicans display an impaired immune response in vivo, causing increased morbidity and mortality in a bloodstream infection model. We identify a critical component in the Candida-host interaction that opens a new avenue to tackle Candida infection and inflammation.


Assuntos
Candida albicans/patogenicidade , Candidíase/imunologia , Proteínas de Transporte/imunologia , Proteínas de Transporte/metabolismo , Proteínas com Domínio LIM/imunologia , Proteínas com Domínio LIM/metabolismo , Receptor 2 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Animais , Proteínas de Transporte/genética , Quimiocinas/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Imunidade Inata , Inflamação/imunologia , Proteínas com Domínio LIM/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Células RAW 264.7 , Alinhamento de Sequência , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Receptores Toll-Like/imunologia
14.
J Exp Clin Cancer Res ; 38(1): 120, 2019 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-30850009

RESUMO

BACKGROUND: Cysteine-rich intestinal protein 1 (CRIP1) is highly expressed in human intestine and aberrantly expressed in several types of tumor. However, studies on CRIP1 are limited and its role on tumor development and progression remains controversial and elusive. METHODS: Immunohistochemistry was performed to evaluate the expression of CRIP1 in paired normal and colorectal tumor specimens, as well as colorectal cell lines. Functional assays, such as CCK8, TUNEL assay and in vivo tumor growth assay, were used to detect the proliferation, apoptosis and response to 5-FU of CRIP1. Western blot was used to analyze Fas-mediated pathway induced by CRIP1. Rescue experiments were performed to evaluate the essential role of CRIP1 for Fas-mediated apoptosis. RESULTS: We demonstrated that CRIP1 is overexpressed in CRC tissues compared with adjacent normal mucosa. CRIP1 could dramatically recover the 5-Fluorouracil (5-FU) inhibited CRC cell proliferation in vitro and stimulate the tumor formation of CRC in vivo, probably through inhibiting CRC cell apoptosis. Moreover, CRIP1 also dramatically recovered the 5-Fluorouracil (5-FU) induced tumor cell apoptosis in vitro. Further study demonstrated that CRIP1 down-regulated the expression of Fas protein and proteins related to Fas-mediated apoptosis. CRIP1 could interact with Fas protein and stimulate its ubiquitination and degradation. In addition, a negative correlation was detected between the expression of CRIP1 and Fas protein in most of the clinical human CRC samples. CONCLUSION: The current research reveals a vital role of CRIP1 in CRC progression, which provide a novel target for clinical drug resistance of colorectal cancer and undoubtedly contributing to the therapeutic strategies in CRC.


Assuntos
Proteínas de Transporte/genética , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas com Domínio LIM/genética , Receptor fas/genética , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Fluoruracila/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , MicroRNAs/genética , Proteólise , Ubiquitina/genética , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Int J Mol Sci ; 20(6)2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30875867

RESUMO

LIM proteins have been found to play important roles in many life activities, including the regulation of gene expression, construction of the cytoskeleton, signal transduction and metabolic regulation. Because of their important roles in many aspects of plant development, LIM genes have been studied in many plant species. However, the LIM gene family has not yet been characterized in foxtail millet. In this study, we analyzed the whole genome of foxtail millet and identified 10 LIM genes. All LIM gene promoters contain MYB and MYC cis-acting elements that are related to drought stress. Based on the presence of multiple abiotic stress-related cis-elements in the promoter of SiWLIM2b, we chose this gene for further study. We analyzed SiWLIM2b expression under abiotic stress and hormone treatments using qRT-PCR. We found that SiWLIM2b was induced by various abiotic stresses and hormones. Under drought conditions, transgenic rice of SiWLIM2b-overexpression had a higher survival rate, higher relative water content and less cell damage than wild type (WT) rice. These results indicate that overexpression of the foxtail millet SiWLIM2b gene enhances drought tolerance in transgenic rice, and the SiWLIM2b gene can potentially be used for molecular breeding of crops with increased resistance to abiotic stress.


Assuntos
Secas , Proteínas com Domínio LIM/genética , Setaria (Planta)/crescimento & desenvolvimento , Sequenciamento Completo do Genoma/métodos , Adaptação Fisiológica , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Família Multigênica , Oryza/genética , Oryza/crescimento & desenvolvimento , Filogenia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Regiões Promotoras Genéticas , Setaria (Planta)/genética
16.
Nat Commun ; 10(1): 1117, 2019 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-30850599

RESUMO

Sensory hair cells, the mechanoreceptors of the auditory and vestibular systems, harbor two specialized elaborations of the apical surface, the hair bundle and the cuticular plate. In contrast to the extensively studied mechanosensory hair bundle, the cuticular plate is not as well understood. It is believed to provide a rigid foundation for stereocilia motion, but specifics about its function, especially the significance of its integrity for long-term maintenance of hair cell mechanotransduction, are not known. We discovered that a hair cell protein called LIM only protein 7 (LMO7) is specifically localized in the cuticular plate and the cell junction. Lmo7 KO mice suffer multiple cuticular plate deficiencies, including reduced filamentous actin density and abnormal stereociliar rootlets. In addition to the cuticular plate defects, older Lmo7 KO mice develop abnormalities in inner hair cell stereocilia. Together, these defects affect cochlear tuning and sensitivity and give rise to late-onset progressive hearing loss.


Assuntos
Células Ciliadas Auditivas/fisiologia , Audição/fisiologia , Proteínas com Domínio LIM/deficiência , Fatores de Transcrição/deficiência , Actinas/metabolismo , Animais , Cóclea/fisiologia , Modelos Animais de Doenças , Células Ciliadas Auditivas/ultraestrutura , Células Ciliadas Auditivas Internas/fisiologia , Células Ciliadas Auditivas Internas/ultraestrutura , Audição/genética , Perda Auditiva/etiologia , Perda Auditiva/genética , Perda Auditiva/fisiopatologia , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Microscopia Eletrônica de Varredura , Estereocílios/genética , Estereocílios/fisiologia , Estereocílios/ultraestrutura , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia
17.
Cancer Commun (Lond) ; 39(1): 3, 2019 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-30728082

RESUMO

BACKGROUND: In our previous study, we identified a candidate tumor suppressor gene, testin LIM domain protein (TES), in primary gastric cancer (GC). TES contains three LIM domains, which are specific interacting regions for the cell adhesion and cytoskeleton regulatory proteins. Mena is a known cytoskeleton regulator that regulates the assembly of actin filaments and modulates cell adhesion and motility by interacting with Lamellipodin (Lpd). Therefore, we hypothesized that TES plays a role as tumor suppressor in GC through interacting with Mena. This study aimed to investigate the tumor suppressive functions of TES in GC. METHODS: We explored the tumor suppressive effect of TES in GC by in vitro cell proliferation assay, colony formation assay, cell cycle analysis, Transwell assays, and in vivo tumorigenicity and metastasis assays. The interaction of TES and Mena was investigated through immunoprecipitation-based mass spectrometry. We also analyzed the expression of TES and Mena in 172 GC specimens using immunohistochemistry and investigated the clinicopathological and prognostic significance of TES and Mena in GC. RESULTS: TES suppressed GC cell proliferation and colony formation, induced cell cycle arrest, and inhibited tumorigenicity in vitro. Additionally, it inhibited GC cell migration and invasion in vitro and suppressed metastasis in vivo. TES interacted with Mena, and inhibited the interaction of Mena with Lpd. Transwell assays suggested that TES suppressed migration and invasion of GC cells in a Mena-dependent fashion. In GC patients with high Mena expression, the expression of TES was associated with tumor infiltration (P = 0.005), lymph node metastasis (P = 0.003), TNM stage (P = 0.003), and prognosis (P = 0.010). However, no significant association was observed in GC patients with low Mena expression. CONCLUSIONS: We believe that TES functions as a Mena-dependent tumor suppressor. TES represents a valuable prognostic marker and potential target for GC treatment.


Assuntos
Adenocarcinoma/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas com Domínio LIM/metabolismo , Proteínas dos Microfilamentos/metabolismo , Neoplasias Gástricas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Adenocarcinoma/patologia , Animais , Proteínas de Transporte/metabolismo , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Proteínas do Citoesqueleto/genética , Humanos , Proteínas com Domínio LIM/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Nus , Processos Neoplásicos , Neoplasias Gástricas/patologia , Proteínas Supressoras de Tumor/genética
18.
Cell Mol Life Sci ; 76(15): 2987-3004, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30701284

RESUMO

Mechanosensors govern muscle tissue integrity and constitute a subcellular structure known as costameres. Costameres physically link the muscle extracellular matrix to contractile and signaling 'hubs' inside muscle fibers mainly via integrins and are localized beneath sarcolemmas of muscle fibers. Costameres are the main mechanosensors converting mechanical cues into biological events. However, the fiber type-specific costamere architecture in muscles is unexplored. We hypothesized that fiber types differ in the expression of genes coding for costamere components. By coupling laser microdissection to a multiplex tandem qPCR approach, we demonstrate that type 1 and type 2 fibers indeed show substantial differences in their mechanosensor complexes. We confirmed these data by fiber type population-specific protein analysis and confocal microscopy-based localization studies. We further show that knockdown of the costamere gene integrin-linked kinase (Ilk) in muscle precursor cells results in significantly increased slow-myosin-coding Myh7 gene, while the fast-myosin-coding genes Myh1, Myh2, and Myh4 are downregulated. In parallel, protein synthesis-enhancing signaling molecules (p-mTORSer2448, p < 0.05; p-P70S6KThr389, tendency with p < 0.1) were reduced upon Ilk knockdown. However, overexpression of slow type-inducing NFATc1 in muscle precursor cells did not change Ilk or other costamere gene expressions. In addition, we demonstrate fiber type-specific costamere gene regulation upon mechanical loading and unloading conditions. Our data imply that costamere genes, such as Ilk, are involved in the control of muscle fiber characteristics. Further, they identify costameres as muscle fiber type-specific loading management 'hubs' and may explain adaptation differences of muscle fiber types to mechanical (un)loading.


Assuntos
Costâmeros/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Actinina/genética , Actinina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Linhagem Celular , Costâmeros/genética , Regulação da Expressão Gênica , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Estresse Mecânico , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
19.
Dev Biol ; 448(1): 16-35, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30721665

RESUMO

The neural crest-a key innovation of the vertebrates-gives rise to diverse cell types including melanocytes, neurons and glia of the peripheral nervous system, and chondrocytes of the jaw and skull. Proper development of the cephalic region is dependent on the tightly-regulated specification and migration of cranial neural crest cells (NCCs). The core PCP proteins Frizzled and Disheveled have previously been implicated in NCC migration. Here we investigate the functions of the core PCP proteins Prickle1a and Prickle1b in zebrafish cranial NCC development. Using analysis of pk1a and pk1b mutant embryos, we uncover similar roles for both genes in facilitating cranial NCC migration. Disruption of either gene causes pre-migratory NCCs to cluster together at the dorsal aspect of the neural tube, where they adopt aberrant polarity and movement. Critically, in investigating Pk1-deficient cells that fail to migrate ventrolaterally, we have also uncovered roles for pk1a and pk1b in the epithelial-to-mesenchymal transition (EMT) of pre-migratory NCCs that precedes their collective migration to the periphery. Normally, during EMT, pre-migratory NCCs transition from a neuroepithelial to a bleb-based and subsequently, mesenchymal morphology capable of directed migration. When either Pk1a or Pk1b is disrupted, NCCs continue to perform blebbing behaviors characteristic of pre-migratory cells over extended time periods, indicating a block in a key transition during EMT. Although some Pk1-deficient NCCs transition successfully to mesenchymal, migratory morphologies, they fail to separate from neighboring NCCs. Additionally, Pk1b-deficient NCCs show elevated levels of E-Cadherin and reduced levels of N-Cadherin, suggesting that Prickle1 molecules regulate Cadherin levels to ensure the completion of EMT and the commencement of cranial NCC migration. We conclude that Pk1 plays crucial roles in cranial NCCs both during EMT and migration. These roles are dependent on the regulation of E-Cad and N-Cad.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Movimento Celular/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Proteínas com Domínio LIM/metabolismo , Crista Neural/embriologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Caderinas/genética , Caderinas/metabolismo , Técnicas de Silenciamento de Genes , Proteínas com Domínio LIM/genética , Crista Neural/citologia , Tubo Neural/citologia , Tubo Neural/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
20.
Gene ; 693: 16-24, 2019 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-30690182

RESUMO

Ajuba dysregulation has been reported in several human cancers. However, its expression patterns and biological roles in human gastric cancers have not yet been characterized. In the current study, we found that Ajuba protein was increased in gastric cancer tissues and in cell lines. High Ajuba expression positively correlated with the tumor-node-metastasis (TNM) stage, lymph node metastasis and poor prognosis. The Cancer Genome Atlas (TCGA) and Oncomine microarray data mining also suggested that Ajuba mRNA upregulation in gastric cancer tissues. We used SGC-7901 and NCI-N87 cell lines for Ajuba overexpression and siRNA knockdown respectively. MTT and colony formation assays indicated that Ajuba overexpression increased proliferation rate and colony formation ability while Ajuba siRNA inhibited proliferation rate and colony formation ability. AnnexinV and JC1 staining showed that Ajuba downregulated cisplatin induced apoptosis while it upregulated mitochondrial membrane potential. Ajuba overexpression also inhibited caspase-3 and PARP cleavage, while Ajuba depletion showed the opposite effects. Notably, Ajuba enhanced glucose metabolism by upregulating glucose uptake, glucose consumption, lactate production and ATP production. We further revealed that Ajuba positively regulated cyclin D1, Bcl-xL and GLUT1 at both mRNA and protein levels. Analysis of TCGA dataset revealed that there were positive correlations between Ajuba and cyclin D1, Bcl-xL, GLUT1 at the mRNA levels. Further investigation demonstrated that Ajuba overexpression inhibited Hippo signaling by upregulating YAP protein expression. Depletion of YAP by siRNA abolished the effect of Ajuba on cyclin D1, Bcl-xL and GLUT1. Together, our study showed that Ajuba was overexpressed in human gastric cancers, where it increased cell growth and chemoresistance. Our data also identified novel roles of Ajuba in gastric cancer progression involving regulating glucose uptake and mitochondrial function through the YAP-GLUT1/Bcl-xL axis, making it a potential therapeutic target.


Assuntos
Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Neoplasias Gástricas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose/genética , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Glucose/metabolismo , Transportador de Glucose Tipo 1/metabolismo , Humanos , Masculino , Potencial da Membrana Mitocondrial/fisiologia , Pessoa de Meia-Idade , Mitocôndrias/genética , Mitocôndrias/metabolismo , Fosfoproteínas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/genética , Proteína bcl-X/metabolismo
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