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This corrects the article DOI: 10.1038/mi.2017.45.
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The airway epithelium secretes proteins that function in innate defense against infection. Bactericidal/permeability-increasing fold-containing family member A1 (BPIFA1) is secreted into airways and has a protective role during bacterial infections, but it is not known whether it also has an antiviral role. To determine a role in host defense against influenza A virus (IAV) infection and to find the underlying defense mechanism, we developed transgenic mouse models that are deficient in BPIFA1 and used these, in combination with in vitro three-dimensional mouse tracheal epithelial cell (mTEC) cultures, to investigate its antiviral properties. We show that BPIFA1 has a significant role in mucosal defense against IAV infection. BPIFA1 secretion was highly modulated after IAV infection. Mice deficient in BPIFA1 lost more weight after infection, supported a higher viral load and virus reached the peripheral lung earlier, indicative of a defect in the control of infection. Further analysis using mTEC cultures showed that BPIFA1-deficient cells bound more virus particles, displayed increased nuclear import of IAV ribonucleoprotein complexes, and supported higher levels of viral replication. Our results identify a critical role of BPIFA1 in the initial phase of infection by inhibiting the binding and entry of IAV into airway epithelial cells.
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Glicoproteínas/genética , Virus de la Influenza A/fisiología , Gripe Humana/inmunología , Infecciones por Orthomyxoviridae/inmunología , Fosfoproteínas/genética , Mucosa Respiratoria/inmunología , Animales , Células Cultivadas , Regulación de la Expresión Génica , Glicoproteínas/metabolismo , Interacciones Huésped-Patógeno , Humanos , Inmunidad Innata , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfoproteínas/metabolismo , Mucosa Respiratoria/virología , Replicación ViralRESUMEN
BACKGROUND: Oral squamous cell carcinoma (OSCC) is the most frequently occurring malignant tumour in the oral cavity. OSCC arises because of multiple genetic alterations. Cell cycle aberrations and aneuploidy are reportedly among the main characteristics of cancer cells and are associated with aggressive growth and poor prognosis. METHODS: The study sample included 47 non-metastasised and 39 metastasised primary OSCC, with matched positive cervical lymph nodes and 17 normal oral mucosa samples. Tissue microarrays (TMAs) were prepared with a minimum of three cores from each case. TMA sections were cut and immunostained with MCM2, Ki-67, geminin and cyclin D1 antibodies. DNA image analysis was performed on the whole tissue section before TMAs were created. RESULTS: The results revealed that there were no differences in cell cycle protein expression in different areas of the tumours or between the metastatic and non-metastatic carcinomas. None of the cell cycle proteins showed significant differences between the lymph node metastasis and the primary OSCC, except for Ki-67. Geminin/Ki-67 ratio showed significant difference between metastatic and non-metastatic tumours. Aneuploidy was detected in all (100%) cases of OSCC. Similarly, all lymph node samples (39 cases) were aneuploid. CONCLUSION: The results suggest that although there was dysregulation of cell cycle regulatory proteins, only Ki-67 and the MCM2/Ki-67 and geminin/Ki-67 ratios may have prognostic significance in oral cancer. DNA ploidy alone was not specific and may not be a good tool to evaluate prognosis or metastatic progression in oral cavity carcinomas.
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Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Proteínas de Ciclo Celular/biosíntesis , Neoplasias de la Boca/genética , Neoplasias de la Boca/metabolismo , Ploidias , Carcinoma de Células Escamosas/secundario , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Metástasis Linfática , Masculino , Persona de Mediana Edad , Neoplasias de la Boca/patologíaRESUMEN
A previous study involving a proteomic screen of induced sputum from smokers and patients with chronic obstructive pulmonary disease (COPD) demonstrated elevated levels of bactericidal/permeability-increasing fold-containing protein B1 (BPIFB1). The aim of the present study was to further evaluate the association of sputum BPIFB1 levels with smoking and longitudinal changes in lung function in smokers with COPD. Sputum BPIFB1 was characterized by two-dimensional gel electrophoresis and mass spectrometry. The expression of BPIFB1 in COPD was investigated by immunoblotting and immunohistochemistry using sputum and lung tissue samples. BPIFB1 levels were also assessed in induced sputum from nonsmokers (n = 31), smokers (n = 169), and patients with COPD (n = 52) via an ELISA-based method. The longitudinal changes in lung function during the 4-year follow-up period were compared with the baseline sputum BPIFB1 levels. In lung tissue samples, BPIFB1 was localized to regions of goblet cell metaplasia. Secreted and glycosylated BPIFB1 was significantly elevated in the sputum of patients with COPD compared with that of smokers and nonsmokers. Sputum BPIFB1 levels correlated with pack-years and lung function as measured by forced expiratory volume in 1 s (FEV1) % predicted and FEV1/FVC (forced vital capacity) at baseline and after the 4-year follow-up in all participants. The changes in lung function over 4 years were significantly associated with BPIFB1 levels in current smokers with COPD. In conclusion, higher sputum concentrations of BPIFB1 were associated with changes of lung function over time, especially in current smokers with COPD. BPIFB1 may be involved in the pathogenesis of smoking-related lung diseases.
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Autoantígenos/metabolismo , Proteínas/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Fumar/efectos adversos , Esputo/química , Autoantígenos/biosíntesis , Proteínas de Unión a Ácidos Grasos , Femenino , Volumen Espiratorio Forzado , Glicoproteínas/biosíntesis , Glicoproteínas/metabolismo , Células Caliciformes/metabolismo , Humanos , Estudios Longitudinales , Pulmón/metabolismo , Masculino , Persona de Mediana Edad , Fosfoproteínas/biosíntesis , Fosfoproteínas/metabolismoRESUMEN
OBJECTIVE: To analyse and compare the expression of Palate, Lung, and Nasal Epithelium Clone (PLUNC) proteins in salivary glands from patients with and without AIDS (control group) using autopsy material. METHODS: We analysed the expression of PLUNCs using immunohistochemistry in parotid (n = 45), submandibular (n = 47) and sublingual gland (n = 37) samples of AIDS patients [30 with normal histology, 21 with mycobacteriosis, 14 with cytomegalovirus (CMV) infection, 30 with chronic non-specific sialadenitis, and 30 HIV-negative controls. In situ hybridization (ISH) for SPLUNC 2 in the HIV-negative group was performed. RESULTS: SPLUNC 1 expression was detected in the mucous acini of submandibular and sublingual glands, and SPLUNC 2 were seen in the serous cells. LPLUNC 1 expression was only positive in the salivary ducts. There was a higher expression of SPLUNC 2 in AIDS patients with CMV infection and mycobacteriosis when compared with all other groups. The intensity of staining for SPLUNC 2 was greater around the lesions than the peripheral ones. ISH for SPLUNC 2 showed perinuclear positivity in the serous cells in all HIV-negative cases. CONCLUSIONS: SPLUNC 1 and LPLUNC 1 proteins were similarly expressed in the salivary glands of AIDS patients and non-HIV patients. CMV infection and mycobacteriosis increase SPLUNC 2 expression in serous cells in the salivary gland of AIDS patients.
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Síndrome de Inmunodeficiencia Adquirida/patología , Glicoproteínas/análisis , Fosfoproteínas/análisis , Glándulas Salivales/patología , Infecciones Oportunistas Relacionadas con el SIDA/patología , Adolescente , Adulto , Anciano , Infecciones por Citomegalovirus/patología , Femenino , Seronegatividad para VIH , Humanos , Inmunohistoquímica , Hibridación in Situ , Masculino , Persona de Mediana Edad , Enfermedades de las Parótidas/patología , Glándula Parótida/patología , Conductos Salivales/patología , Enfermedades de las Glándulas Salivales/patología , Membrana Serosa/patología , Sialadenitis/patología , Glándula Sublingual/patología , Glándula Submandibular/patología , Enfermedades de la Glándula Submandibular/patología , Tuberculosis Bucal/patología , Adulto JovenRESUMEN
Shared care of military and civilian patients has resulted in transmission of multidrug-resistant Acinetobacter baumannii (MDR-Aci) from military casualties to civilians. Current typing technologies have been useful in revealing relationships between isolates of A. baumannii but they are unable to resolve differences between closely related isolates from small-scale outbreaks, where chains of transmission are often unclear. In a recent hospital outbreak in Birmingham, six patients were colonised with MDR-Aci isolates indistinguishable using standard techniques. We used whole-genome sequencing to identify single nucleotide polymorphisms in these isolates, allowing us to discriminate between alternative epidemiological hypotheses in this setting.
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Infecciones por Acinetobacter/epidemiología , Acinetobacter baumannii/clasificación , Técnicas de Tipificación Bacteriana , Infección Hospitalaria/epidemiología , Brotes de Enfermedades , Genoma Bacteriano , Análisis de Secuencia de ADN , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/genética , Acinetobacter baumannii/aislamiento & purificación , Infección Hospitalaria/microbiología , Farmacorresistencia Bacteriana Múltiple , Genotipo , Humanos , Masculino , Polimorfismo de Nucleótido Simple , Reino Unido/epidemiologíaRESUMEN
OBJECTIVES: The aim of this study was to determine the expression of PLUNC proteins in benign and malignant salivary gland tumours and thus their potential use as diagnostic and / or prognostic tools. MATERIALS AND METHODS: A tissue microarray was assembled from 64 salivary gland tumours including adenoid cystic carcinoma, carcinoma ex-pleomorphic adenoma, mucoepidermoid carcinoma, polymorphous low-grade adenocarcinoma, pleomorphic adenoma, acinic cell carcinoma, myoepithelial carcinoma and papillary cystadenocarcinoma. Clinicopathological data were collected retrospectively and immunohistochemical analysis of three PLUNC proteins (SPLUNC1, SPLUNC2 and LPLUNC1) was performed. Immunoreactivity was assessed as positive or negative. RESULTS: PLUNC expression was only found in mucoepidermoid carcinomas and papillary cystadenocarcinoma; all other tumours studied were negative. Mucin plugs, mucous and intermediate cells of mucoepidermoid carcinomas were positive for LPLUNC1 and SPLUNC2, but areas composed of epidermoid and clear cells were negative for all PLUNCs. Papillary cystadenocarcinoma was positive for all PLUNCs. No correlation was found with tumour grade or outcome. CONCLUSIONS: Intense expression of two PLUNC proteins in mucous cells and mucin plugs of mucoepidermoid carcinoma and papillary cystadenocarcinoma indicate that they could be used as additional diagnostic tools in some equivocal cases, but further studies are needed to understand the biological processes involved in PLUNC expression.
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Biomarcadores de Tumor , Carcinoma Mucoepidermoide/metabolismo , Cistadenocarcinoma Papilar/metabolismo , Glicoproteínas/biosíntesis , Fosfoproteínas/biosíntesis , Neoplasias de las Glándulas Salivales/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/biosíntesis , Carcinoma Mucoepidermoide/química , Cistadenocarcinoma Papilar/química , Femenino , Humanos , Técnicas para Inmunoenzimas , Masculino , Persona de Mediana Edad , Análisis por Matrices de Proteínas , Estudios Retrospectivos , Neoplasias de las Glándulas Salivales/química , Adulto JovenAsunto(s)
Proteínas Bacterianas/química , Proteínas Represoras/química , Aminoácidos/análisis , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Isótopos de Carbono , Hidrógeno , Isótopos de Nitrógeno , Resonancia Magnética Nuclear Biomolecular , Plásmidos , Conformación Proteica , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Transcripción GenéticaRESUMEN
An in vivo model has been established to study the role of macrophages in the initiation of angiogenesis by human breast tumour spheroids in vivo. The extent of the angiogenic response induced by T47D spheroids implanted into the dorsal skinfold chamber in nude mice was measured in vivo and compared to that induced by spheroids infiltrated with human macrophages prior to implantation. Our results indicate that the presence of macrophages in spheroids resulted in at least a three-fold upregulation in the release of vascular endothelial growth factor (VEGF) in vitro when compared with spheroids composed only of tumour cells. The angiogenic response measured around the spheroids, 3 days after in vivo implantation, was significantly greater in the spheroids infiltrated with macrophages. The number of vessels increased (macrophages vs no macrophages 34+/-1.9 vs 26+/-2.5, P<0.01), were shorter in length (macrophages vs no macrophages 116+/-4.92 vs 136+/-6.52, P<0.008) with an increased number of junctions (macrophages vs no macrophages 14+/-0.93 vs 11+/-1.25, P<0.025) all parameters indicative of new vessel formation. This is the first study to demonstrate a role for macrophages in the initiation of tumour angiogenesis in vivo.
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Neoplasias de la Mama/patología , Macrófagos/fisiología , Neovascularización Patológica , Animales , Femenino , Humanos , Ratones , Ratones Desnudos , Esferoides Celulares , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Parotid secretory protein (PSP) and palate-lung-nasal epithelium clone (PLUNC) are novel secretory proteins that are expressed in the oral cavity and upper airways. Both proteins are related to bactericidal/permeability increasing protein (BPI). Cationic peptides derived from BPI exhibit anti-inflammatory activity. To test if PSP (C20orf70 gene product) also contains anti-inflammatory peptides, we designed 3 cationic peptides based on the predicted structure of PSP and known active regions of BPI. Each peptide inhibited the lipopolysaccharide (LPS)-stimulated secretion of TNFalpha from RAW 264.7 macrophage cells. At 200 microg/mL, the peptide GK-7 exhibited inhibition similar to that achieved with 10 microg/mL of polymyxin B. PSP peptides directly inhibited the binding of LPS to LPS-binding protein. The cationic peptide Substance P had no inhibitory effect in these assays, confirming the specificity of the PSP peptides. These findings suggest that PSP peptides can serve as templates for the design of novel anti-inflammatory peptides.
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Péptidos Catiónicos Antimicrobianos/síntesis química , Proteínas Sanguíneas/química , Glicoproteínas/química , Proteínas de la Membrana/química , Fosfoproteínas/química , Ingeniería de Proteínas/métodos , Proteínas y Péptidos Salivales/química , Secuencia de Aminoácidos , Análisis de Varianza , Antibacterianos/química , Péptidos Catiónicos Antimicrobianos/farmacología , Proteínas Sanguíneas/genética , Línea Celular , Clonación Molecular , Glicoproteínas/genética , Glicoproteínas/farmacología , Humanos , Macrófagos/citología , Macrófagos/efectos de los fármacos , Proteínas de la Membrana/genética , Datos de Secuencia Molecular , Fosfoproteínas/genética , Fosfoproteínas/farmacología , ARN/análisis , Proteínas y Péptidos Salivales/genética , Proteínas y Péptidos Salivales/farmacología , Alineación de Secuencia , Análisis de Secuencia de Proteína , Factor de Necrosis Tumoral alfa/efectos de los fármacosRESUMEN
Palate, lung, and nasal epithelium clone (Plunc, now renamed Splunc1) is a small secreted protein expressed in the oropharynx and upper airways of humans, mice, rats, and cows. This protein is structurally homologous to known mediators of host defense against gram-negative bacteria. We have characterized the genomic sequence and expression of a novel but closely related gene from rodents, which we call splunc5. Mouse Splunc5 sequence is 60% identical to mouse Splunc1. The genes also share highly conserved genomic elements including intron-exon structure and intronic sequence. Strikingly, splunc5 is expressed exclusively in the interpapillary epithelium of the tongue's dorsal surface. By comparing the expression profiles of splunc5, splunc1, and a third related sequence, lplunc1, in mice, we show that these genes are expressed in unique domains along a continuous corridor of oral, lingual, pharyngeal, and respiratory epithelia. This expression pattern is consistent with the hypothesis that these proteins protect epithelial surfaces colonized by potentially pathogenic microorganisms.
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Células Epiteliales/metabolismo , Regulación de la Expresión Génica , Glicoproteínas/biosíntesis , Glicoproteínas/genética , Fosfoproteínas/biosíntesis , Fosfoproteínas/genética , Biosíntesis de Proteínas , Proteínas/genética , Lengua/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , Exones , Hibridación in Situ , Intrones , Ratones , Ratones Endogámicos C57BL , Modelos Genéticos , Datos de Secuencia Molecular , Familia de Multigenes , ARN/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Glándulas Salivales/metabolismo , Homología de Secuencia de Ácido Nucleico , Distribución TisularRESUMEN
The role of macrophages in tumour growth and development is complex and multifaceted. Whilst there is limited evidence that tumour-associated macrophages (TAMs) can be directly tumouricidal and stimulate the anti-tumour activity of T cells, there is now contrasting evidence that tumour cells are able to block or evade the activity of TAMs at the tumour site. In some cases, tumour-derived molecules even redirect TAM activities to promote tumour survival and growth. Indeed, evidence has emerged for a symbiotic relationship between tumour cells and TAMs, in which tumour cells attract TAMs and sustain their survival, with TAMs then responding to micro-environmental factors in tumours such as hypoxia (low oxygen tension) by producing important mitogens as well as various growth factors and enzymes that stimulate tumour angiogenesis. This review presents evidence for the number and/or distribution of TAMs being linked to prognosis in different types of human malignancy. It also outlines the range of pro- and anti-tumour functions performed by TAMs, and the novel therapies recently devised using TAMs to stimulate host immune responses or deliver therapeutic gene constructs to solid tumours.
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Macrófagos/inmunología , Neoplasias/inmunología , Recuento de Células , Citocinas/inmunología , Progresión de la Enfermedad , Humanos , Hipoxia , Inmunoterapia , Activación de Macrófagos , Neoplasias/terapia , Pronóstico , Linfocitos T/inmunologíaRESUMEN
Trypanosoma brucei undergoes genetic exchange in its insect vector by an unknown mechanism. To visualize the production of hybrids in the fly, a tetracycline (Tet)-inducible expression system was adapted. One parental trypanosome clone was transfected with the gene encoding Green Fluorescent Protein (GFP) under control of the Tet repressor in trans; transfection with these constructs also introduced genes for resistance to hygromycin and phleomycin, respectively. An experimental cross with a second parental clone carrying a gene for geneticin resistance produced fluorescent hybrids with both hygromycin and geneticin resistance. These results are consistent with the meiotic segregation and reassortment of the GFP and repressor genes. Fluorescent hybrids were visible in the salivary glands of the fly, but not the midgut, confirming that genetic exchange occurs among the trypanosome life cycle stages present in (or possibly en route to) the salivary glands. In conclusion, the experimental design has successfully produced fluorescent hybrids which can be observed directly in the salivary glands of the fly, and it has been shown that the recombinant genotypes were most probably the result of meiosis.
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Genes Reporteros , Trypanosoma brucei brucei/genética , Moscas Tse-Tse/parasitología , Animales , Quimera , Cruzamientos Genéticos , Electroforesis en Gel de Campo Pulsado , Regulación de la Expresión Génica , Proteínas Fluorescentes Verdes , Insectos Vectores/parasitología , Proteínas Luminiscentes , Proteínas Represoras , Glándulas Salivales/parasitologíaRESUMEN
Elafin, a low molecular-weight proteinase inhibitor, is a member of the recently described trappin gene family. These proteins are thought to play important roles in the regulation of inflammation and are expressed in multiple epithelia. Elafin is found within the lung, and its expression can be induced by inflammatory mediators. The molecular mechanisms that mediate its induction are not understood. In this study we investigated the transcriptional regulation of the elafin gene in pulmonary epithelial cell lines. Transfection of elafin promoter constructs into the elafin-expressing pulmonary epithelial cell line A549 identified a number of positive-acting elements. Cytokine-mediated inducibility of the elafin gene promoter was shown to occur through a nuclear factor (NF)-kappaB site present within the minimal promoter. This site was shown to bind to NF-kappaB proteins within nuclear extracts from cytokine stimulated cell lines as well as to in vitro-translated RelA. Cotransfection with both RelA and NF-kappaB-inducing kinase induced reporter gene activation via this site, and mutagenesis experiments confirmed that it was crucial for induction of elafin gene activity. These results clearly identify a role for elafin in the inflammatory response of the airway epithelium to pathogenic insult and show that this response is mediated by an NF-kappaB site within the proximal promoter.
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Regulación de la Expresión Génica/fisiología , Interleucina-1/fisiología , Pulmón/metabolismo , FN-kappa B/fisiología , Proteínas/genética , Factor de Necrosis Tumoral alfa/fisiología , Secuencia de Bases , Línea Celular , Sondas de ADN , Células Epiteliales/metabolismo , Humanos , Ligasas/genética , Pulmón/citología , Regiones Promotoras Genéticas , Unión Proteica , Proteínas Inhibidoras de Proteinasas Secretoras , Proteínas/metabolismo , Transcripción Genética , Activación Transcripcional , TransfecciónRESUMEN
Few genes have been isolated which display specific expression in the proximal airways. A recently identified mouse cDNA, plunc, appears to be confined to the upper airways and nasopharyngeal epithelium, and may prove a useful marker for these regions. We now report the genomic cloning and characterization of the mouse plunc gene as well as its developmental expression in the nasal and airway epithelium. We also report the novel finding that plunc is also expressed in the medullary compartment of the murine thymus. The mouse gene contains nine exons and the intron-exon boundaries are conserved with those in the human homologue. At e14.5 plunc is expressed in the nasal epithelium and several days later is seen in the thymic lobes, but not in the lining of the tracheobronchial tree. Expression in the trachea and main-stem bronchi first appears at 1--2 days after birth. Tracheobronchial expression persists at high levels throughout adulthood, as do regional areas of nasal and thymic expression. Finally, we show that the human homologue is expressed in bronchial epithelium, suggesting a transcript that is evolutionarily conserved in the mammalian airway.
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Glicoproteínas/genética , Pulmón/embriología , Fosfoproteínas/genética , Timo/embriología , Animales , Bronquios/metabolismo , Glicoproteínas/metabolismo , Humanos , Hibridación in Situ , Pulmón/metabolismo , Ratones , Nasofaringe/embriología , Nasofaringe/metabolismo , Fosfoproteínas/metabolismo , Sitios de Empalme de ARN , ARN Mensajero/biosíntesis , Mucosa Respiratoria/embriología , Mucosa Respiratoria/metabolismo , Timo/metabolismo , Tráquea/embriología , Tráquea/metabolismoRESUMEN
Bacterial plasmids deploy a diverse range of regulatory mechanisms to control expression of the functions they need to survive in the host population. Understanding of the mechanisms by which autoregulatory circuits control plasmid survival functions, in particular plasmid replication, has been advanced by recent studies. At a molecular level, structural understanding of how certain antisense RNAs control replication and stability functions is almost complete. Control circuits linking plasmid transfer functions to the status of the bacterial population have been dissected, uncovering a complex and hierarchical organisation. Coordinate or global regulation of plasmid replication, transfer and stable maintenance functions is becoming apparent across a range of plasmid families.
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Replicación del ADN/fisiología , ADN Bacteriano , Plásmidos/genética , Regulación Bacteriana de la Expresión Génica , ARN sin Sentido/genéticaRESUMEN
The KorB and TrbA proteins of broad-host-range plasmid RK2 are key regulators of the plasmid genes required for conjugative transfer. trbBp is the primary promoter responsible for expression of mating pair formation genes. We show that despite the targets for KorB and TrbA at trbBp being about 165 bp apart, 189 bp upstream of the transcription start point and overlapping the -10 region, respectively, these two proteins show up to 10-fold cooperativity for the repression of trbBp. Deletion analysis of TrbA showed that the C-terminal domain (CTD), which has a high degree of sequence conservation with the CTD of KorA, is required for this cooperativity with KorB. Western blotting demonstrated that the apparently mutual enhancement of repression is not due simply to elevation of repressor level by the presence of the second protein, suggesting that the basis for cooperativity is interaction between KorB and TrbA bound at their respective operators.
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Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proteínas de Escherichia coli , Escherichia coli/genética , Factores R/genética , Proteínas Represoras/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Sitios de Unión , Conjugación Genética , Regulación Bacteriana de la Expresión Génica , Datos de Secuencia Molecular , Unión Proteica , Secuencias Reguladoras de Ácidos NucleicosRESUMEN
Here we report the cloning and characterization of the human homologue of plunc, a murine gene expressed specifically in the upper airways and nasopharyngeal regions. The human plunc cDNA codes for a leucine-rich protein of 256 amino acids which is 72% identical to the murine protein. RNA blot analysis suggests that expression of plunc is restricted to the trachea, upper airway, nasopharyngeal epithelium and salivary gland. The human plunc gene contains nine exons and is localised to chromosome 20q11.2. The unique expression pattern of the human plunc suggest that it may prove a useful model gene with which to study the regulatory mechanisms which direct expression of genes specifically to the upper airways.
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Glicoproteínas/genética , Pulmón/metabolismo , Nasofaringe/metabolismo , Fosfoproteínas/genética , Tráquea/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , ADN Complementario/química , Epitelio/metabolismo , Expresión Génica , Glicoproteínas/química , Humanos , Datos de Secuencia Molecular , Fosfoproteínas/química , Homología de Secuencia de AminoácidoRESUMEN
We have assessed the potential of using the spliced leader (SL) or mini-exon gene as a marker for molecular phylogenetic analysis of genus Trypanosoma. A total of 27 trypanosome sequences were compared, 18 of these being newly reported. In contrast to genus Leishmania, we found the non-transcribed spacer region of the SL locus in trypanosomes to be far too variable for informative comparison of all but the most closely related species. At the other extreme, the short (39 nt) SL exon was usually completely conserved and hence uninformative. The SL RNA showed variation in both length (97-152 nt) and sequence among different trypanosome species, with most variation occurring in stem-loop II. Consequently, this region could not be aligned with confidence in multiple sequence alignment, severely reducing the number of phylogenetically informative nucleotide positions. In computer simulation, most of the SL RNAs readily folded into the 3 stem-loop secondary structure predicted previously, but again stem-loop II was highly variable. No obvious correlation could be seen between the length of this stem-loop and trypanosome biology. We conclude that the SL repeat is not an informative phylogenetic marker for long range evolutionary studies of genus Trypanosoma.
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Variación Genética , ARN Lider Empalmado/genética , Trypanosoma/genética , Animales , Secuencia de Bases , ADN Protozoario/genética , Exones/genética , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Reacción en Cadena de la Polimerasa , ARN Lider Empalmado/química , Análisis de Secuencia de ADN , Transcripción Genética , Tripanosomiasis/parasitología , Tripanosomiasis/veterinariaRESUMEN
This paper presents a re-evaluation of the taxonomic position and evolutionary relationships of Trypanosoma (Herpetosoma) rangeli based on the phylogenetic analysis of ssrRNA sequences of 64 Trypanosoma species and comparison of mini-exon sequences. All five isolates of T. rangeli grouped together in a clade containing Trypanosoma (Schizotrypanum) cruzi and a range of closely related trypanosome species from bats [Trypanosoma (Schizotrypanum) dionisii, Trypanosoma (Schizotrypanum) vespertilionis] and other South American mammals [Trypanosoma (Herpetosoma) leeuwenhoeki, Trypanosoma (Megatrypanum) minasense, Trypanosoma (Megatrypanum) conorhini] and an as yet unidentified species of trypanosome from an Australian kangaroo. Significantly T. rangeli failed to group with (a) species of subgenus Herpetosoma, other than those which are probably synonyms of T. rangeli, or (b) species transmitted via the salivarian route, although either of these outcomes would have been more consistent with the current taxonomic and biological status of T. rangeli. We propose that use of the names Herpetosoma and Megatrypanum should be discontinued, since these subgenera are clearly polyphyletic and lack evolutionary and taxonomic relevance. We hypothesise that T. rangeli and T. cruzi represent a group of mammalian trypanosomes which completed their early evolution and diversification in South America.