RESUMEN
OBJECTIVE: It remains unknown whether individuals with a family history (FH) of gastric cancer (GC) are associated with aberrant DNA methylation. The aim of this study was to investigate the association between aberrant DNA methylation and FH of GC. DESIGN: Using quantitative MethyLight assay, MOS, miR124a-3, NKX6-1, EMX1, CDH1, and TWIST1 methylation levels in the noncancerous gastric mucosa was compared between subjects with and without FH based on GC and Helicobacter pylori (Hp) infection. Changes in the methylation levels were evaluated over time after Hp eradication. RESULTS: In Hp-positive GC patients, MOS (P < 0.001), CDH1 (P < 0.001), and TWIST1 (P = 0.004) methylation were decreased in subjects with FH (n = 64) than in those without FH (n = 58). In Hp-positive controls, MOS methylation was lower in subjects with FH (n = 73) than in those without FH (n = 50) (P = 0.042), while miR124a-3 (P = 0.006), NKX6-1 (P < 0.001), and CDH1 (P < 0.001) methylation were higher in subjects with FH. CDH1 methylation constantly decreased from 2 years in GC patients and 3-4 years in controls after Hp eradication (all P < 0.001). A persistent decrease in methylation over time was not observed in other genes after eradication. CONCLUSION: The methylation of MOS and CDH1 provided an association between aberrant DNA methylation and gastric carcinogenesis in FH of GC, a useful marker for GC risk in individuals with FH. Furthermore, CDH1 methylation decreased after Hp eradication.
Asunto(s)
Metilación de ADN/genética , Infecciones por Helicobacter/genética , Anamnesis , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiología , Antígenos CD/genética , Biomarcadores de Tumor/genética , Cadherinas/genética , Estudios de Casos y Controles , Femenino , Mucosa Gástrica/metabolismo , Genes mos/genética , Helicobacter pylori , Proteínas de Homeodominio/genética , Humanos , Masculino , MicroARNs/genética , Persona de Mediana Edad , Proteínas Nucleares/genética , Factores de Transcripción/genética , Proteína 1 Relacionada con Twist/genéticaRESUMEN
The aims of this study were to investigate the effects of epidermal growth factor (EGF) and progesterone on the development, viability and the gene expression of bovine secondary follicle culture in vitro for 18 days. Secondary follicles (â¼0.2â¯mm) were isolated from ovarian cortex and individually cultured at 38.5⯰C, with 5% CO2 in air, for 18 days, in TCM-199+ (nâ¯=â¯63) alone (control medium) or supplemented with 10â¯ng/mL progesterone (nâ¯=â¯64), 10â¯ng/mL EGF (nâ¯=â¯61) or both EGF and progesterone (nâ¯=â¯66). The effects of these treatments on growth, antrum formation, viability, ultrastructure and mRNA levels for GDF-9, c-MOS, H1foo and cyclin B1 were evaluated, significantly different (pâ¯<â¯0.05). The results showed that there was a progressive increase in follicular diameter in all treatments, but only follicles cultured in medium supplemented with EGF had increased significantly in diameter when compared to follicles cultured in the control medium at the end of the culture period, significantly different (pâ¯<â¯0.05). A positive interaction between EGF and progesterone was not observed. In addition, the presence of EGF, progesterone or both in culture medium did not influence the rate of follicle survival and antrum formation. However, the presence of only progesterone in cultured medium increased the expression of mRNAs for GDF9 and cyclin B1 in oocytes. EGF also significantly increased the levels of mRNAs for cMOS and GDF9 when compared to follicles cultured in control medium. Ultrastructural analyzes showed that cultured follicles in all treatments maintained the integrity of granulosa cells. In conclusion, the EGF promotes the development of secondary follicles cultured in vitro for 18 days and increases the expression of cMOS and GDF9, while progesterone alone or in association with EGF have not a positive effect on follicular growth. However, progesterone increases the expression of GDF9 and cyclin B1 in oocytes.
Asunto(s)
Factor de Crecimiento Epidérmico/farmacología , Expresión Génica/efectos de los fármacos , Folículo Ovárico/efectos de los fármacos , Progesterona/farmacología , Animales , Bovinos , Células Cultivadas , Femenino , Genes mos/efectos de los fármacos , Genes mos/genética , Células de la Granulosa/efectos de los fármacos , Células de la Granulosa/fisiología , Factor 9 de Diferenciación de Crecimiento/genética , Folículo Ovárico/fisiologíaRESUMEN
BACKGROUND/AIMS: To identify the risk factors for metachronous gastric neoplasms in patients who underwent an endoscopic resection of a gastric neoplasm. METHODS: We prospectively collected clinicopathologic data and measured the methylation levels of HAND1, THBD, APC, and MOS in the gastric mucosa by methylation-specific real-time polymerase chain reaction in patients who underwent endoscopic resection of gastric neoplasms. RESULTS: A total of 257 patients with gastric neoplasms (113 low-grade dysplasias, 25 highgrade dysplasias, and 119 early gastric cancers) were enrolled. Metachronous gastric neoplasm developed in 7.4% of patients during a mean follow-up of 52 months. The 5-year cumulative incidence of metachronous gastric neoplasm was 4.8%. Multivariate analysis showed that moderate/severe corpus intestinal metaplasia and family history of gastric cancer were independent risk factors for metachronous gastric neoplasm development; the hazard ratios were 4.12 (95% confidence interval [CI], 1.23 to 13.87; p=0.022) and 3.52 (95% CI, 1.09 to 11.40; p=0.036), respectively. The methylation level of MOS was significantly elevated in patients with metachronous gastric neoplasms compared age- and sex-matched patients without metachronous gastric neoplasms (p=0.020). CONCLUSIONS: In patients who underwent endoscopic resection of gastric neoplasms, moderate/severe corpus intestinal metaplasia and a family history of gastric cancer were independent risk factors for metachronous gastric neoplasm, and MOS was significantly hypermethylated in patients with metachronous gastric neoplasms.
Asunto(s)
Neoplasias Primarias Secundarias/genética , Neoplasias Gástricas/patología , Anciano , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Metilación de ADN , Femenino , Gastrectomía/métodos , Genes APC/fisiología , Genes mos/genética , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Análisis Multivariante , Neoplasias Primarias Secundarias/epidemiología , Neoplasias Primarias Secundarias/patología , Modelos de Riesgos Proporcionales , Factores de Riesgo , Neoplasias Gástricas/genética , Neoplasias Gástricas/cirugía , Trombomodulina/genéticaRESUMEN
BACKGROUND/AIMS: To identify the risk factors for metachronous gastric neoplasms in patients who underwent an endoscopic resection of a gastric neoplasm. METHODS: We prospectively collected clinicopathologic data and measured the methylation levels of HAND1, THBD, APC, and MOS in the gastric mucosa by methylation-specific real-time polymerase chain reaction in patients who underwent endoscopic resection of gastric neoplasms. RESULTS: A total of 257 patients with gastric neoplasms (113 low-grade dysplasias, 25 high-grade dysplasias, and 119 early gastric cancers) were enrolled. Metachronous gastric neoplasm developed in 7.4% of patients during a mean follow-up of 52 months. The 5-year cumulative incidence of metachronous gastric neoplasm was 4.8%. Multivariate analysis showed that moderate/severe corpus intestinal metaplasia and family history of gastric cancer were independent risk factors for metachronous gastric neoplasm development; the hazard ratios were 4.12 (95% confidence interval [CI], 1.23 to 13.87; p=0.022) and 3.52 (95% CI, 1.09 to 11.40; p=0.036), respectively. The methylation level of MOS was significantly elevated in patients with metachronous gastric neoplasms compared age- and sex-matched patients without metachronous gastric neoplasms (p=0.020). CONCLUSIONS: In patients who underwent endoscopic resection of gastric neoplasms, moderate/severe corpus intestinal metaplasia and a family history of gastric cancer were independent risk factors for metachronous gastric neoplasm, and MOS was significantly hypermethylated in patients with metachronous gastric neoplasms.
Asunto(s)
Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Metilación de ADN , Gastrectomía/métodos , Genes APC/fisiología , Genes mos/genética , Incidencia , Análisis Multivariante , Neoplasias Primarias Secundarias/epidemiología , Modelos de Riesgos Proporcionales , Factores de Riesgo , Neoplasias Gástricas/genética , Trombomodulina/genéticaRESUMEN
Aberrant DNA methylation is frequently found during gastric carcinogenesis. Recently, we identified potential methylation markers important for Helicobacter pylori-induced gastric carcinogenesis using an Illumina methylation chip assay. In this study, we evaluated the candidate genes as markers for gastric cancer (GC) in a large Korean population. DNA methylation of PTPN6, MOS, DCC, CRK, and VAV1 was evaluated in non-neoplastic gastric specimens using quantitative methylation-specific PCR in patients with GC (n = 207) and their age- and gender-matched controls (n = 207). Methylation levels in 125 GC samples were also compared. H. pylori infection status was categorized as negative, active, or past infection according to the results of endoscopy-based tests (CLOtest, histology, and culture), H. pylori serology, and serum pepsinogen test. In the controls, active H. pylori infection increased methylation levels in DCC, CRK, MOS, and VAV1 but decreased methylation levels in PTPN6 (all p < 0.05); the methylation levels in MOS remained increased in patients with past H. pylori infection compared to H. pylori-negative subjects (p < 0.001). Methylation levels in MOS in non-neoplastic gastric mucosae increased in the presence of GC, regardless of H. pylori infection status (p < 0.01). Methylation levels in all genes but DCC decreased significantly in GC specimens compared to neoplastic gastric mucosae (p < 0.01); however, methylation levels in GC tissues were not correlated with those in their background gastric mucosae. Hypomethylation of MOS in GC tissues was associated with tumour invasion, nodal metastasis, and undifferentiated histology (p < 0.05). To summarize, among the candidate genes, DNA methylation of MOS may reflect the duration of H. pylori exposure and may be a marker for the development of GC.
Asunto(s)
Metilación de ADN , Mucosa Gástrica/patología , Gastritis/patología , Predisposición Genética a la Enfermedad , Lesiones Precancerosas/patología , Neoplasias Gástricas/patología , Femenino , Mucosa Gástrica/microbiología , Gastritis/genética , Gastritis/microbiología , Genes mos/genética , Infecciones por Helicobacter/genética , Infecciones por Helicobacter/microbiología , Infecciones por Helicobacter/patología , Helicobacter pylori , Humanos , Masculino , Persona de Mediana Edad , Lesiones Precancerosas/genética , Lesiones Precancerosas/microbiología , Valor Predictivo de las Pruebas , Riesgo , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiologíaRESUMEN
Transposable elements (TEs) are highly abundant in the genome and capable of mobility, two properties that make them particularly prone to transfer horizontally between organisms. Although the impact of horizontal transfer (HT) of TEs is well recognized in prokaryotes, the frequency of this phenomenon and its contribution to genome evolution in eukaryotes remain poorly appreciated. Here, we provide evidence that a DNA transposon called SPIN has colonized the genome of 17 species of reptiles representing nearly every major lineage of squamates, including 14 families of lizards, snakes, and amphisbaenians. Slot blot analyses indicate that SPIN has amplified to high copy numbers in most of these species, ranging from 2,000-28,000 copies per haploid genome. In contrast, we could not detect the presence of SPIN in any of the turtles (seven species from seven families) and crocodiles (four species) examined. Genetic distances between SPIN sequences from species belonging to different squamate families are consistently very low (average = 0.1), considering the deep evolutionary divergence of the families investigated (most are >100 My diverged). Furthermore, these distances fall below interfamilial distances calculated for two genes known to have evolved under strong functional constraint in vertebrates (RAG1, average = 0.24 and C-mos, average = 0.27). These data, combined with phylogenetic analyses, indicate that the widespread distribution of SPIN among squamates is the result of at least 13 independent events of HTs. Molecular dating and paleobiogeographical data suggest that these transfers took place during the last 50 My on at least three different continents (North America, South America and, Africa). Together, these results triple the number of known SPIN transfer events among tetrapods, provide evidence for a previously hypothesized transoceanic movement of SPIN transposons during the Cenozoic, and further underscore the role of HT in the evolution of vertebrate genomes.
Asunto(s)
Elementos Transponibles de ADN/genética , Evolución Molecular , Transferencia de Gen Horizontal , Reptiles/genética , Animales , Genes RAG-1/genética , Genes mos/genética , Datos de Secuencia Molecular , Filogenia , Reptiles/clasificación , Análisis de Secuencia de ADNRESUMEN
After DNA or spindle damage, p53-defective tumor cells undergo a complex cycle of reversible polyploidy. How this process occurs and more importantly, why, has recently become the focus of several research groups, prompting this review in which we discuss two related phenomena that accompany the reversible polyploidy of tumor cells: the induction of meiosis genes such as MOS and the decrease in genomic instability observed during the reversion from polyploidy to para-diploidy. The reversible polyploidy likely provides the means through which the balance between increased chromosome instability (CIN), driving genetic variation and decreased CIN, necessary for perpetuating these malignant clones, is maintained. These concepts are integrated with recent findings that many meiotic and self-renewal genes become activated during reversible polyploidy and lead us to the hypothesis that tumor cell immortality may be achieved through germline-like transmission.
Asunto(s)
Aneuploidia , Regulación Neoplásica de la Expresión Génica , Genes mos/genética , Neoplasias/genética , Ploidias , Animales , HumanosRESUMEN
In oocytes, many maternally supplied products are stored, and these products play important roles in cell cycle regulation and early development. Mos protein, which is coded on the c-mos gene, promotes oocyte maturation and is involved in MAP-kinase signaling pathway. In Xenopus, maternally supplied c-mos mRNA undergoes poly(A) addition, and translational activation via CPE (cytoplasmic polyadenylation element) and CPEB (CPE binding protein). The elongated poly(A) is shortened and the c-mos mRNA is degraded during early embryogenesis via EDEN (embryo deadenylation element) and EDEN-BP (EDEN-binding protein). We cloned the full-length zebrafish c-mos gene, which is conserved at the protein coding region in vertebrates. c-mos mRNA has two putative CPE sequences in its 3'UTR, which binds to zebrafish CPEB homologous protein, Zor-1. We could not observe EDEN sequence, and could not detect interaction between c-mos mRNA and zebrafish EDEN-BP homologous protein, Brul, even though immuno precipitation and RT-PCR experiments suggested that c-mos mRNA interacts with Zor-1 in vivo. Interestingly, we found c-mos mRNA is located in the animal cortex of zebrafish oocyte, where Zor-1 protein exists. Taken together, these results suggest that the animal cortex is the central core of oocyte maturation in zebrafish.
Asunto(s)
Genes mos/genética , Oocitos/metabolismo , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/embriología , Animales , Northern Blotting , Polaridad Celular/fisiología , Clonación Molecular , Cartilla de ADN/genética , ADN Complementario/genética , Inmunoprecipitación , Hibridación in Situ , Oocitos/citología , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The Mos proto-oncogene is a critical regulator of vertebrate oocyte maturation. The maturation-dependent translation of Mos protein correlates with the cytoplasmic polyadenylation of the maternal Mos mRNA. However, the precise temporal requirements for Mos protein function differ between oocytes of model mammalian species and oocytes of the frog Xenopus laevis. Despite the advances in model organisms, it is not known if the translation of the human Mos mRNA is also regulated by cytoplasmic polyadenylation or what regulatory elements may be involved. We report that the human Mos 3' untranslated region (3' UTR) contains a functional cytoplasmic polyadenylation element (CPE) and demonstrate that the endogenous Mos mRNA undergoes maturation-dependent cytoplasmic polyadenylation in human oocytes. The human Mos 3' UTR interacts with the human CPE-binding protein and exerts translational control on a reporter mRNA in the heterologous Xenopus oocyte system. Unlike the Xenopus Mos mRNA, which is translationally activated by an early acting Musashi/polyadenylation response element (PRE)-directed control mechanism, the translational activation of the human Mos 3' UTR is dependent on a late acting CPE-dependent process. Taken together, our findings suggest a fundamental difference in the 3' UTR regulatory mechanisms controlling the temporal induction of maternal Mos mRNA polyadenylation and translational activation during Xenopus and mammalian oocyte maturation.
Asunto(s)
Regiones no Traducidas 3'/genética , Regulación del Desarrollo de la Expresión Génica/genética , Genes mos/genética , Oocitos/crecimiento & desarrollo , Poliadenilación/genética , ARN Mensajero/metabolismo , Animales , Western Blotting , Cartilla de ADN/genética , Ensayo de Cambio de Movilidad Electroforética , Humanos , Mutagénesis Sitio-Dirigida , Oocitos/metabolismo , Proto-Oncogenes Mas , ARN Mensajero/genética , Especificidad de la Especie , Factores de Transcripción/metabolismo , Xenopus , Factores de Escisión y Poliadenilación de ARNm/metabolismoRESUMEN
AIM: To immunolocalize the c-mos gene product and to investigate its spatial and temporal expression in mouse testis during postnatal development. METHODS: Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization techniques were used to examine c-mos mRNA and indirect immunofluorescence was used to localize c-Mos protein in mouse testis on postnatal days 14, 21, 25, 28, 30, 35, 49 and 70. RESULTS: c-mos mRNA remained low on postnatal days 14-21, increased abruptly from day 25 and peaked on day 30. Its levels decreased a little on day 35 and became almost stable thereafter until day 70. c-mos mRNA was localized in the nucleus and cytoplasm of the spermatocytes and round spermatids. The nuclear staining was much stronger than the cytoplasmic staining. Using a polyclonal anti-c-Mos antibody, Western blotting detected a single band at 43 kDa in testis lysate. c-Mos protein was exclusively localized to the elongating spermatids and was first detected on postnatal day 30. The number of c-Mos-positive spermatids increased progressively till day 49 and stabilized thereafter. CONCLUSION: The c-mos gene displays a spatial and temporal expression pattern in the mouse testis during postnatal development at both the mRNA and protein level. This suggests that c-mos might play important roles in spermatogenesis.
Asunto(s)
Genes mos/genética , Espermatocitos/metabolismo , Espermatogénesis/genética , Testículo/metabolismo , Animales , Técnica del Anticuerpo Fluorescente Indirecta , Expresión Génica , Masculino , Ratones , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Espermatocitos/crecimiento & desarrolloRESUMEN
Gekkonid geckos, representing more than 85 percent of the gekkotan genera, are found on all major land masses and almost all oceanic islands in the tropics and subtropics. Intergeneric relationships of the Gekkonidae have been far more difficult to resolve than those among other gekkotan families. Our data set consists of a large number of complete mitochondrial 12S rRNA gene sequences and partial nuclear C-mos gene sequences for 33 genera of geckos, two genera of pygopods and two genera of eublepharids. Maximum parsimony (MP) and maximum likelihood (ML) trees were generated based on unweighted analysis using PAUP 4.0b10. Bayesian inference (BI) analyses trees were generated by MrBayes 3.0B4. All phylogenetic trees supported the monophyly of Gekkonidae with great confidence. The 12S data and combined data (12S and C-mos) place Sphaerodactylus deeply within gekkonine geckos, whereas Teratoscincus+Pristurus are the sister group of the remaining gekkonids. However, it is known that 12S may be positively misleading when dealing with older divergences. Therefore, the conflict between the results in this study and the latest conclusions based on C-mos points to the need for future focus on the phylogenetic position of both Sphaerodactylus and Teratoscincus.
Asunto(s)
Genes mos/genética , Lagartos/clasificación , Lagartos/genética , Filogenia , ARN Ribosómico/genética , Animales , Secuencia de Bases , Cartilla de ADN/química , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa/veterinaria , Alineación de Secuencia/veterinariaRESUMEN
The extant crocodylians comprise 23 species divided among three families, Alligatoridae, Crocodylidae, and Gavialidae. Currently, based on morphological data sets, Tomistoma schlegelii (false gharial) is placed within the family Crocodylidae. Molecular data sets consistently support a sister-taxon relationship of T. schlegelii with Gavialis gangeticus (Indian Gharial), which is the sole species in Gavialidae. To elucidate the placement of T. schlegelii within the extant crocodylians, we have sequenced 352bp of the dentin matrix protein 1 (DMP1) nuclear gene in 30 individuals and 424bp of the nuclear gene C-mos in 74 individuals. Molecular analysis of the DMP1 data set indicates that it is highly conserved within the Crocodylia. Of special note is a seven base-pair indel (GTGCTTT) shared by T. schlegelii and G. gangeticus, that is absent in the genus Crocodylus, Osteolaemus, and Mecistops. To date, C-mos is the largest molecular data set analyzed for any crocodylian study including multiple samples from all representatives of the eight extant genera. Analysis of these molecular data sets, both as individual gene sequences and concatenated sequences, support the hypothesis that T. schlegelii should be placed within the family Gavialidae.
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Caimanes y Cocodrilos/genética , Filogenia , Caimanes y Cocodrilos/clasificación , Animales , Núcleo Celular/genética , Proteínas de la Matriz Extracelular/genética , Genes mos/genética , Datos de Secuencia Molecular , Análisis de Secuencia de ADNRESUMEN
We use approximately 1900bp of mitochondrial (ND2) and nuclear (c-mos and Rag-1) DNA sequence data to recover phylogenetic relationships among 58 species and 26 genera of Eugongylus group scincid lizards from New Caledonia, Lord Howe Island, New Zealand, Australia and New Guinea. Taxon sampling for New Caledonian forms was nearly complete. We find that the endemic skink genera occurring on New Caledonia, New Zealand and Lord Howe Island, which make up the Gondwanan continental block Tasmantis, form a monophyletic group. Within this group New Zealand and New Zealand+Lord Howe Island form monophyletic clades. These clades are nested within the radiation of skinks in New Caledonia. All of the New Caledonian genera are monophyletic, except Lioscincus. The Australian and New Guinean species form a largely unresolved polytomy with the Tasmantis clade. New Caledonian representatives of the more widespread genera Emoia and Cryptoblepharus are more closely related to the non-Tasmantis taxa than to the endemic New Caledonian genera. Using ND2 sequences and the calibration estimated for the agamid Laudakia, we estimate that the diversification of the Tasmantis lineage began at least 12.7 million years ago. However, using combined ND2 and c-mos data and the calibration estimated for pygopod lizards suggests the lineage is 35.4-40.74 million years old. Our results support the hypothesis that skinks colonized Tasmantis by over-water dispersal initially to New Caledonia, then to Lord Howe Island, and finally to New Zealand.
Asunto(s)
Lagartos/genética , Filogenia , Animales , Australia , ADN Mitocondrial/genética , Complejo I de Transporte de Electrón/genética , Evolución Molecular , Genes mos/genética , Lagartos/clasificación , Proteínas Mitocondriales/genética , Datos de Secuencia Molecular , Nueva Zelanda , Análisis de Secuencia de ADNRESUMEN
The analysis of a 581 bp section of the nuclear gene c-mos revealed multiple copies of putative functional sequences as well as pseudogenes in three closely related lacertid species Lacerta laevis, L. kulzeri and L. cyanisparsa. A phylogenetic analysis of c-mos in comparison with a molecular phylogeny based on the mitochondrial cytochrome b gene supports our findings. The study also provides new insights into the phylogenetic relationships of L. cyanisparsa and L. laevis. Pseudogenes of the three species share 11 single-nucleotide substitutions, a 1 bp deletion and a premature stop codon but differ by group-specific mutations. This result suggests that the c-mos gene has become duplicated and subsequently silenced already in the common ancestor of the three species. Sequence divergence suggests that the duplication and the loss of function occurred in the late Miocene/early Pliocene, i.e., about 5 million years ago. Indications of gene conversion are discussed. We suggest that future studies using c-mos for phylogenetic studies should provide evidence for the orthology of the sequences compared.
Asunto(s)
Dosificación de Gen/genética , Genes mos/genética , Variación Genética , Lagartos/genética , Filogenia , Seudogenes/genética , Animales , Secuencia de Bases , Teorema de Bayes , Cartilla de ADN , Lagartos/clasificación , Modelos Genéticos , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADN , Especificidad de la EspecieRESUMEN
Recently, the phylogenetic placement of the African slender snouted crocodile, Crocodylus cataphractus, has come under scrutiny and herein we address this issue using molecular and morphological techniques. Although it is often recognized as being a "basal" form, morphological studies have traditionally placed C. cataphractus within the genus Crocodylus, while molecular studies have suggested that C. cataphractus is very distinct from other Crocodylus. To address the relationship of this species to its congeners we have sequenced portions of two nuclear genes (C-mos 302bp and ODC 294bp), and two mitochondrial genes (ND6-tRNA(glu)-cytB 347bp and control region 457bp). Analyses of these molecular datasets, both as individual gene sequences and as concatenated sequences, support the hypothesis that C. cataphractus is not a member of Crocodylus or Osteolaemus. Examination of 165 morphological characters supports and strengthens our resurrection of an historic genus, Mecistops (Gray 1844) for cataphractus.
Asunto(s)
Caimanes y Cocodrilos/clasificación , Filogenia , Aldehído Oxidorreductasas/genética , Caimanes y Cocodrilos/anatomía & histología , Caimanes y Cocodrilos/genética , Animales , Núcleo Celular/genética , Citocromos b/genética , ADN Mitocondrial/genética , Genes mos/genética , NADH Deshidrogenasa/genética , Ornitina Descarboxilasa/genéticaRESUMEN
Xenopus oocyte maturation is analogous to G2/M transition and characterized by germinal vesicle breakdown (GVBD), spindle formation, activation of MPF and Mos-Xp42(Mpk1) pathways. It is accompanied prior to GVBD by a transient increase in intracellular pH. We determined that a well known acidifying compound, NH(4)Cl, delayed progesterone-induced GVBD in a dose-dependent manner. GVBD(50) was delayed up to 2.3-fold by 10 mM NH(4)Cl. Cyclin B2 phosphorylation, Cdk1 Tyr15 dephosphorylation as well as p39(Mos) accumulation, Xp42(Mpk1) and p90(Rsk) phosphorylation induced by progesterone were also delayed by incubation of oocyte in NH(4)Cl. The delay induced by NH(4)Cl was prevented by injection of MOPS buffer pH 7.7. In contrast to acidifying medium, alkalyzing treatment such as Tris buffer pH 9 injections, accelerated GVBD, MPF and Xp42(Mpk1) activation, indicating that pHi changes control early steps of G2/M dynamics. When injected in an immature recipient oocyte, egg cytoplasm triggers GVBD through MPF auto-amplification, independently of protein synthesis. In these conditions, GVBD and Xp42(Mpk1) activation were delayed by high concentration of NH(4)Cl, which never prevented or delayed MPF activation. Strickingly, NH(4)Cl strongly inhibited thiophosphorylated active MAPK-induced GVBD and MPF activation. Nevertheless, Tris pH 9 did not have any effects on egg cytoplasm- or active MAPK-induced GVBD. Taken together, our results suggest that dynamic of early events driving Xp42(Mpk1) and MPF activation induced by progesterone may be negatively or positively regulated by pH(i) changes. However Xp42(Mpk1) pathway was inhibited by acidification alone. Finally, MPF auto-amplification loop was not sensitive to pH(i) changes.
Asunto(s)
Equilibrio Ácido-Base/fisiología , Fase G2/fisiología , Factor Promotor de Maduración/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Oocitos/citología , Oocitos/metabolismo , Progesterona/metabolismo , Proteínas de Xenopus/metabolismo , Cloruro de Amonio/metabolismo , Cloruro de Amonio/farmacología , Animales , Citoplasma/efectos de los fármacos , Citoplasma/metabolismo , Femenino , Fase G2/efectos de los fármacos , Genes mos/genética , Sistema de Señalización de MAP Quinasas/fisiología , Meiosis/efectos de los fármacos , Metafase/efectos de los fármacos , Fosforilación , Progesterona/farmacología , Proteínas Proto-Oncogénicas c-mos/metabolismo , Transducción de Señal , Xenopus laevisRESUMEN
During oogenesis, mRNA is actively transcribed and accumulated in growing oocytes, but this transcription stops before the oocytes grow to their full size. The accumulated maternal mRNA is used for protein synthesis in the oocytes during meiotic maturation and even in the embryos to sustain development after fertilization. Therefore, the degradation of accumulated maternal mRNA starts during meiotic maturation, but its rate is slow. Nevertheless, some mRNA species should rapidly degrade after fertilization if they encode proteins that play a role in specific events during meiosis and are detrimental for development after fertilization. In this study, to identify the selective degradation of maternal transcripts after fertilization, we sought mRNAs that are degraded in the early hours after fertilization by constructing an oocyte cDNA library after subtracting the cDNA of embryos at the mid one-cell stage. H1oo, c-mos, tPA (tissue type plasminogen activator gene), and Gdf9 were identified as genes whose transcripts undergo rapid degradation after fertilization. RT-PCR analysis showed that none of these transcripts was expressed during pre-implantation development once they were eliminated, suggesting that the mRNA species that are required for oogenesis, but not for early pre-implantation development, are degraded rapidly after fertilization. Microinjection of chimeric mRNAs in which the coding and 3'-untranslated regions (3'UTR) were exchanged between c-mos and hypoxanthine phosphoribosyltransferase mRNAs revealed that the 3'UTR plays a role in the rapid degradation that occurs after fertilization. Cytoplasmic polyadenylation elements (CPEs) was found near a poly(A) signal in the 3'UTR of all the mRNA species identified as rapidly degrading mRNA. The mechanism for the selective degradation is discussed, in relation to its biological significance.
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Embrión de Mamíferos/fisiología , Fertilización/fisiología , Regulación del Desarrollo de la Expresión Génica , Estabilidad del ARN/fisiología , Regiones no Traducidas 3'/genética , Animales , Northern Blotting , Proteína Morfogenética Ósea 15 , Cartilla de ADN , Embrión de Mamíferos/metabolismo , Femenino , Fertilización/genética , Biblioteca de Genes , Genes mos/genética , Factor 9 de Diferenciación de Crecimiento , Histonas , Péptidos y Proteínas de Señalización Intercelular/genética , Ratones , Oocitos/química , Embarazo , Proteínas/genética , Estabilidad del ARN/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo , Activador de Tejido Plasminógeno/genética , Factores de Escisión y Poliadenilación de ARNm/metabolismoRESUMEN
A 480-bp fragment of nuclear c-mos gene was sequenced in nine bird species representing four genera of the family Tetraonidae. It was demonstrated that nuclear genome region examined was highly conservative. The data were used to construct phylogenetic relationships among the c-mos gene sequences in Tetraonidae. The data obtained point to a paraphyletic origin of hazel grouse (Bonasa bonasia) and ruffed grouse (Bonasa umbellus).
Asunto(s)
Galliformes/genética , Genes mos/genética , Filogenia , Animales , Marcadores Genéticos/genéticaRESUMEN
Squamate reptiles (snakes, lizards, and amphisbaenians) serve as model systems for evolutionary studies of a variety of morphological and behavioral traits, and phylogeny is crucial to many generalizations derived from such studies. Specifically, the traditional dichotomy between Iguania (anoles, iguanas, chameleons, etc.) and Scleroglossa (skinks, geckos, snakes, etc.) has been correlated with major evolutionary shifts within Squamata. We present a molecular phylogenetic study of 69 squamate species using approximately 4600 (2876 parsimony-informative) base pairs (bp) of DNA sequence data from the nuclear genes RAG-1(approximately 2750 bp) and c-mos(approximately 360 bp) and the mitochondrial ND2 region (approximately 1500 bp), sampling all major clades and most major subclades. Under our hypothesis, species previously placed in Iguania, Anguimorpha, and almost all recognized squamate families form strongly supported monophyletic groups. However, species previously placed in Scleroglossa, Varanoidea, and several other higher taxa do not form monophyletic groups. Iguania, the traditional sister group of Scleroglossa, is actually highly nested within Scleroglossa. This unconventional rooting does not seem to be due to long-branch attraction, base composition biases among taxa, or convergence caused by similar selective forces acting on nonsister taxa. Studies of functional tongue morphology and feeding mode have contrasted the similar states found in Sphenodon(the nearest outgroup to squamates) and Iguania with those of Scleroglossa, but our findings suggest that similar states in Sphenodonand Iguania result from homoplasy. Snakes, amphisbaenians, and dibamid lizards, limbless forms whose phylogenetic positions historically have been impossible to place with confidence, are not grouped together and appear to have evolved this condition independently. Amphisbaenians are the sister group of lacertids, and dibamid lizards diverged early in squamate evolutionary history. Snakes are grouped with iguanians, lacertiforms, and anguimorphs, but are not nested within anguimorphs.
Asunto(s)
Filogenia , Reptiles/clasificación , Reptiles/genética , Animales , Composición de Base , Secuencia de Bases , Teorema de Bayes , Simulación por Computador , Cartilla de ADN , ADN Mitocondrial/genética , Genes RAG-1/genética , Genes mos/genética , Funciones de Verosimilitud , Modelos Genéticos , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADN , Especificidad de la EspecieRESUMEN
Phylogenetic studies incorporating multiple loci, and multiple genomes, are becoming increasingly common. Coincident with this trend in genetic sampling, model-based likelihood techniques including Bayesian phylogenetic methods continue to gain popularity. Few studies, however, have examined model fit and sensitivity to such potentially heterogeneous data partitions within combined data analyses using empirical data. Here we investigate the relative model fit and sensitivity of Bayesian phylogenetic methods when alternative site-specific partitions of among-site rate variation (with and without autocorrelated rates) are considered. Our primary goal in choosing a best-fit model was to employ the simplest model that was a good fit to the data while optimizing topology and/or Bayesian posterior probabilities. Thus, we were not interested in complex models that did not practically affect our interpretation of the topology under study. We applied these alternative models to a four-gene data set including one protein-coding nuclear gene (c-mos), one protein-coding mitochondrial gene (ND4), and two mitochondrial rRNA genes (12S and 16S) for the diverse yet poorly known lizard family Gymnophthalmidae. Our results suggest that the best-fit model partitioned among-site rate variation separately among the c-mos, ND4, and 12S + 16S gene regions. We found this model yielded identical topologies to those from analyses based on the GTR+I+G model, but significantly changed posterior probability estimates of clade support. This partitioned model also produced more precise (less variable) estimates of posterior probabilities across generations of long Bayesian runs, compared to runs employing a GTR+I+G model estimated for the combined data. We use this three-way gamma partitioning in Bayesian analyses to reconstruct a robust phylogenetic hypothesis for the relationships of genera within the lizard family Gymnophthalmidae. We then reevaluate the higher-level taxonomic arrangement of the Gymnophthalmidae. Based on our findings, we discuss the utility of nontraditional parameters for modeling among-site rate variation and the implications and future directions for complex model building and testing.