20,332 research outputs found
While K-ras is essential for mouse development, expression of the K-ras 4A splice variant is dispensable
No full textIn mammals, the three classical ras genes encode four highly homologous proteins, N-Ras, H-Ras, and the isoforms K-Ras 4A and 4B. Previous studies have shown that K-ras is essential for mouse development and that while K-ras 4A and 4B are expressed during development, K-ras 4A expression is regulated temporally and spatially and occurs in adult kidney, intestine, stomach, and liver. In the present study, the pattern of K-ras 4A expression was examined in a wide range of wild-type adult mouse tissues, and gene targeting was used to generate K-ras 4A-deficient mice to examine its role in development. It was found that K-ras 4A is also expressed in uterus, lung, pancreas, salivary glands, seminal vesicles, bone marrow cells, and cecum, where it was the major K-Ras isoform expressed. Mating between K-ras(tmDelta4A/+) mice produced viable K-ras(tmDelta4A/tmDelta4A) offspring with the expected Mendelian ratios of inheritance, and these mice expressed the K-ras 4B splice variant only. K-ras(tm-Delta4A/t-Delta4A) mice were fertile and showed no histopathological abnormalities on inbred (129/Ola) or crossbred (129/Ola x C57BL/6) genetic backgrounds. The results demonstrate that K-Ras 4A, like H- and N-Ras, is dispensable for normal mouse development, at least in the presence of functional K-Ras 4B
Ras-mediated phosphorylation of a conserved threonine residue enhances the transactivation activities of c-Ets1 and c-Ets2
No full textThe Ras oncogene products regulate the expression of genes in transformed cells, and members of the Ets family of transcription factors have been implicated in this process. To determine which Ets factors are the targets of Ras signaling pathways, the abilities of several Ets factors to activate Ras- responsive enhancer (RRE) reporters in the presence of oncogenic Ras were examined. In transient transfection assays, reporters containing RREs composed of Ets-AP-1 binding sites could be activated 30-fold in NIH 3T3 fibroblasts and 80-fold in the macrophage-like line RAW264 by the combination of Ets1 or Ets2 and Ras but not by several other Ets factors that were tested in the assay. Ets2 and Ras also superactivated an RRE composed of Ets-Ets binding sites, but the Ets-responsive promoter of the c-fms gene was not superactivated. Mutation of a threonine residue to alanine in the conserved amino-terminal regions of Ets1 and Ets2 (threonine 38 and threonine 72, respectively) abrogated the ability of each of these proteins to superactivate reporter gene expression. Phosphoamino acid analysis of radiolabeled Ets2 revealed that Ras induced normally absent threonine- specific phosphorylation of the protein. The Ras-dependent increase in threonine phosphorylation was not observed in Ets2 proteins that had the conserved threonine 72 residue mutated to alanine or serine. These data indicate that Ets1 and Ets2 are specific nuclear targets of Ras signaling events and that phosphorylation of a conserved threonine residue is a necessary molecular component of Ras-mediated activation of these transcription factors
Barbara Ras - Sowell Conference 2017
No full textBarbara Ras, San Antonio, Poet, author of "Bite Every Sorrow" and "The Last Skin
An increase in N-Ras expression is associated with development of hormone refractory prostate cancer in a subset of patients
No full textProtein expression of H, K and N-Ras was assessed in hormone sensitive and hormone refractory prostate tumour pairs from 61 patients by immunohistochemistry. Expression of H-Ras and K- Ras was not associated with any known clinical parameters. In contrast an increase in N-Ras membrane expression in the transition from hormone sensitive to hormone refractory prostate cancer was associated with shorter time to relapse (p=0.01) and shorter disease specific survival (p=0.008). In addition, patients with an increase in N-Ras membrane expression had lower levels of PSA at relapse (p=0.02) and expression correlated with phosphorylated MAP kinase (p=0.010) and proliferation index (Ki67, p=0.02). These results suggest that in a subgroup patients N-Ras expression is associated with development of hormone refractory prostate cancer via activation of the MAP kinase cascade
Ras signalling in the fission yeast Schizosaccharomyces pombe
No full textRas signalling is vital to many cellular processes. Ras proteins mediate a
vast array of cellular signalling networks, and are conserved from humans to
unicellular eukaryotes. The study of ras signalling in higher eukaryotes presents
a number of technical challenges, due to the presence of multiple ras isoforms,
regulatory proteins and activators. The fission yeast Sz. pombe represents an
ideal system for the investigation of ras signalling, as it contains a single, nonessential
ras protein (Ras1). In addition, Ras1 is involved in the regulation of
a number of downstream pathways.
A number of studies in recent years have highlighted the role of subcellular
localisation in ras signalling output. The localisation of Ras1 in Sz. pombe
has also been described as key in effector selection, with Ras1 at the plasma
membrane regulating mating and Ras1 at the endomembranes regulating cell
morphology. This thesis describes a series of studies utilising Ras1 mutants
and chimeric Ras1 proteins which display differing localisation patterns to determine
the role of Ras1 localisation in signalling. The data presented herein
support the notion of a revised model for the role of Ras1 localisation in signalling,
suggesting that the localisation of Ras1 to the plasma membrane is
key to all signalling events downstream of Ras1.
This thesis also describes the characterisation of oncogenic mutants of Ras1,
demonstrating the importance of signalling magnitude in functional output. In
addition, the importance of Ras1 regulation in cell viability and chromosome
stability is also demonstrated. Finally, the functional expression of three human
ras isoforms is described, validating the use of Sz. pombe as a model
system for the heterologous expression of human ras signalling components
K-Ras mediated murine epidermal tumorigenesis Is dependent upon and associated with elevated Rac1 activity
No full textA common goal for potential cancer therapies is the identification of differences in protein expression or activity that would allow for the selective targeting of tumor vs. normal cells. The Ras proto-oncogene family (K-Ras, H-Ras and N-Ras) are amongst the most frequently mutated genes in human cancers. As a result, there has been substantial effort dedicated to determining which pathways are activated by Ras signaling and, more importantly, which of these contribute to cancer. Although the most widely studied Ras-regulated signaling pathway is the Raf/mitogen-activated protein kinase cascade, previous research in model systems has revealed that the Rac1 GTP-binding protein is also required for Ras-induced biological responses. However, what have been lacking are rigorous in vivo Rac1 target validation data and a clear demonstration that in Ras-driven hyperplastic lesions, Rac1 activity is increased. Using a combination of genetically-modified mouse models that allow for the tissue-selective activation or deletion of signaling molecules and an activation-state sensitive Rac1 antibody that detects GTP-bound Rac1, we found that Rac1 contributes to K-Ras induced epidermal papilloma initiation and growth and that Rac1 activity is elevated by oncogenic K-Ras in vivo. Previously, it was not practical to assess Rac1 activation status in the most commonly used format for clinical tumor specimens, formalin-fixed paraffin embedded (FFPE) tissues samples. However, this study clearly demonstrates that Rac1 is essential for K-Ras driven epithelial cell hyperproliferation and that Rac1 activity is elevated in tissues expressing mutant oncogenic K-Ras, while also characterizing the activation-state specific Rac1-GTP antibody as a probe to examine Rac1 activation status in FFPE samples. Our findings will facilitate further research on the status of Rac1 activity in human tumors and will help to define the tumor types of the patient population that could potentially benefit from therapies targeting Rac activation or downstream effector signaling pathway
Cigarette smoking and K-ras mutations in pancreas, lung and colorectal adenocarcinomas: etiopathogenic similarities, differences and paradoxes.
No full textSurprisingly different frequencies and patterns of K-ras mutations are observed in human adenocarcinomas of the pancreas, colorectum and lung. Their respective relationships with smoking are apparently paradoxical. We evaluated all the available types of clinical and epidemiological studies on the relationship between tobacco smoking and the occurrence of K-ras mutations in human adenocarcinomas of the pancreas, colorectum and lung. We identified 8, 7 and 12 studies that analyzed the relationship between K-ras mutations and tobacco smoking in human neoplasms of the pancreas, colorectum and lung, respectively. A meta-analysis was undertaken for each site separately. In pancreatic adenocarcinomas lifetime history of tobacco consumption was not significantly associated with the frequency of K-ras mutations (OR=1.26; 95% CI=0.82-1.94). Similarly, no association was observed between smoking and K-ras mutations in colorectal adenocarcinomas (OR=0.94; CI=0.79-1.12), neither when colorectal adenomas and adenocarcinomas were jointly analyzed (OR=0.96; 95% CI=0.83-1.13). In lung adenocarcinoma, where only 15-25% of cases harbor a K-ras mutation, tumors from smokers were more likely to have K-ras mutations than tumors from non-smokers (OR=3.67; 95% CI=2.47-5.45). Furthermore, in lung adenocarcinomas K-ras mutations have a pattern different from that in pancreatic and colorectal adenocarcinomas. Results support the hypothesis that smoking influences the risk of pancreatic cancer - and possibly colorectal cancer - through events other than K-ras mutations. In adenocarcinoma of the lung, smoking may play a role in the occurrence of K-ras mutations. If the influence of tobacco products in the induction, acquisition and persistence of K-ras mutations had some tissue specificity, or was dependent on different factors in different organs, the corresponding mechanisms would deserve detailed research
Transcription factor 8 activates R-Ras to regulate angiogenesis
No full textWe have recently reported that transcription factor 8 (TCF8) negatively regulates pathological angiogenesis by regulating endothelial invasiveness by acting as a transcriptional attenuator of matrix metalloproteinase 1. TCF8 also modulates cell–matrix and cell–cell adhesion; however molecular mechanism of this TCF8 function remains obscure. Here, we provide evidence that TCF8 activates R-Ras, another class of angiogenic regulator, to suppress angiogenesis by a mechanism other than a transcriptional attenuator. Tube formation by human umbilical vein endothelial cells (HUVECs) facilitated by TCF8 suppression was significantly inhibited by the expression of onstitutive active mutant of R-Ras. When we examined the mRNA expression levels of R-Ras regulators, no significant changes were observed to explain the R-Ras activation by TCF8. Interestingly, we found that TCF8 bound to CalDAG-GEFIII, an R-Ras activator, in the cytosol, indicating that TCF8 emanates signaling for R-Ras activation from cytosol to regulate angiogenesis negatively
R-Ras ihon karsinogeneesissä
No full textR-Ras on Ras-perheeseen kuuluva pieni GTPaasi, joka muistuttaa rakenteellisesti muita Ras-perheen geenejä. Useat näistä muista Ras-perheen geeneistä ovat tunnettuja proto-onkogeenejä, joiden aktivoivat mutaatiot kykenevät aiheuttamaan normaalin solun transformaation syöpäsoluksi. Näiden Ras-geenien mutaatioiden on arvioitu olevan keskeisessä osassa useissa eri syövissä, n. 50 % kaikista syöpätapauksista.
Huolimatta rakenteellisista yhtäläisyyksistä ko. tunnettuihin Ras onkogeeneihin, R-Ras geeniä aktivoivien mutaatioiden ei ole kuitenkaan osoitettu aiheuttavan syöpää. Sen sijaan proteiinin aktivaation on esitetty hillitsevän syöpäsoluille tyypillistä proliferaatiota ja invasiivistä kasvua. Tämän tutkimuksen tarkoituksena oli selvittää R-Ras:n roolia kemiallisesti indusoidussa ihon kasvainmallissa. Käyttämällä poistogeenistä hiirikantaa (R-Ras KO) ja vertailemalla sitä normaaliin hiirikantaan havaitsimme, että R-Ras on merkittävä tekijä epidermaalisten kasvainten kehittymisessä.
19 viikkoa kestäneessä kokeessa perimältään normaaleilla (villi-tyyppi, WT) eläimille kehittyi lähes kuusinkertainen määrä kasvaimia verrattuna poistogeenisiin eläimiin. Kasvaimet myös ilmaantuivat aiemmin ja kasvoivat suuremmiksi villi-tyypin eläimillä. Vaikka R-Ras ilmentyi voimakkaasti ihon dermaalisissa verisuonissa, sitä ei havaittu sekä normaalissa että karsinogeeneillä käsitellyssä epidermiksessä, jossa kasvaimet manifestoituivat. Nämä seikat antavat ymmärtää, että huolimatta aiemmin kuvatuista antionkogeenisistä ominaisuuksistaan, R-Ras vaikuttaisi olevan merkittävässä roolissa kasvaimen kehittymisessä (ihon kokeellisessä, kemiallisessa karsinogeneesi-mallissa) mekanismeillä, jotka mahdollisesti liittyvät kasvainkudoksen immunologiseen säätelyyn
Comparison of K-Ras and N-Ras Mutagenic Hot Spots for UVC Damage
No full textIt has been well established that mutations in K-Ras and N-Ras proto-oncogenes can convert them into active oncogenes. Current molecular cancer research has been focused on determining the key steps by which cellular genes become oncogenes and not on the underlying and fundamental chemical damage mechanism and susceptibility to damage. In this study, we investigate the damage hot spots present in the N-Ras and K-Ras genes upon exposure to UVC radiation. Detection of damage is accomplished by a simple, sensitive, mix-and-read assay using an EvaGreen probe in a 96-well microtiter plate. Our results show that, although there is high degree of sequential similarities among K-Ras and N-Ras genes, they show different degrees of UV damage in different portions of their genomes. Our experiments demonstrate that overall, the K-Ras genome is more prone to UVC damage than the N-Ras genome. We observe that the extent of damage increases with increasing number of TTs in a sequence, consistent with previous results that show that thymine cyclobutyl photodimers are the primary DNA damage photoproducts upon UVC irradiation. This understanding of the effect of UVC radiation on various codons of K-Ras and N-Ras genes will help to increase our understanding about hot spots of DNA damage and the chemical damage mechanism
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