Search

A new operational ocean forecast system, the Atlantic Margin Model implementation of the Forecast Ocean Assimilation Model (FOAM-AMM), has been developed for the European North West Shelf (NWS). An overview of the system is presented including shelf specific developments of the physical model, the Nucleus for European Modelling of the Ocean (NEMO), and the Sea Surface Temperature (SST) data assimilation scheme. Initial validation is presented of the tides and model SST. The SST skill of the system is significantly improved by the data assimilation scheme. Finally, an analysis of the seasonal tidal mixing fronts shows that these, in general, agree well with observation, but data assimilation does not significantly alter their positions.

The breast cancer predisposition genes, BRCA1 and BRCA2, are responsible for the vast majority of hereditary breast cancer. Although BRCA2 functions to help the cell repair double-stranded DNA breaks, the function of BRCA1 remains enigmatic. Here, we develop a human genetic system to study the role of BRCA1 in oxidative DNA damage. We show that human cancer cells containing mutated BRCA1 are hypersensitive to ionizing radiation. This hypersensitivity can be reversed by the expression of forms of BRCA1 that are not growth suppressing. Reversal of hypersensitivity requires the ring finger of BRCA1, its transactivation domain, and its BRCT domain. Lastly, we show that unlike BRCA2, BRCA1 does not function in the repair of double-stranded DNA breaks. Instead, it functions in transcription-coupled DNA repair (TCR). TCR ability correlated with radioresistance as cells containing BRCA1 showed both increased TCR and radioresistance, whereas cells without BRCA1 showed decreased TCR and radiosensitivity. These findings give physiologic significance to the interaction of BRCA1 with the basal transcription machinery.

The effects of upper and lower boundaries on stratified shear flows with horizontally uniform velocity and density profiles are investigated through tilting tank experiments. Two geometries are considered: first horizontal boundaries at unequal distances from the shear layer; and second boundaries sloping at an angle, ø, in the spanwise direction. Asymmetric boundary separation is shown to retard the growth of Kelvin-Helmholtz billows and delay their transition to turbulence. However when the billows do evolve, they are shown to migrate vertically away from the closer boundary. This results in the billows causing mixing to a similar vertical extent as in the case of symmetrically placed boundaries, although this mixing is limited to patches in the streamwise direction. Boundaries tilted in the spanwise direction introduce a cross-tank slope to the orientation of the billows. This slope is found to vary linearly with ø, as predicted by Thorpe and Holt (1995). Of the 102 individual billows observed, only 21 evolve with a purely two-dimensional orientation, and 65 of the 81 sloping billows are oriented in the predicted sense. The across-lank structure of the billows is found to divide between two distinct regimes depending on the values of ø and H, where H is the depth of the tank scaled by the characteristic width of the density interface, L. In cases where H is large and ø small, the billows evolve with approximately spanwise sections displaced in the streamwise direction either side of the centre of the tank. These are connected by a sloping region of reduced amplitude. When H is small the whole of the billows slope across the tank with little Variation in their orientation. In both cases vortex tubes are commonly observed connecting neighbouring billows. These are always observed to slope in the opposite sense to the billow themselves.

An increasing body of evidence suggests that mitogen-induced activation of the RAF/ERK signaling pathway is functionally separate from the stress-induced activation of the SEK/JNK/p38 signaling pathway. In general, stress stimuli strongly activate the p38s and the JNKs while only weakly activating ERK1 and ERK2. However, a number of independent groups have now shown that the RAF/ERK signaling pathway is strongly activated by ionizing radiation. In this work, we examine this paradox. We show that both mitogen-activated protein (MAP) kinase kinase 1 (MEK1) and MAP kinase kinase 2 (MEK2) are activated by ionizing radiation. Blockage of this activation through the use of dominant negative MEK2 increases sensitivity of the cell to ionizing radiation and decreases the ability of a cell to recover from the G2/M cell cycle checkpoint arrest. Blocking MEK2 activation does not affect double-strand DNA break repair, however. Although MEK1 is activated to a lesser extent by ionizing radiation, expression of a dominant negative MEK1 does not affect radiation sensitivity of the cell, the G2/M checkpoint of the cell, or double-strand break repair. Because ionizing radiation leads to a different cell cycle arrest (G2/M arrest) than that typically seen with other stress stimuli, and because we have shown that MEK2 can affect G2/M checkpoint kinetics, these results provide an explanation for the observation that the MEKs can be strongly activated by ionizing radiation and only weakly activated by other stressful stimuli.

A 23-year-old man presented with signs of increased intracranial pressure. CAT scan, cisternogram, and angiogram on admission were reported to be normal. Lumbar puncture revealed elevated pressure and protein of 500 mg-%. CSF cytology revealed malignant tumor cells. A brain biopsy and decompression were carried out to reveal diffuse subarachnoid invasion by malignant tumor cells. Immunohistochemical studies using anti-glial fibrillary acidic protein serum revealed tumor cells to be positive for GFA protein. A lumbosacral CAT scan 9 days after surgery revealed numerous sclerotic densities involving bony pelvis, sacrum, T-12 vertebra and left femoral head. No definite primary site of tumor was found antemortem in the brain or any other organ. Autopsy demonstrated diffuse subarachnoid spread of malignant tumor in brain and spinal cord. Malignant astrocytoma was found in the midline pons projecting into the 4th ventricle. The histology of the neoplasm was identical in all sites including bone. No other tumor was found at autopsy. The vascular invasion by the tumor cells in the pons and distant bony metastasis in this case suggest hematogenous spread. A review of the lumbar X-ray taken 6 weeks prior to admission and the presence of well-established bony lesions within 10 days of craniotomy suggest that this is a rare case of extraneural metastasis of glioma occurring prior to surgery. The midline location of clinically occult pontine glioma and the presence of bony metastasis created difficulty in the diagnosis but immunohistochemical studies proved to be crucial in establishing correct diagnosis antemortem.

Two distinct regions of the transforming growth factor-β1 (TGF-β1) promoter are responsive to autoregulation and activation by phorbol ester (12-O-tetradecanoylphorbol-13-acetate): sequences located between nucleotides −454 to −323 (first promoter) and between the two transcriptional start sites. We have now characterized in detail the induction of the second promoter (sequences between nucleotides +1 to +271) of the TGF-β1 gene by both TGF-β1 and phorbol ester. By assaying progressively deleted mutations in the second promoter, we have found two regions responsible for the induction; each contains a phorbol ester-responsive element. In vitrotranscription of the second promoter-chloramphenicol acetyltransferase chimeric genes using nuclear extracts of A-549 cells showed that deletion of the putative phorbol ester-responsive elements results in a 70–80% decrease in activity. DNase I footprinting and gel mobility shift assays showed that binding to an Sp1 site and the putative TRE elements are required for maximal expression of the second promoter region of the TGF-β1 gene. These results suggest that AP-1, which is capable of conferring phorbol ester or TGF-β1 responsiveness, is the major transcription factor involved in the second promoter-derived transcription of the TGF-β1 gene.

Finkel-Biskis-Reilly (FBR) osteosarcoma virus v-Fos causes tumors of mesenchymal origin, including osteosarcomas, rhabdomyosarcomas, chondrosarcomas, and liposarcomas. Because the cell of origin in all these tumors is a pluripotent mesenchymal cell, the variety of tumors seen in mice which express FBR v-Fos implies that FBR v-Fos inhibits multiple differentiation pathways. To study the mechanism of FBR v-Fos' inhibition of mesenchymal differentiation, we utilized an in vitro model of adipocyte differentiation. We show by both morphological and biochemical means that FBR v-Fos inhibits adipocyte differentiation in vitro. This inhibition is due to FBR v-Fos' inhibition of the growth arrest characteristic of terminal differentiation and FBR v-Fos' inhibition of the expression and activity of a key regulator of this growth arrest, C/EBPalpha. The in vitro inhibition of adipogenesis by FBR v-Fos has in vivo significance as immunostaining of FBR v-Fos-induced tumors shows no CCAAT/enhancer binding protein (EBP)-alpha expression. These data implicate C/EBPalpha as a protein involved in the generation of liposarcomas.

Histopathologic study of human breast biopsy samples has identified specific lesions which are associated with a high risk of development of invasive breast cancer. Presumably, these lesions (collectively termed premalignant breast disease) represent the earliest recognizable morphologic expression of fundamental molecular events that lead to the development of invasive breast cancer. To study molecular events underlying premalignant breast disease, we have developed a method for isolating RNA from histologically identified lesions from frozen human breast tissue. This method specifically obtains mRNA from breast epithelial cells and has identified three genes which are differentially expressed in premalignant breast epithelial lesions. One gene identified by this method is overexpressed in four of five noncomedo ductal carcinoma in situ lesions and appears to be the human homologue of the gene encoding the M2 subunit of ribonucleotide reductase, an enzyme involved in DNA synthesis.

The myristylated v-fos product, FBR murine sarcoma virus (Gag-Fos) protein, exhibits a lower level of transrepression of the serum response element (SRE) than does c-fos protein (Fos). Mutation of the N-terminal myristylation site in FBR protein restored SRE transrepression. Replacement of N-terminal viral Gag sequences with the Fos N terminus also restored this activity, providing additional evidence that myristylation inhibits transrepression by FBR protein. However, the myristylated Gag domain did not inhibit SRE transrepression when fused to Fos, indicating that myristylation of a fos protein is not by itself sufficient to prevent SRE transrepression and that C-terminal mutation is necessary to inhibit transrepression by N myristylation. Comparison of transfection results with Fos C-terminal deletion mutants and the Fos/FBR chimeric mutant revealed that the FBR C terminus retained the potential for transrepression despite deletion of the normal Fos C terminus, whereas similar Fos deletion mutants did not. These results indicate that both N- and C-terminal mutations are required to inhibit transrepression by FBR protein and that multiple structural mutations accompanied by posttranslational protein modification alter gene regulation by FBR protein.

In our studies of indicators and putative precursors of breast cancer we have chosen to use the term 'premalignant' rather than more broadly constructed terms with similar intent such as 'neoplastic' because malignancy has the intrinsic meaning of indicating potential mortality and morbidity in humans. Along with discussions of anatomic determinants of premalignancy in women, we will discuss the potentially important role of the hereditary breast cancer genes, particularly BRCA1. BRCA1 certainly plays a central role in the special hereditary setting where it seems a specific and sufficient cause for the occurrence of breast cancer in women. As a part of this focus on morbidity and mortality in women we will include an emphasis on the earliest and most determinant points in the development of breast cancer derived from many studies of possible precursor lesions and indicators of risk elevation derived from epidemiologic and clinical studies.There are obviously

DNA damage is recognized as a central component of carcinogenesis. DNA-damaging agents activate a number of signal transduction pathways that lead to repair of the DNA, apoptosis, or cell cycle arrest. It is reasoned that a cell deficient in DNA repair is more likely to acquire other cancer-promoting mutations. Despite the recent interest in the link between DNA damage and carcinogenesis, retroviral oncogenes have not yet been shown to affect the DNA damage-signaling pathway. In this report, we show that Finkel-Biskis-Reilly mouse osteosarcoma virus (FBR) v-fos, the retroviral homologue of the c-fos proto-oncogene, inhibits the cellular response to ionizing radiation. Cells that express FBR v-Fos show a decreased ability to repair DNA damage caused by ionizing radiation, and these cells show decreased survival in response to ionizing radiation. In addition, FBR v-Fos inhibits DNA-dependent protein kinase, a kinase specifically activated upon exposure to ionizing radiation. These effects were specific to ionizing radiation, as no effect of FBR v-Fos on the UV light signaling pathway was seen. Last, these effects were dependent on a lipid modification required for FBR v-Fos tumorigenesis, that of myristoylation of FBR v-Fos. A non-myristoylated mutant FBR v-Fos caused none of these effects. This study suggests that a retroviral oncogene can lead to an increased genomic instability, which can ultimately increase the carcinogenic potential of a cell.

Serum induces the expression of a number of proteins with similar transcriptional properties, including those encoded by the proto-oncogenes c-fos and c-jun. This study employs a novel antisense rescue method to determine whether antisense-resistant genes (constructed by deletion of antisense RNA target sequences) can replace c-fos expression during serum-induced DNA synthesis. Immunoprecipitation studies and nuclease protection assays demonstrated that anti-fos RNA inhibited endogenous c-fos expression but did not inhibit expression of transfected antisense-resistant mutant c-fos genes. The results of nuclear-labelling and cellular-proliferation studies indicated that C terminally truncated Fos mutants, including FBR v-fos, could not rescue endogenous Fos, although full-length and minimally truncated c-fos expression vectors could restore serum-induced DNA synthesis in cells expressing anti-fos RNA. Overexpression of c-Jun protein (Jun) could not restore serum-induced DNA synthesis to cells expressing inducible anti-fos RNA despite equivalent transactivation of an AP-1 target gene. Thus, the antisense rescue method defines a specialized function for c-Fos protein which is distinct from the function(s) of Jun and/or transforming FBR v-Fos proteins.

To study the role of a nuclear proto-oncogene in the regulation of cell growth and differentiation, we inhibited HL-60 c-myc expression with a complementary antisense oligomer. This oligomer was stable in culture and entered cells, forming an intracellular duplex. Incubation of cells with the anti-myc oligomer decreased the steady-state levels of c-myc protein by 50 to 80%, whereas a control oligomer did not significantly affect the c-myc protein concentration. Direct inhibition of c-myc expression with the anti-myc oligomer was associated with a decreased cell growth rate and an induction of myeloid differentiation. Related antisense oligomers with 2- to 12-base-pair mismatches with c-myc mRNA did not influence HL-60 cells. Thus, the effects of the antisense oligomer exhibited sequence specificity, and furthermore, these effects could be reversed by hybridization competition with another complementary oligomer. Antisense inhibition of a nuclear proto-oncogene apparently bypasses cell surface events in affecting cell proliferation and differentiation.

In our studies of indicators and putative precursors of breast cancer we have chosen to use the term 'premalignant' rather than more broadly constructed terms with similar intent such as 'neoplastic' because malignancy has the intrinsic meaning of indicating potential mortality and morbidity in humans. Along with discussions of anatomic determinants of premalignancy in women, we will discuss the potentially important role of the hereditary breast cancer genes, particularly BRCA1. BRCA1 certainly plays a central role in the special hereditary setting where it seems a specific and sufficient cause for the occurrence of breast cancer in women. As a part of this focus on morbidity and mortality in women we will include an emphasis on the earliest and most determinant points in the development of breast cancer derived from many studies of possible precursor lesions and indicators of risk elevation derived from epidemiologic and clinical studies.There are obviously

Antisense RNA complementary to c-fos mRNA was produced in mouse 3T3 cells by gene transfer techniques. Transcriptional units were constructed consisting of a steroid-inducible mouse mammary tumor virus (MMTV) promoter, mouse or human 5' c-fos gene fragments in either the sense (5' to 3') or antisense (3' to 5') orientation, and splice and poly(A) signals from the human beta-globin gene. A gene that confers neomycin resistance was included in the vectors to allow isolation of stable transformants. Dexamethasone caused a marked induction of hybrid MMTV-fos-globin RNA. Induction of the hybrid transcript containing antisense c-fos RNA decreased colony formation following DNA transfer and inhibited the proliferation of cells into which the antisense transcriptional unit had been integrated. In contrast, colony formation and cell proliferation were not inhibited by induction of hybrid RNA containing c-fos RNA sequences in the sense orientation. These results indicate that the strategy of generating antisense RNA to inhibit gene expression may be useful in delineating the function of protooncogenes. The c-fos gene product appears to have a required role in normal cell division.

Transforming growth factor beta 1 (TGF-beta 1) is a potent growth inhibitor for many cell types, including most epithelial cells. However, the mechanism of growth inhibition is unknown. In skin keratinocytes, TGF-beta 1 has been shown to inhibit growth and to rapidly reduce c-myc expression. It has been demonstrated that protein synthesis is required for TGF-beta 1 regulation of c-myc in keratinocytes. Here we present evidence that treatment of mouse BALB/MK keratinocyte cells with either antisense c-myc oligonucleotides or TGF-beta 1 inhibited cell entry into S phase. These results suggest that TGF-beta inhibition of c-myc expression may be essential for growth inhibition by TGF-beta 1. The block in c-myc expression by TGF-beta 1 occurred at the level of transcriptional initiation. Studies with a series of 5' deletion c-myc/chloramphenicol acetyltransferase constructs indicated that a cis regulatory element(s), which resides between positions -100 and +71 relative to P1 transcription start site, is responsible for the TGF-beta 1 responsiveness. Based on these data, it is proposed that the mechanism of TGF-beta 1 growth inhibition involves synthesis or modification of a protein that may interact with a specific element(s) in the 5' regulatory region of the c-myc gene, resulting in inhibition of transcriptional initiation.

FBR murine sarcoma virus (gag-fos) protein, a virally transduced Fos protein, exhibits decreased gene transactivation in comparison with the cellular Fos protein. Biochemical analysis suggests that myristylation of the virally encoded N-terminal gag region results in decreased DNA binding and transcriptional activation without affecting heterodimerization with Jun protein. These findings demonstrate that protein myristylation can modulate gene regulation by a DNA-binding protein.