SEARCH · PubMed Health

Results for “Enhancer mapping”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 577 records · Page 32Linked to original sources

PubMed1986

Regulation of microtubule composition and stability during nerve growth factor-promoted neurite outgrowth.

We have used the nerve growth factor (NGF)-responsive line of PC12 pheochromocytoma cells as a model system to study microtubule specializations associated with neurite outgrowth. PC12 cells treated with NGF cease proliferating and extend neurites. Long-term NGF treatment results in a two- to threefold increase in the proportion of total cellular tubulin that is polymerized in PC12 cells. The increase in this parameter first becomes apparent at 2-4 d with NGF and increases steadily thereafter. Several changes in microtubule-associated proteins (MAPs) of PC12 cells also occur after exposure to NGF. In immunoprecipitation assays, we observed the levels of MAP-2 to increase by at least several-fold after treatment with NGF. We also found that the compositions of three MAP classes with apparent Mr of 64K, 67K, and 80K are altered by NGF treatment. These MAPs, recently designated "chartins," are biochemically and immunologically distinct from the similarly-sized tau MAPs (Peng et al., 1985 Brain Res. 361: 200; Magendantz and Solomon, 1985 Proc. Natl. Acad. Sci. 82: 6581). In two-dimensional isoelectric focusing x SDS polyacrylamide gels, each chartin MAP class resolves into a set of proteins of similar apparent Mr but distinct pI. Peptide mapping analyses confirm that the isoelectric variants comprising each chartin MAP class are closely related in primary structure. Several striking differences in the composition of the chartin MAPs of PC12 cells grown with or without NGF were consistently observed. In particular, following longterm NGF treatment, the abundances of the more acidic variants of each chartin MAP class were markedly enhanced relative to the more basic members. This occurs without substantial changes in the abundance of each MAP class as a whole relative to total cell protein. The combined results of in vivo phosphorylation and peptide mapping experiments indicate that the NGF-inducible chartin MAP species are not primary translation products, but are generated posttranslationally, apparently by differential phosphorylation of other chartin MAPs. These observations suggest that NGF treatment of PC12 cells leads to changes in the posttranslational processing of the chartin MAPs. The time course of these changes closely resembles that for the increase in the proportion of cellular tubulin that is polymerized and for neurite outgrowth. One of the important events in the growth and stabilization of neurites appears to be the formation of microtubule bundles that extend from the cell body to the tips of the neurites.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed2001

Effect of vasodilator hydralazine on tumor microvascular random flow and blood volume as measured by intravoxel incoherent motion (IVIM) weighted MRI in conjunction with Gd-DTPA-Albumin enhanced MRI.

We studied the effect of hydralazine on tumor blood volume fraction and microvascular random flow velocity magnitude by IVIM weighted MRI in conjunction with dynamic Gd-DTPA-Albumin enhanced MRI. Blood volume fraction maps were obtained from the dynamic Gd-DTPA-Albumin enhanced MRI measurements. The average blood volume fraction of R3230 AC adenocarcinoma decreased from 0.125 +/- 0.022 (s.d.) ml/g to 0.105 +/- 0.018 (s.d.) ml/g (p < 0.001) after the administration of hydralazine at a dose of 5 mg/kg. The microvascular random flow velocity magnitude maps were obtained from the IVIM weighted MRI signals by utilizing the Gd-DTPA-Albumin measured blood volume fractions as an input in the compartmental modeling analysis of the IVIM weighted MRI signal. The random-directional microvascular flow induced MRI signal attenuation was separated from the molecular diffusion induced signal attenuation. Flow induced attenuation was more significant after the administration of hydralazine. The mean microvascular random flow velocity magnitude increased from 0.52 +/- 0.15 (s.d.) mm/sec to 0.73 +/- 0.23 (s.d.) mm/sec (p < 0.05) in the presence of the above blood volume fraction change.

Adenocarcinoma↗

PubMed2005

Role of nitric oxide in modulating systemic pressor responses to different vasoconstrictors in man.

OBJECTIVE: Animal studies suggest that nitric oxide (NO) attenuates responses to endogenous vasoconstrictors. We investigated whether this also holds true in man by monitoring pressor responses to different vasoconstrictors during nitric oxide synthase (NOS) inhibition. METHODS: Systemic hemodynamic responses to intravenous infusions of three doses (each for 5 min) of angiotensin II (AngII) (2, 4 and 8 ng/kg per min), noradrenaline (NOR) (10, 30 and 70 ng/kg per min) and phenylephrine (PE) (0.5, 1.0 and 1.5 microg/kg per min) were monitored in 44 healthy subjects during saline. A second dose-response curve was obtained during NOS inhibition with a subpressor dose of N- nitro-L-arginine-methyl ester (L-NAME) (5 microg/kg per min) or during a systemic NO clamp using combined systemic infusions of L-NAME (12.5 microg/kg per min) and nitroprusside. Blood pressure was measured in the brachial artery and other hemodynamic parameters were derived from this signal. RESULTS: Mean arterial pressure (MAP) increased 2 +/- 2, 6 +/- 1 and 16 +/- 2 mmHg in response to AngII during saline, 7 +/- 6, 15 +/- 5 and 26 +/- 6 mmHg during the subpressor dose of L-NAME (P < 0.05) and 11 +/- 10, 18 +/- 7 and 25 +/- 6 mmHg during the systemic NO clamp (P < 0.001). These augmented responses of MAP were due to enhanced increments in systemic vascular resistance. Infusions of NOR and PE during saline resulted in dose-dependent increments in MAP and systemic vascular resistance. These increments were of comparable magnitude as those seen during AngII, but were not affected by NOS inhibition. CONCLUSION: Our findings show that the systemic pressor response evoked by AngII, but not by NOR or PE, is enhanced during NOS inhibition, suggesting that AngII is associated with increased NO release that counteracts its blood pressure rising effect.

Adult↗

PubMedDate not supplied

From acquisition to consolidation: on the role of brain-derived neurotrophic factor signaling in hippocampal-dependent learning.

One of the most rigorously investigated problems in modern neuroscience is to decipher the mechanisms by which experience-induced changes in the central nervous system are translated into behavioral acquisition, consolidation, retention, and subsequent recall of information. Brain-derived neurotrophic factor (BDNF) has recently emerged as one of the most potent molecular mediators of not only central synaptic plasticity, but also behavioral interactions between an organism and its environment. Recent experimental evidence indicates that BDNF modulates synaptic transmission and plasticity by acting across different spatial and temporal domains. BDNF signaling evokes both short- and long-term periods of enhanced synaptic physiology in both pre- and postsynaptic compartments of central synapses. Specifically, BDNF/TrkB signaling converges on the MAP kinase pathway to enhance excitatory synaptic transmission in vivo, as well as hippocampal-dependent learning in behaving animals. Emerging concepts of the intracellular signaling cascades involved in synaptic plasticity induced through environmental interactions resulting in behavioral learning further support the contention that BDNF/TrkB signaling plays a fundamental role in mediating enduring changes in central synaptic structure and function. Here we review recent literature showing the involvement of BDNF/TrkB signaling in hippocampal-dependent learning paradigms, as well as in the types of cellular plasticity proposed to underlie learning and memory.

Animals↗

PubMed1982

In vivo mapping of local cerebral blood flow by xenon-enhanced computed tomography.

A noninvasive technique has been developed to measure and display local cerebral blood flow (LCBF) in vivo. In this procedure, nonradioactive xenon gas is inhaled and the temporal changes in radiographic enhancement produced by the inhalation are measured by sequential computerized tomography. The time-dependent xenon concentrations in various anatomical units in the brain are used to derive both the local partition coefficient and the LCBF. Functional mapping of blood flow with excellent anatomical specificity has been obtained in the baboon brain. The response of LCBF to stimuli such as changes in carbon dioxide concentrations as well as the variability in LCBF in normal and diseased tissue can be easily demonstrated. This method is applicable to the study of human physiology and pathologic blood flow alterations.

Animals↗

PubMed2001

An inhibitory role of the phosphatidylinositol 3-kinase-signaling pathway in vascular endothelial growth factor-induced tissue factor expression.

Vascular endothelial growth factor (VEGF) is not only essential for vasculogenesis and angiogenesis but is also capable of inducing tissue factor, the prime initiator of coagulation, in endothelial cells. In this study we have analyzed the VEGF-elicited pathways involved in the induction of tissue factor in human umbilical cord vein endothelial cells. Using specific low molecular weight inhibitors we could demonstrate a crucial role of the p38 and Erk-1/2 mitogen-activated protein (MAP) kinases. In contrast, treatment with wortmannin or LY294002, inhibitors of phosphatidylinositol 3 (PI3)-kinase, resulted in a strong enhancement of the VEGF-induced tissue factor production, indicating a negative regulatory role of the PI3-kinase on tissue factor-inducing pathways. Accordingly, transduction with constitutively active Akt led to a reduction of VEGF-induced tissue factor production. Western blot analyses using antibodies specific for phosphorylated p38 showed an enhanced activation of this MAP kinase in human umbilical cord vein endothelial cells when stimulated with VEGF in the presence of wortmannin in comparison to either agent alone. Thus, the negative regulation of the PI3-kinase pathway on endothelial tissue factor activity can be explained at least in part by a suppression of this MAP kinase-signaling pathway. This is the first demonstration of a reciprocal relationship between procoagulant activity and the PI3-kinase-Akt signaling pathway, and it reveals a novel mechanism by which tissue factor expression can be controlled in endothelial cells.

Adenoviridae↗

PubMed1994

Role of IRS-1-GRB-2 complexes in insulin signaling.

GRB-2 is a small SH2- and SH3 domain-containing adapter protein that associates with the mammalian SOS homolog to regulate p21ras during growth factor signaling. During insulin stimulation, GRB-2 binds to the phosphorylated Y895VNI motif of IRS-1. Substitution of Tyr-895 with phenylalanine (IRS-1F-895) prevented the IRS-1-GRB-2 association in vivo and in vitro. The myeloid progenitor cell line, 32-D, is insensitive to insulin because it contains few insulin receptors and no IRS-1. Coexpression of IRS-1 or IRS-1F-895 with the insulin receptor was required for insulin-stimulated mitogenesis in 32-D cells, while expression of the insulin receptor alone was sufficient to mediate insulin-stimulated tyrosine phosphorylation of Shc and activation of p21ras and mitogen-activated protein (MAP) kinase. The Shc-GRB-2 complex formed during insulin stimulation is a possible mediator of p21ras and MAP kinase activation in IRS-1-deficient 32-D cells. Interestingly, IRS-1, but not IRS-1F-895, enhanced the stimulation of MAP kinase by insulin in 32-D cells expressing insulin receptors. Thus, IRS-1 contributes to the stimulation of MAP kinase by insulin, probably through formation of the IRS-1-GRB-2 complex at Tyr-895. Our results suggest that the Shc-GRB-2 complex and the activation of p21ras-dependent signaling pathways, including MAP kinase, are insufficient for insulin-stimulated mitogenesis and that the essential function(s) of IRS-1 in proliferative signaling is largely unrelated to IRS-1-GRB-2 complex formation.

Adaptor Proteins, Signal Transducing↗

PubMed2004

Location of the epitope for an anti-CD8alpha antibody 53.6.7 which enhances CD8alpha-MHC class I interaction indicates antibody stabilization of a higher affinity CD8 conformation.

MHC class I tetramers are widely used, usually in combination with an antibody to CD8, to detect antigen specific T cells. Some anti-CD8alpha antibodies block the interaction of murine MHC class I tetramers with CD8 T cells, while others such as 53.6.7, enhance. To understand the molecular basis for this effect, we mapped the epitope for the enhancing antibody 53.6.7 and three other blocking antibodies using a panel of murine CD8alpha (Lyt-2) mutants expressed on COS-7 transfectants. Mutations in residues that contact MHC class I affected binding of the blocking antibodies. In contrast, antibody 53.6.7 was affected by a mutation in the residue T81A located on the D-E loop. In the cocrystal of CD8alphaalpha with MHC class I, two different complexes (A and B) were observed, indicating the existence of different CD8 conformations. The T81 residue does not make contact with MHC class I in either complex, however, neighboring residues in the D-E loop make very different contacts in the two different complexes. The most likely explanation for antibody enhancement of tetramer bindings is that binding of 53.6.7 to CD8alphabeta stabilizes a conformation with a higher affinity for interaction with MHC class I and suggests that the CD8 binding site is flexible.

Animals↗

PubMed1994

Molecular genetic methods for mapping disease genes.

Recent technological enhancements in molecular genetics have significantly improved our ability to locate disease genes by linkage analysis. Despite the introduction of molecular methods such as PCR and the discovery of highly polymorphic SSRs, genotyping is still rate-limiting for localizing disease genes by linkage. These methods have been, until recently, highly technical, time-consuming, and expensive. The challenge of mapping the major genes that render one susceptible to environmentally initiated lung disorders such as asthma will require efficient and highly accurate methods of genotyping. We have shown that genotyping by semi-automated fluorescence-based techniques is both highly accurate and efficient. These methods may improve productivity by more than an order of magnitude and can be easily adapted to most linkage studies.

Asthma↗

PubMed2003

Development and genetic mapping of 127 new microsatellite markers in barley.

To enhance the marker density of existing genetic maps of barley ( Hordeum vulgare L.), a new set of microsatellite markers containing dinucleotide motifs was developed from genomic clones. Out of 254 primer pairs tested, a total of 167 primer pairs were classifed as functional in a panel of six barley cultivars and three H. spontaneum accessions, and of those, 127 primer pairs resulting in 133 loci were either mapped or located onto chromosomes. The polymorphism information content (PIC) ranged from 0.05 to 0.94 with an average of 0.60. The number of alleles per locus varied from 1 to 9. On average, 3.9 alleles per primer pair were observed. The RFLP frameworks of two previously published linkage maps were used to locate a total of 115 new microsatellite loci on at least one mapping population. The chromosomal assignment of 48 mapped loci was corroborated on a set of wheat-barley chromosome addition lines; 18 additional loci which were not polymorphic in the mapping populations were assigned to chromosomes by this method. The microsatellites were located on all seven linkage groups with four significant clusters in the centromeric regions of 2H, 3H, 6H and 7H. These newly developed microsatellites improve the density of existing barley microsatellite maps and can be used in genetic studies and breeding research.

Chromosome Mapping↗

PubMed2000

Predictors of success in radiofrequency catheter ablation of atrial flutter.

Radiofrequency catheter ablation of typical atrial flutter at the isthmus between the tricuspid annulus and the inferior vena cava is established. However in selected patients, the creation of a continuous linear lesion at the targeted isthmus requires a lengthened procedure or is not feasible at all and atrial flutter recurrences are common. In a retrospective analysis, we found that an intraoperatively determined distance between the tricuspid annulus and the inferior vena cava of <.2.5 cm is an independent predictor of a lengthened or failed ablation procedure. Additional equipment, e.g., long introducer sheaths, adapted ablation catheter design, or irrigated tip ablation, as well as alternative ablation approaches, e.g., linear lesions between the tricuspid annulus and Eustachian ridge, have been invented in order to increase the acute success rate or decrease fluoroscopy and procedure time. In a prospective study on the effects of various conduction properties at the isthmus between tricuspid annulus and inferior vena cava following radiofrequency ablation of atrial flutter, we showed previously that others than a complete bidirectional conduction block predicts a high recurrence rate of atrial flutter. For determination of transisthmal conduction properties following ablation, established mapping approaches are documentation of double potentials at the ablation line and right atrial activation sequence following posteroseptal and low lateral right atrial pacing. Novel threedimensional mapping systems, i.e., Carto(R) and EnSite(R), may further enhance the accuracy of conventional mapping techniques.

Atrial Flutter↗

PubMed1994

Perceptual sensitivity maps within globally defined visual shapes.

An unsolved problem of biology is the processing of global shape in natural vision. The known processes of early vision are spatially restricted (or local) operations, and little is known about their interactions in organizing the visual image into functionally coherent (or global) objects. Here we introduce a human psychophysical method which allows us to measure the effect of perceptual organization on the activity pattern of local visual detectors. We map differential contrast sensitivity for a target across regions enclosed by a boundary. We show that local contrast sensitivity is enhanced within the boundary even for large distances between the boundary and the target. Furthermore, the locations of maximal sensitivity enhancement in the sensitivity maps are determined by global shape properties. Our data support a class of models which describe shapes by the means of a medial axis transformation, implying that the visual system extracts 'skeletons' as an intermediate-level representation of objects. The skeletal representation offers a structurally simplified shape description which can be used for higher-level operations and for coding into memory.

Contrast Sensitivity↗

PubMed1993

Enhanced cytochrome-oxidase staining of the cuneate nucleus in the rat reveals a modifiable somatotopic map.

Existing cytochrome oxidase (CO)-staining techniques were modified to enhance sensitivity and contrast in order to examine patterns of CO-activity in the dorsal column nuclei (DCN) of adult Long-Evans rats. Within a rostrocaudally limited region in the middle of the cuneate nucleus (CN) distinctive blotches of intense CO-activity were observed. The CO-staining was maximally differentiated approximately 0.3-0.7 mm caudal to the obex. No CO-blotches were observed anywhere else in the DCN. Transganglionic labelling (WGA-HRP) demonstrated that some of the CO-blotches in the rat CN are related to the terminal projection fields of primary afferents from the skin of the forepaws. The corresponding location of primary afferent termination fields and CO-staining patterns supports a tripartite rostrocaudal division in the rat CN, similar to that described by other investigators in cats, monkeys and raccoons. Comparing the patterns of CO-staining to (1) the cytoarchitecture (Nissl-stained sections), or to (2) the dendritoarchitecture (distribution of microtubule-associated protein 2 (MAP2) or to (3) the organization of retrogradely labelled (WGA-HRP/HRP) cuneothalamic cells, revealed no topographical organization corresponding to the CO-blotches. Postnatal (at least up to 11 days postpartum) forepaw deafferentation or removal disrupted the CO-staining pattern in the CN.

Animals↗

PubMed2004

Enhanced virtual bronchoscopy using the pulmonary artery: improvement in route mapping for ultraselective transbronchial lung biopsy.

OBJECTIVE: We evaluated a new simulation method for ultraselective transbronchial lung biopsy using the pulmonary artery. MATERIALS AND METHODS: A new method for enhanced virtual bronchoscopy using the pulmonary artery was developed for ultraselective transbronchial lung biopsy. In a volunteer study of healthy adults, three radiologists with different levels of experience independently reconstructed conventional virtual bronchoscopy and enhanced virtual bronchoscopy using the pulmonary artery until reaching the farthest point of the bronchus and pulmonary artery. The bronchovascular branch order and the minimum luminal diameter (e.g., for bronchus and artery) for reconstruction were compared. In a clinical study, virtual bronchoscopy and enhanced virtual bronchoscopy using the pulmonary artery were compared with regard to accessibility to target lesions in 40 patients with small pulmonary nodules or ground-glass opacities. A comparison between the simulated bronchial route reconstructions and actual bronchoscopic routes on biopsy was made to determine the efficacy of each reconstruction method. RESULTS: In the volunteer study, quality of enhanced virtual bronchoscopy using the pulmonary artery was not significantly affected by the experience levels of the radiologists. In the clinical study, bronchial reconstruction was successful in guiding to a bronchoscopic tumor in 35 (87.5%) of 40 cases. The maximum bronchial order on reconstruction was the sixth for the virtual bronchoscopy group and the eighth for the group with enhanced virtual bronchoscopy using the pulmonary artery (p < 0.001, Wilcoxon's signed rank test). The bronchial route reconstructed on enhanced virtual bronchoscopy using the pulmonary artery agreed with the actual biopsy routes in 30 cases (85.7%), but those reconstructed on virtual bronchoscopy alone agreed in only eight cases (22.9%) (p < 0.001, chi-square test). CONCLUSION: Enhanced virtual bronchoscopy using the pulmonary artery is feasible and helpful for ultraselective transbronchial lung biopsy of small nodules in the lung.

Aged↗

PubMed1987

Enhancer of seizure: a new genetic locus in Drosophila melanogaster defined by interactions with temperature-sensitive paralytic mutations.

Mutations in the enhancer of seizure (e(sei] locus have been isolated on the basis of their ability to cause temperature-induced paralysis of alleles at the seizure (sei) locus at temperatures at which these mutations ordinarily do not paralyze. This enhancer is specific to the seizure locus and is without effect on other temperature-sensitive paralytic mutants including para, nap, tip-E and shi. This suggests that the enhancer responds specifically to the mechanism of paralysis mediated by the seizure mutations. The e(sei) is a recessive mutation which maps to 39.0 on the left arm of chromosome 3. Deficiency mapping has placed it at 69A4-B5 on the salivary gland polytene chromosome map. When a new enhancer allele was isolated following P-M hybrid dysgenesis, there was a concomitant P-element insertion at 69B. In the absence of seizure mutations, the enhancer mutation causes non-temperature dependent hyperactivity when agitated and interferes with the climbing response. Electrophysiological studies examined the effects of increasing temperature on electrical activity in the adult giant fiber/flight muscle system. Neuronal hyperactivity was seen in both e(sei) and sei single mutant homozygotes, but not in wild type. The hyperactivity was more severe in the sei;e(sei) double mutants. The correlation between the physiological effects and the mutant behavior suggests that both sei and e(sei) cause membrane excitability defects. Since previous work has shown that seizure mutants affect [3H]saxitoxin binding to the voltage-sensitive sodium channel, e(sei) may code for a gene product which interacts with this channel.

Alleles↗

PubMed1995

End-to-end annealing of plant microtubules by the p86 subunit of eukaryotic initiation factor-(iso)4F.

The p86 subunit of eukaryotic initiation factor-(iso)4F from wheat germ exhibits saturable and substoichiometric binding to maize microtubules, induces microtubule bundling in vitro, and is colocalized or closely associated with cortical microtubule bundles in maize root cells, indicating its function as a microtubule-associated protein (MAP). The effects of p86 on the growth of short, taxol-stabilized maize microtubules were investigated. Pure microtubules underwent a gradual length redistribution, an increase in mean length, and a decrease in number concentration consistent with an end-to-end annealing mechanism of microtubule growth. Saturating p86 enhanced the microtubule length distribution and produced significantly longer and fewer microtubules than the control, indicating a facilitation of annealing by p86. Confirmation of endwise annealing rather than of dynamic instability as the mechanism for microtubule growth was made using mammalian MAP2, which also promoted the redistribution of length, increase in mean length, and decrease in number concentration of taxol-stabilized maize microtubules. Enhancement of microtubule growth occurred concomitant with bundling by p86, indicating that an alignment of microtubules in bundles facilitated endwise annealing kinetics. The results demonstrate that nonfacile plant microtubules can spontaneously elongate by endwise annealing and that MAPs enhance the rate of annealing. The p86 subunit of eukaryotic initiation factor-(iso)4F may be an important regulator of microtubule dynamics in plant cells.

Animals↗

PubMed2004

Health and environment information systems for exposure and disease mapping, and risk assessment.

A large number of chemicals are used on a regular basis in modern society. Thousands of new chemicals are added each year, many of which may have toxic properties constituting potential health hazards. Rapid assessment of the risk associated with the use of these chemicals is therefore essential to protect people from exposure to potentially harmful substances. Exposures to chemicals (and physical agents) are typically unevenly distributed geographically as well as temporally. Disease occurrence also shows geographically varying patterns. Geographic information systems (GIS) may be used to produce maps of exposure and/or disease to reveal spatial patterns. Exposure mapping using advanced GIS modeling may enhance exposure assessment in environmental epidemiology studies. Disease maps can be valuable tools in risk assessment to explore changes in disease patterns potentially associated with changes in environmental exposures. Spatial variations in risk and trends related to distance from pollution sources may be studied using software tools such as the Rapid Inquiry Facility, developed by the U.K. Small Area Health Statistics Unit and enhanced in the European Health and Environment Information System project, for an initial quick evaluation of any potential health hazards associated with an environmental pollutant.

Cross-Sectional Studies↗

PubMed1995

Rapid activation of mitogen-activated protein kinase and p21ras by prolactin and interleukin 2 in rat Nb2 node lymphoma cells.

Several serine/threonine and tyrosine kinase signal transduction pathways have been recently linked to prolactin (PRL) action in lymphoid cells. Utilizing the lactogen-dependent, rat pre-T lymphoma cell line, Nb2-11, and the autonomous subline, Nb2-SFJCD1, studies were conducted to determine whether PRL- or interleukin-2 (IL-2)-stimulated Nb2 cell proliferation is coupled to the activation of p21ras and mitogen-activated protein (MAP) kinase. Stimulation of Nb2-11 cells, growth-arrested in the early G1 phase of the cell cycle, with PRL or IL-2 rapidly (5-10 min) provoked GTP binding to Ras, enhanced tyrosyl phosphorylation of MAP kinase, significantly increased its enzymatic activity, and caused its nuclear translocation. The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), which directly activates protein kinase C, similarly activated Ras and MAP kinase but failed to cause its nuclear translocation. Tyrosine kinase antagonism with genistein inhibited PRL-stimulated Ras and MAP kinase activation. In other experiments, Ras and MAP kinase were each found to be constitutively active in the Nb2-SFJCD1 line. The addition of PRL to these cultures enhanced the activity of these signaling proteins. Finally, the effects of PRL, IL-2, TPA, and phosphatase inhibition on Nb2-11 cell population density and [3H]thymidine uptake were compared. The addition of PRL, IL-2, and TPA significantly stimulated[3H] thymidine incorporation, while only the polypeptide growth factors augmented cell density. Phosphatase inhibition had no effect on either parameter. These results indicate that Nb2 cell proliferation is associated with the early activation of Ras and MAP kinase. Moreover, tyrosyl phosphorylation upstream of Ras activation appears to be required for its subsequent stimulation of mediators, which activate MAP kinase. Protein kinase C activation may be coupled to MAP kinase activation but is not sufficient for Nb2 cell proliferation.

Animals↗