R-Ras promotes focal adhesion formation through focal adhesion kinase and p130(Cas) by a novel mechanism that differs from integrins

R-Ras regulates integrin function, but its effects on integrin signaling pathways have not been well described. We demonstrate that activation of R-Ras promoted focal adhesion formation and altered localization of the alpha2beta1 integrin from cell-cell to cell-matrix adhesions in breast epithelial cells. Constitutively activated R-Ras(38V) dramatically enhanced focal adhesion kinase (FAK) and p130(Cas) phosphorylation upon collagen stimulation or clustering of the alpha2beta1 integrin, even in the absence of increased ligand binding. Signaling events downstream of R-Ras differed from integrins and K-Ras, since pharmacological inhibition of Src or disruption of actin inhibited integrin-mediated FAK and p130(Cas) phosphorylation, focal adhesion formation, and migration in control and K-Ras(12V)-expressing cells but had minimal effect in cells expressing R-Ras(38V). Therefore, signaling from R-Ras to FAK and p130(Cas) has a component that is Src independent and not through classic integrin signaling pathways and a component that is Src dependent. R-Ras effector domain mutants and pharmacological inhibition suggest a partial role for phosphatidylinositol 3-kinase (PI3K), but not Raf, in R-Ras signaling to FAK and p130(Cas). However, PI3K cannot account for the Src-independent pathway, since simultaneous inhibition of both PI3K and Src did not completely block effects of R-Ras on FAK phosphorylation. Our results suggest that R-Ras promotes focal adhesion formation by signaling to FAK and p130(Cas) through a novel mechanism that differs from but synergizes with the alpha2beta1 integrin.     

Environmental and hormonal control of vernal migration in redheaded bunting (Emberiza bruniceps)

This paper reviews the factors and mechanisms which result in the development of the metabolic state characteristic of migration with special reference to a palaeotropic migrant the redheaded bunting,Emberiza bruniceps. Changes in climatic conditions and food supply act as proximate triggers of migratory behaviour in partial migrants. Typical migrants like buntings use daylength as a cue but the exact mechanism of how photoperiodic information is translated in terms of migratory events is still not known. Almost entirely the photoperiodic effects have been explained on the basis of the involvement of hypothalamo/hypophyseal system. We feel mechanism(s) other than those acting through neuroendocrine system may be equally important. Furthermore the role of temperature has not been adequately explored so far. Our observations indicate the possibility that redheaded buntings might integrate the information received from photoperiod with environmental temperature (and other factors?) resulting in the development of migratory state. The physiological control of avian migration is much less understood. Majority of papers have centered around the ‘gonadal hypothesis’ of Rowan supporting or contradicting it without providing conclusive evidence. Pituitary prolactin has also been shown to be implicated although the mechanism of action is only speculative. Conclusive evidence for the involvement of thyroid hormones (thyroxine, T4; triiodothyronine, T3) in the physiological timing of migration has been produced attributing independent roles to T4 and T3. It is suggested that seasonal variation in peripheral conversion of T4 to T3 could serve as an effective strategy to render available the required thyroid hormones T4 and/or T3 during different phases of the year thus accounting for the metabolic switch over from T4-dependent moult to T3-dependent migratory fat deposition and zugunruhe and also ensuring preclusion of simultaneous occurrence of these mutually incompatible events. Considering that the number of environmental and physiological factors influence this mechanism and considering that thyroid hormone molecule has been put to a wide range of usage during the course of evolution the mechanism(s) of peripheral conversion of T4-T3 may assume great flexibility and have selective value-especially in migration which is known to have evolved several times in diverse avian families. The attractiveness of this hypothesis lies in the fact that it has potential to explain the both physiological development of the metabolic state of migration and at the same time the physiological timing of migration not only with respect to the cycle of environment but also with respect to other conflicting seasonal events (moult and reproduction).     

Tuning of intercalation and electron-transfer processes between DNA and acridinium derivatives through steric effects

A series of acridinium derivatives 1-6, wherein steric factors have been varied systematically through substitution at the 9 position of the acridine ring, have been synthesized and their DNA interactions have been investigated by various biophysical techniques. The unsubstituted and methylacridinium derivatives 1 and 2 and the o-tolylacridinium derivative 6 exhibited high fluorescence quantum yields (Phi(f)() congruent with 1) and lifetimes (tau = 35, 34, and 25 ns, respectively), when compared with the arylacridinium derivatives 3-5. The acridinium derivatives 1 and 2 showed high DNA binding affinity (K = 7.3-7.7 x 10(5) M(-)(1)), when compared to the arylacridinium derivatives 3-5 (K = 6.9-10 x 10(4) M(-)(1)). DNA melting and viscosity studies establish that in the case of the aryl-substituted systems, the efficiency of DNA binding is in the order, phenyl > p-tolyl > m-tolyl > o-tolyl derivative. The increase in steric crowding around the acridine ring hinders the DNA binding interactions and thereby leads to negligible binding as observed in the case of 6 (o-tolyl derivative). These results indicate that a subtle variation in the substitution pattern has a profound influence on the photophysical and DNA interactions. Further, they demonstrate that pi-stacking interactions of the ligands with DNA are essential for efficient electron transfer between the DNA bases and the ligands. These water soluble and highly fluorescent molecules which differ in their DNA binding mode can act as models to study various DNA-ligand interactions.     

Drug-resistant breast carcinoma (MCF-7) cells are paradoxically sensitive to apoptosis

The purpose of this study was to determine whether expression of tissue transglutaminase (TG2) and caspase-3 proteins in drug-resistant breast carcinoma MCF-7/DOX cells would render these cells selectively susceptible to apoptotic stimuli. Despite high resistance to multidrug resistance (MDR)-related drug, doxorubicin (> or =150-fold), the MCF-7/DOX cells were extremely sensitive to apoptotic stimuli. Thus, calcium ionophore, A23187 (A23187) and the protein kinase C inhibitor staurosporine (STS) each induced rapid and time-dependent apoptosis in MCF-7/DOX cells. The apoptosis induced by either agent was accompanied by caspase-3 activation and other downstream changes that are typical of cells undergoing apoptosis. The alterations upstream of caspase-3 activation, however, such as loss in mitochondrial membrane potential (DeltaPsi), release of cytochrome c, and activation of caspase-8, and caspase-9, were detected only in STS-treated cells. The A12387 failed to induce any of the caspase-3 upstream changes, implying that A23187-induced apoptosis may utilize one or more novel upstream pathways leading to the activation of caspase 3. In summary, these data demonstrate that MCF-7/DOX cells are much more sensitive to apoptotic stimuli than previously thought and that A23187-induced apoptosis may involve some novel, yet unidentified, upstream pathway that leads to the activation of caspase-3 and other downstream events.     

Bilateral high upper limb replantation in a child

Bilateral high amputation of upper limbs in a child is a very unusual injury. In the present case, although the amputation was high and significant avulsion was present, the age of the child (6 years) made the case both challenging and encouraging--challenging because of the anticipated systemic effects of reperfusion, and encouraging because the long-term prognosis is always more encouraging in a child.     

An enzyme immunoassay for detection of Japanese encephalitis virus-induced chemotactic cytokine

Japanese encephalitis virus (JEV) induces human peripheral blood monocytes to secrete a chemotactic cytokine [human macrophage-derived factor (hMDF)] which causes chemotaxis of neutrophils. The only known assay for hMDF cannot quantify its level in samples, so an enzyme immunoassay has been standardized for detection of hMDF and hMDF-specific antibodies in test samples. The reported enzyme linked immunosorbent assay (ELISA) was found to be sensitive (89%), specific (91%), accurate (92 2%) and reproducible and was able to detect a minimum concentration of 23 ng hMDF/ml in test samples. The chemotactic factor could be detected in JEV inoculated mouse sera and JEV infected culture fluids. Significant finding of the test was the detection of hMDF in sera of human cases of JE.     

Interactions with pocket proteins contribute to the role of human papillomavirus type 16 E7 in the papillomavirus life cycle

Human papillomaviruses (HPVs), most commonly the HPV16 genotype, are the principle etiological determinant for cervical cancer, a common cancer worldwide resulting in over 200,000 deaths annually. The oncogenic properties of HPVs are attributable in part to the virally encoded protein E7, best known for its ability to bind to and induce the degradation of the retinoblastoma tumor suppressor, pRb, and related "pocket proteins" p107 and p130. Previously, we defined a role for E7 in the productive stage of the HPV16 life cycle, which takes place in stratified squamous epithelia. HPV perturbs the normal processes of cell growth and differentiation of stratified squamous epithelia. HPVs reprogram cells to support continued DNA synthesis and inhibit their differentiation in the suprabasal compartment of the epithelia, where cells normally have withdrawn from the cell cycle and initiated a well-defined pattern of terminal differentiation. These virus-induced perturbations, which contribute to the production of progeny HPVs, are dependent on E7. In this study, we define the mechanism of action by which E7 contributes to the productive stage of the HPV16 life cycle. We found that the ability of HPV16 to reprogram suprabasal cells to support DNA synthesis correlates with E7's ability to bind pocket proteins but not its ability to induce their degradation. In contrast, the ability of HPV16 to perturb differentiation correlated with both E7's binding to and degradation of pocket proteins. These data indicate that different hallmarks of the productive stage of the HPV16 life cycle rely upon different sets of requirements for E7.     

The probable significance of tracheal tufts in the 8th abdominal segment of Heliothis virescens (F.) on the development of its parasitoid, Toxoneuron nigriceps (Viereck)

The 8th abdominal segment of Heliothis virescens (Fabricius) larvae contains aerating trachea and tracheole tufts that end in the hemocoel of the 8th segment, unlike the tracheae that invade tissues in other segments. These tracheal tufts from the 8th abdominal segment extend to the tokus region, which along with the telson cavity is known to act as a “lung” for hemocytes in Calpodes ethlius and a few other lepidopteran larvae. The goal of this research was to study the effects of these tracheal tufts in the 8th abdominal segment on parasitoid development inside the host larvae, H. virescens. The first objective was to determine if the eggs of the parasitoid, Toxoneuron nigriceps, are predominantly located among the tracheal tufts of the 8th abdominal segment compared to other body cavity regions irrespective of their oviposition site or the position of the host larvae. The results showed that several hours after oviposition most of the eggs are found in the 8th abdominal segment irrespective of the oviposition site or the position of the host larvae. The second objective was to study the effect of varying oxygen concentrations in vitro on various developmental stages of the egg. The results showed that decreasing oxygen concentrations adversely affects the parasitoid egg development in vitro. A third objective was to determine the oxygen concentration in 8th abdominal segment of the host larvae and compare it to other regions of the body using an oxygen sensor placed in vivo. The results suggested relatively high concentration of oxygen in the 8th abdominal segment compared to other regions of the host, thus supporting our hypothesis that the increased oxygen level in the 8th abdominal segment is important to the development of the parasitoid eggs.