Transmembrane Helix Packing of ErbB/Neu Receptor in Membrane Environment: A Molecular Dynamics Study

Abstract

Dimerization or oligomerization of the ErbB/Neu receptors are necessary but not sufficient for initiation of receptor signaling. The two intracellular domains must be properly oriented for the juxtaposition of the kinase domains allowing trans-phosphorylation. This suggests that the transmembrane (TM) domain acts as a guide for defining the proper orientation of the intracellular domains.

Two structural models, with the two helices either in left-handed or in right-handed coiling have been proposed as the TM domain structure of the active receptor. Because experimental data do not distinguish clearly helix-helix packing, molecular dynamics (MD) simulations are used to investigate the energetic factors that drive Neu TM-TM interactions of the wild and the oncogenic receptor (Val664/Glu mutation) in DMPC or in POPC environments. MD results indicate that helix-lipid interactions in the bilayer core are extremely similar in the two environments and raise the role of the juxtamembrane residues in helix insertion and helix-helix packing. The TM domain shows a greater propensity to adopt a left-handed structure in DMPC, with helices in optimal position for strong inter-helical Hbonds induced by the Glu mutation. In POPC, the right-handed structure is preferentially formed with the participation of water in inter-helical Hbonds. The two structural arrangements of the NeuTM helices both with GG4 residue motif in close contact at the interface are permissible in the membrane environment. According to the hypothesis of a monomer-dimer equilibrium of the proteins it is likely that the bilayer imposes structural constraints that favor dimerization- competent structure responsible of the proper topology necessary for receptor activation.     

Stagnant immigrant social networks and cycles of exploitation

Based on over four years of ethnographic research among street vendors in Los Angeles and on interviews with family members of vendors and former vendors living in Mexico, this article examines the influence of a sending community and its social networks on migrant outcomes in the USA. These social networks affect migration patterns, ease entry into the fruit-vending business but also facilitate exploitation. Furthermore, these social networks do not always function as effective conduits of information because its members, due to feelings of shame or embarrassment, often fail to add to the existing body of knowledge. As a result, international migration patterns, job placement and exploitative practices do not change or improve for subsequent migrants. This creates a cycle in which social networks become stagnant and successively fail to function as effective conduits of information and resources in ways that might help network members equally and in the aggregate.     

The Archaeal Diversity and Population in a Drained Alkaline Saline Soil of the Former Lake Texcoco (Mexico)

Draining soil of the former Lake Texcoco, Mexico with pH > 10.0 and electrolytic conductivity (EC) > 100 dS m^?1 for 17 years has reduced pH to 7.8 and EC to 0.68 dS m^?1. Metagenomic DNA from the archaeal community was extracted directly from this soil and used as template to amplify the 16S ribosomal genes by PCR to construct gene libraries. Most of the cloned Archaea were related to mesophilic crenarchaeota and were not-yet-cultured. Sequence and phylogenetic analyses of these clones identified a group of Archaea with close affiliation to the ammonia-oxidizing Archaea. The cloned sequences from the drained soil diverged clearly from Haloarchaea found in the undrained soil from the lake.     

Isolation and Characterization of Mercury Resistant Bacillus sp. from Soils with an Extensive History as Substrates for Mercury Extraction in Mexico

Metal phytoextraction assisted by bacteria plays an important role in bioremediation systems. In this work, mercury-resistant bacterial strains were isolated from soils with high levels of mercury (San Joaquin, Queretaro State, Mexico) and identified as Bacillus sp. based on the 16S rDNA gene sequence analysis. The bacterial strains were found to exhibit different multiple mercury-resistance and carbon source utilization characteristics. The mercury reduction ability was tested through a volatilization assay. The bacterial isolates were also evaluated for their ability to promote growth and mercury uptake in tomato plants. In a roll towel assay, the maximum vigor index of tomato plants was obtained with the inoculation of Bacillus sp. A2, A12, B11, B15 and C1, while in a pot assay, the maximum vigor index was obtained with the inoculation of Bacillus sp. A6, A7 and B20, compared with un-inoculated controls in the presence of HgCl₂. Maximum Hg accumulation in the roots and shoots of tomato plants was obtained only with Bacillus sp. A7 in the roll towel assay, whereas in the pot assay, maximum accumulation was obtained with Bacillus sp. A12 compared with un-inoculated controls. Our results show that mercury accumulation in tissue is enhanced by these plant growth promoting bacterial strains, which recommends their possible use as microbe-assisted phytoremediation systems in mercury-polluted soils.     

cGMP-independent inhibition of integrin alphaIIbbeta3-mediated platelet adhesion and outside-in signalling by nitric oxide

We examined the influence of S-nitrosoglutathione (GSNO) on alpha(IIb)beta(3) integrin-mediated platelet adhesion to immobilised fibrinogen. GSNO induced a time- and concentration-dependent inhibition of platelet adhesion. Inhibition was cGMP-independent and associated with both reduced platelet spreading and protein tyrosine phosphorylation. To investigate the cGMP-independent effects of NO we evaluated integrin beta(3) phosphorylation. Adhesion to fibrinogen induced rapid phosphorylation of beta(3) on tyrosines 773 and 785, which was reduced by GSNO in a cGMP independent manner. Similar results were observed in suspended platelets indicating that NO-induced effects were independent of spreading-induced signalling. This is the first demonstration that NO directly regulates integrin beta(3) phosphorylation.     

Inflammation: a way to understanding the evolution of portal hypertension

Background

Portal hypertension is a clinical syndrome that manifests as ascites, portosystemic encephalopathy and variceal hemorrhage, and these alterations often lead to death.

Hypothesis

Splanchnic and/or systemic responses to portal hypertension could have pathophysiological mechanisms similar to those involved in the post-traumatic inflammatory response. The splanchnic and systemic impairments produced throughout the evolution of experimental prehepatic portal hypertension could be considered to have an inflammatory origin. In portal vein ligated rats, portal hypertensive enteropathy, hepatic steatosis and portal hypertensive encephalopathy show phenotypes during their development that can be considered inflammatory, such as: ischemia-reperfusion (vasodilatory response), infiltration by inflammatory cells (mast cells) and bacteria (intestinal translocation of endotoxins and bacteria) and lastly, angiogenesis. Similar inflammatory phenotypes, worsened by chronic liver disease (with anti-oxidant and anti-enzymatic ability reduction) characterize the evolution of portal hypertension and its complications (hepatorenal syndrome, ascites and esophageal variceal hemorrhage) in humans.

Conclusion

Low-grade inflammation, related to prehepatic portal hypertension, switches to high-grade inflammation with the development of severe and life-threatening complications when associated with chronic liver disease.     

Allelochemical Stress Can Trigger Oxidative Damage in Receptor Plants: Mode of Action of Phytotoxicity

Plants can interact with other plants through the release of chemical compounds or allelochemicals. These compounds released by donor plants influence germination, growth, development, and establishment of receptor plants; having an important role on the pattern of vegetation, i.e as invasive strategy, and on crop productivity. This phytotoxic or negative effect of the released allelochemicals (allelochemical stress) is caused by modifying or altering diverse metabolic processes, having many molecular targets in the receptor plants. Recently, using an aggressive and allelopathic plant Sicyos deppei as the donor plant, and Lycopersicon esculentum as the receptor plant, we showed that the allelochemicals released by S. deppei caused oxidative damage through an increase in reactive oxygen species (ROS) and activation or modification of antioxidant enzymes. Based on this study, we proposed that oxidative stress is one of the mechanisms, among others, by which an allelopathic plant causes phytotoxicity to other plants.Key Words: allelochemical stress, Sicyos deppei, Lycopersicon esculentum, plant allelochemicals, phytotoxicity, ROS, lipid peroxidationIt is well known that plants interact with many organisms, including co-habitation with other plants. Among these relations are the ones referred to as allelochemical interactions. Allelopathy can be defined as a mechanism of interference in plant growth and development mediated by the addition of plant-produced secondary products (allelochemicals) to the soil rhizosphere. Allelochemicals are present in all types of plants and tissues and are released into the soil rhizosphere by a variety of mechanisms, including decomposition of residues, volatilization, and root exudation.^1–3 These released allelochemicals become stressful only when they are toxic or when they affect the growth and development of surrounding plants (phytotoxicity). Studies on allelochemical stress have been expanding; recently the phenomenon has taken on increased importance, since it can help explain plant growth inhibition in interspecies interactions and in structuring the plant community. It appears to be one mechanism or strategy used by invasive plants to become successful and replace other native ones.^4–6On the other hand, the chemical diversity of the organic compounds that mediate these allelochemical interactions is as diverse as their modes of action. Many studies have shown that allelochemicals interfere with several physiological processes in the receptor organism.^3,7,8 The physiological effects on receptor plants or other organisms are useful in determining the role of the allelochemicals in the system. Recently, it has been proposed that allelochemicals can cause oxidative stress in target plants and therefore activate the antioxidant mechanism.^3,8–12 In particular; our studies have been focused on knowing the physiological targets of the phytotoxic compounds released by a noxious and endemic weed Sicyos deppei G. Don (Cucurbitaceae). We have taken as the model the receptor or damaged plant Lycopersicon esculentum Mill (Solanaceae), since in Mexican crop-fields, it is common to find both plants. We have observed the strong allelopathic potential of S. deppei and are exploring the potential metabolic target that could be involved in the strong phytotoxic effect of this weed.^13–16 We recently documented the oxidative damage that an aqueous leachate of S. deppei caused in the target plant L. esculentum.¹⁶ In this work we explored in seeds and in primary roots the antioxidant mechanism of tomato to determine whether or not the inhibitory effect of S. deppei was due to oxidative damage. We analyzed the activity and expression of some antioxidant enzymes involved in the detoxification of ROS, and found an imbalance in its activity as well as an increase in the levels of H₂O₂ at 24 h of treatment. Additional studies on the levels of ROS, including hydrogen peroxide, were monitored in primary roots from germinating seeds under allelochemical stress by imaging the ROS-sensitive fluorescent dye dichlorofluorescein (_H2DCFDA, carboxy-2′, 7′-diclhlorofluorescein diacetate) in a confocal microscope (BIORAD 1024, 488 nm dichroic and 510–560 nm emission). DCFDA fluorescence increases as the dye is oxidized by ROS to dichlorofluorescein (DCF). Figure 1 shows a marked increase in fluorescence at 48 h and 72 h of treatment (Fig. 1A–C) compared with the same treatment at 24 h, and with the corresponding control. This fluorescence was more evident at the root cap and at the zone of root hairs in treated seeds.Open in a separate windowFigure 1Allelochemical stress caused by S. deppei elicits ROS generation in tomato germinating seeds. Panels show control (left) and treatment (right) at 24 h (A), 48 h (B), and 72 h (C). Lower panels show higher magnification (40X) of the corresponding time. Seedlings with primary roots were stained for 10–15 minutes with 25 µM DCFDA in distilled water.Clearly, allelochemical stress caused by S. deppei is producing an oxidative imbalance as evidenced by generation of ROS and alteration of activity of antioxidant enzymes. Another result that supports this observation is the high level of lipid peroxidation that we observed at 48 and 72 h, which correlates with the inhibition of two membrane-associated enzymes, H⁺-ATPase¹⁵ and NADPH oxidase.¹⁶ We believe, however, that the oxidative damage we observed is not solely responsible for the phytotoxic effect of S. deppei on tomato growth. In other words, we suggest that its inhibitory effect represents the sum of many metabolic processes affected at different times. Currently we are studying the dynamics of carbohydrate mobilization, cell wall loosing of the endosperm to allow the protrusion of the radicle, and ABA content. Preliminary results have shown that there is a delay in expression of some enzyme activities and a high content of ABA.     

Kluyveromyces lactis sexual pheromones. Gene structures and cellular responses to alpha-factor

The Kluyveromyces lactis genes for sexual pheromones have been analyzed. The alpha-factor gene encodes a predicted polypeptide of 187 amino acid residues containing four tridecapeptide repeats (WSWITLRPGQPIF). A nucleotide blast search of the entire K. lactis genome sequence allowed the identification of the nonannotated putative a-pheromone gene that encodes a predicted protein of 33 residues containing one copy of the dodecapeptide a-factor (WIIPGFVWVPQC). The role of the K. lactis structural genes KlMFalpha1 and KlMFA1 in mating has been investigated by the construction of disruption mutations that totally eliminate gene functions. Mutants of both alleles showed sex-dependent sterility, indicating that these are single-copy genes and essential for mating. MATalpha, Klsst2 mutants, which, by analogy to Saccharomyces cerevisiae, are defective in Galpha-GTPase activity, showed increased sensitivity to synthetic alpha-factor and increased capacity to mate. Additionally, Klbar1 mutants (putatively defective in alpha-pheromone proteolysis) showed delay in mating but sensitivity to alpha-pheromone. From these results, it can be deduced that the K. lactis MATa cell produces the homolog of the S. cerevisiaealpha-pheromone, whereas the MATalpha cell produces the a-pheromone.