The Relative Expression of Mig6 and EGFR Is Associated with Resistance to EGFR Kinase Inhibitors

The sensitivity of only a few tumors to anti-epidermal growth factor receptor EGFR tyrosine kinase inhibitors (TKIs) can be explained by the presence of EGFR tyrosine kinase (TK) domain mutations. In addition, such mutations were rarely found in tumor types other than lung, such as pancreatic and head and neck cancer. In this study we sought to elucidate mechanisms of resistance to EGFR-targeted therapies in tumors that do not harbor TK sensitizing mutations in order to identify markers capable of guiding the decision to incorporate these drugs into chemotherapeutic regimens. Here we show that EGFR activity was markedly decreased during the evolution of resistance to the EGFR tyrosine kinase inhibitor (TKI) erlotinib, with a concomitant increase of mitogen-inducible gene 6 (Mig6), a negative regulator of EGFR through the upregulation of the PI3K-AKT pathway. EGFR activity, which was more accurately predicted by the ratio of Mig6/EGFR, highly correlated with erlotinib sensitivity in panels of cancer cell lines of different tissue origins. Blinded testing and analysis in a prospectively followed cohort of lung cancer patients treated with gefitinib alone demonstrated higher response rates and a marked increased in progression free survival for patients with a low Mig6/EGFR ratio (approximately 100 days, P = 0.01).     

Nonlinear optical imaging to evaluate the impact of obesity on mammary gland and tumor stroma

Obesity is an established risk factor for breast cancer incidence and mortality. However, the mechanism that links obesity to tumorigenesis is not well understood. Here we combined nonlinear optical imaging technologies with an early-onset diet-induced obesity breast cancer animal model to evaluate the impact of obesity on the composition of mammary gland and tumor stroma. Using coherent anti-Stokes Raman scattering and second harmonic generation on the same platform, we simultaneously imaged mammary adipocytes, blood capillaries, collagen fibrils, and tumor cells without any labeling. We observed that obesity increases the size of lipid droplets of adipocytes in mammary gland and collagen content in mammary tumor stroma, respectively. Such impacts of obesity on mammary gland and tumor stroma could not be analyzed using standard two-dimensional histologic evaluation. Given the importance of mammary stroma to the growth and migration of tumor cells, our observation provides the first imaging evidence that supports the relationship between obesity and breast cancer risk.     

Foliar application of urea to “Sauvignon Blanc” and “Merlot” vines: doses and time of application

A careful control of the N nutritional status of grapevines can have a determining effect on wine characteristics; therefore a suitable management of N fertilization might allow some wine parameters to be modified, thereby improving product quality. The aim of this study was to determine the effect of foliar application of urea at different doses and different times of the growing season on the parameters of Sauvignon Blanc and Merlot grape juice. The research described herein involved Sauvignon Blanc and Merlot grapevines (V. vinifera L.) at a commercial vineyard and was conducted over 2 years. In the first year, N treatment involved a foliar application at a dose of 10 kg N ha^?1 during veraison, whereas in the second year it involved a foliar urea application at two doses (10 and 50 kg N ha^?1) and at three different times—3 weeks before veraison, during veraison and 3 weeks after veraison. In this second year, the urea applied at a dose of 10 kg N ha^?1 was isotopically labelled with 1% ¹⁵N. Chemical parameters, yeast assimilable N, amino acid content, amino acid profile and N isotopic composition were determined for all treatments. Grape and grape-juice parameters for Merlot were found to be more affected by N fertilization than for Sauvignon Blanc and were also more affected during the second year than during the first year, thus indicating that the climatic characteristics of each campaign could affect these parameters. The yeast assimilable N in grape juice was found to be higher for late applications of foliar urea, with application of the higher dose of urea during veraison increasing the amino acid and proline contents in both varieties. The isotopic analysis data showed that the urea applied to leaves was transferred to the berries, with the maximum translocation in Sauvignon Blanc occurring for the post-veraison treatment and in Merlot for the veraison treatment. We can therefore conclude that foliar application of urea could modify grape juice quality and could therefore be used as a tool for obtaining quality wines.     

An Adaptive Complex Network Model for Brain Functional Networks

Brain functional networks are graph representations of activity in the brain, where the vertices represent anatomical regions and the edges their functional connectivity. These networks present a robust small world topological structure, characterized by highly integrated modules connected sparsely by long range links. Recent studies showed that other topological properties such as the degree distribution and the presence (or absence) of a hierarchical structure are not robust, and show different intriguing behaviors. In order to understand the basic ingredients necessary for the emergence of these complex network structures we present an adaptive complex network model for human brain functional networks. The microscopic units of the model are dynamical nodes that represent active regions of the brain, whose interaction gives rise to complex network structures. The links between the nodes are chosen following an adaptive algorithm that establishes connections between dynamical elements with similar internal states. We show that the model is able to describe topological characteristics of human brain networks obtained from functional magnetic resonance imaging studies. In particular, when the dynamical rules of the model allow for integrated processing over the entire network scale-free non-hierarchical networks with well defined communities emerge. On the other hand, when the dynamical rules restrict the information to a local neighborhood, communities cluster together into larger ones, giving rise to a hierarchical structure, with a truncated power law degree distribution.     

Endogenous circadian rhythms in pigment composition induce changes in photochemical efficiency in plant canopies

There is increasing evidence that the circadian clock is a significant driver of photosynthesis that becomes apparent when environmental cues are experimentally held constant. We studied whether the composition of photosynthetic pigments is under circadian regulation, and whether pigment oscillations lead to rhythmic changes in photochemical efficiency. To address these questions, we maintained canopies of bean and cotton, after an entrainment phase, under constant (light or darkness) conditions for 30–48 h. Photosynthesis and quantum yield peaked at subjective noon, and non‐photochemical quenching peaked at night. These oscillations were not associated with parallel changes in carbohydrate content or xanthophyll cycle activity. We observed robust oscillations of Chl a/b during constant light in both species, and also under constant darkness in bean, peaking when it would have been night during the entrainment (subjective nights). These oscillations could be attributed to the synthesis and/or degradation of trimeric light‐harvesting complex II (reflected by the rhythmic changes in Chl a/b), with the antenna size minimal at night and maximal around subjective noon. Considering together the oscillations of pigments and photochemistry, the observed pattern of changes is counterintuitive if we assume that the plant strategy is to avoid photodamage, but consistent with a strategy where non‐stressed plants maximize photosynthesis.     

Goodpasture antigen-binding protein is a soluble exportable protein that interacts with type IV collagen. Identification of novel membrane-bound isoforms

Goodpasture-antigen binding protein (GPBP) is a nonconventional Ser/Thr kinase for basement membrane type IV collagen. Various studies have questioned these findings and proposed that GPBP serves as transporter of ceramide between the endoplasmic reticulum and the Golgi apparatus. Here we show that cells expressed at least two GPBP isoforms resulting from canonical (77-kDa) and noncanonical (91-kDa) mRNA translation initiation. The 77-kDa polypeptide interacted with type IV collagen and localized as a soluble form in the extracellular compartment. The 91-kDa polypeptide and its derived 120-kDa polypeptide associated with cellular membranes and regulated the extracellular levels of the 77-kDa polypeptide. A short motif containing two phenylalanines in an acidic tract and the 26-residue Ser-rich region were required for efficient 77-kDa polypeptide secretion. Removal of the 26-residue Ser-rich region by alternative exon splicing rendered the protein cytosolic and sensitive to the reduction of sphingomyelin cellular levels. These and previous data implicate GPBPs in a multicompartmental program for protein secretion (i.e. type IV collagen) that includes: 1) phosphorylation and regulation of protein molecular/supramolecular organization and 2) interorganelle ceramide trafficking and regulation of protein cargo transport to the plasma membrane.     

N-Benzoyl amino acids as ICAM/LFA-1 inhibitors. Part 2: structure-activity relationship of the benzoyl moiety

o-Bromobenzoyl l-tryptophan 1 inhibits the association of LFA-1 with ICAM-1 with an IC(50) of 1.7microM. Evaluation of the structure-activity relationship of the benzoyl moiety shows that 2,6-di-substitutions greatly enhance potency of this class of inhibitors. Electronegative substitutions that favor a 90 degrees angle between the benzoyl ring and the amide bond yield the most potent compounds. There is a strong correlation between the potency of the compounds and the difference between the ab initio energy at 90 degrees and the global minima energy for given compounds. Combining the favored benzoyl substitutions with l-histidine and l-asparagine resulted in a 15-fold increase in potency over compound 1.     

Efficient Transformation of <Emphasis Type="Italic">Cellulomonas flavigena</Emphasis> by Electroporation and Conjugation with <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis>

The conjugative self-transmissible plasmid pHT73, harbored in Bacillus thuringiensis var. kurstaki, was demonstrated to be transferred to Cellulomonas flavigena, a cellulolytic bacterium. Both conjugation and transformation procedures yielded resistant colonies; however, chromosomal integration was observed only when bacterial conjugation occurred. The efficiency of conjugation was 10% of recipient strain, which is considered a very efficient process. When the plasmid pHT73 was introduced by transformation, erythromycin-resistant cells contained the plasmid as an episome with no arrangements, as assayed by Southern blot analysis. In contrast, conjugated-resistant cells harbor the plasmid integrated into the chromosome. These data suggest a common mechanism of cell communication between nonrelated bacterial species with similar ecological habitats, and also that both electroporation and conjugation can be used to transform C. flavigena efficiently.     

Autoinhibitory regulation of TrwK, an essential VirB4 ATPase in type IV secretion systems

Type IV secretion systems (T4SS) mediate the transfer of DNA and protein substrates to target cells. TrwK, encoded by the conjugative plasmid R388, is a member of the VirB4 family, comprising the largest and most conserved proteins of T4SS. In a previous work we demonstrated that TrwK is able to hydrolyze ATP. Here, based on the structural homology of VirB4 proteins with the DNA-pumping ATPase TrwB coupling protein, we generated a series of variants of TrwK where fragments of the C-terminal domain were sequentially truncated. Surprisingly, the in vitro ATPase activity of these TrwK variants was much higher than that of the wild-type enzyme. Moreover, addition of a synthetic peptide containing the amino acid residues comprising this C-terminal region resulted in the specific inhibition of the TrwK variants lacking such domain. These results indicate that the C-terminal end of TrwK plays an important regulatory role in the functioning of the T4SS.