Type I collagen abrogates the clathrin-mediated internalization of membrane type 1 matrix metalloproteinase (MT1-MMP) via the MT1-MMP hemopexin domain

Type I collagen (Col I)-stimulated matrix metalloproteinase-2 (MMP-2) activation via membrane type 1 MMP (MT1-MMP) involves both a transcriptional increase in MT1-MMP expression and a nontranscriptional response mediated by preexisting MT1-MMP. In order to identify which MT1-MMP domains were required for the nontranscriptional response, MCF-7 cells that lack endogenous MT1-MMP were transfected with either wild type or domain mutant MT1-MMP constructs. We observed that mutant constructs lacking the MT1-MMP cytoplasmic tail were able to activate MMP-2 in response to Col I but not a construct lacking the MT1-MMP hemopexin domain. Col I did not alter total MT1-MMP protein levels; nor did it appear to directly induce MT1-MMP oligomerization. Col I did, however, redistribute preexisting MT1-MMP to the cell periphery compared with unstimulated cells that displayed amore diffuse staining pattern. In addition, Col I blocked the internalization of MT1-MMP in a dynamin-dependent manner via clathrin-coated pit-mediated endocytosis. This mechanism of impaired internalization is different from that reported for concanavalin A, since it is not mediated by the cytoplasmic tail of MT1-MMP but rather by the hemopexin domain. In summary, upon Col I binding to its cell surface receptor, MT1-MMP internalization via clathrin-coated pit-mediated endocytosis is impaired through interactions with the hemopexin domain, thereby regulating its function and ability to activate MMP-2.

Transmembrane/cytoplasmic domain-mediated membrane type 1-matrix metalloprotease docking to invadopodia is required for cellinvasion

The invasion of human malignant melanoma cells into the extracellular matrix (ECM) involves the accumulation of proteases at sites of ECM degradation where activation of matrix metalloproteases (MMP) occurs. Here, we show that when membrane type 1 MMP (MT-MMP) was overexpressed in RPMI7951 human melanoma cells, the cells made contact with the ECM, activated soluble and ECM-bound MMP-2, and degraded and invaded the ECM. Further experiments demonstrated the importance of localization of the MT-MMP to invadopodia. Overexpression of MT-MMP without invadopodial localization caused activation of soluble MMP-2, but did not facilitate ECM degradation or cell invasiveness. Up-regulation of endogenous MT-MMP with concanavalin A caused activation of MMP-2. However, concanavalin A treatment prevented invadopodial localization of MT-MMP and ECM degradation. Neither a truncated MT-MMP mutant lacking transmembrane (TM) and cytoplasmic domains (ΔTM(MT-MMP)), nor a chimeric MT-MMP containing the interleukin 2 receptor α chain (IL-2R) TM and cytoplasmic domains (ΔTM(MT-MMP)/TM(IL-2R)) were localized to invadopodia or exhibited ECM degradation. Furthermore, a chimera of the TM/cytoplasmic domain of MT-MMP (TM(MT-MMP)) with tissue inhibitor of MMP 1 (TIMP-1/TM(MT- MMP)) directed the TIMP-1 molecule to invadopodia. Thus, the MT-MMP TM/cytoplasmic domain mediates the spatial organization of MT-MMP into invadopodia and subsequent degradation of the ECM.

ST7-mediated suppression of tumorigenicity of prostate cancer cells is characterized by remodeling of the extracellular matrix

Multiple lines of evidence have provided compelling evidence for the existence of a tumor suppressor gene (TSG) on chromosome 7q31.1. ST7 may be the target of this genetic instability but its designation as a TSG is controversial. In this study, we show that, functionally, ST7 behaves as a tumor suppressor in human cancer. ST7 suppressed growth of PC-3 prostate cancer cells inoculated subcutaneously into severe combined immunodeficient mice, and increased the latency of tumor detection from 13 days in control tumors to 23 days. Re-expression of ST7 was also associated with suppression of colony formation under anchorage-independent conditions in MDA-MB-231 breast cancer cells and ST7 mRNA expression was downregulated in 44% of primary breast cancers. Expression profiling of PC-3 cells revealed that ST7 predominantly induces changes in genes involved in re-modeling the extracellular matrix such as SPARC, IGFBP5 and several matrix metalloproteinases. These data indicate that ST7 may mediate tumor suppression through modification of the tumor microenvironment.

The type I collagen induction of MT1-MMP-mediated MMP-2 activation is repressed by αVβ3 integrin in human breast cancer cells

The influence of αVβ3 integrin on MT1-MMP functionality was studied in human breast cancer cells of differing β3 integrin status. Overexpression of β3 integrin caused increased cell surface expression of αV integrin and increased cellular adhesion to extracellular matrix (ECM) substrates in BT-549, MDA-MB-231 and MCF-7 cells. β3 integrin expression also enhanced the migration of breast cancer cells on ECM substrates and enhanced collagen gel contraction. In vivo, αVβ3 cooperated with MT1-MMP to increase the growth of MCF-7 cells after orthotopic inoculation in immunocompromised mice, but had no influence on in vitro proliferation. Despite these stimulatory effects, overexpression of β3 integrin suppressed the type I collagen (Col I) induced MMP-2 activation in all breast cancer cell lines analyzed. This was also evident in extracts from the MCF-7 tumors in vivo, where MMP-2 activation was stimulated by MT1-MMP transfection, but attenuated with β3 integrin expression. Although our studies confirm important biological effects of αVβ3 integrin on enhancing cell adhesion and migration, ECM remodeling and tumor growth, β3 integrin caused reduced MMP-2 activation in response to Col I in vitro, which appears to be physiologically relevant, as it was also seen in tumor xenografts in vivo. The reduction of MMP-2 activation (and thus MT1-MMP activity) by αVβ3 in response to Col I may be important in scenarios where cells which are activated for matrix degradation need to preserve some pericellular collagen, perhaps as a substrate for cell adhesion and migration, thus maintaining a balanced level of proteolysis required for efficient tumor growth.

Self assembly of plasmonic core-satellite nano-assemblies mediated by hyperbranched polymer linkers

The morphology of plasmonic nano-assemblies has a direct influence on optical properties, such as localised surface plasmon resonance (LSPR) and surface enhanced Raman scattering (SERS) intensity. Assemblies with core-satellite morphologies are of particular interest, because this morphology has a high density of hot-spots, while constraining the overall size. Herein, a simple method is reported for the self-assembly of gold NPs nano-assemblies with a core-satellite morphology, which was mediated by hyperbranched polymer (HBP) linkers. The HBP linkers have repeat units that do not interact strongly with gold NPs, but have multiple end-groups that specifically interact with the gold NPs and act as anchoring points resulting in nano-assemblies with a large (~48 nm) core surrounded by smaller (~15 nm) satellites. It was possible to control the number of satellites in an assembly which allowed optical parameters such as SPR maxima and the SERS intensity to be tuned. These results were found to be consistent with finite-difference time domain (FDTD) simulations. Furthermore, the multiplexing of the nano-assemblies with a series of Raman tag molecules was demonstrated, without an observable signal arising from the HBP linker after tagging. Such plasmonic nano-assemblies could potentially serve as efficient SERS based diagnostics or biomedical imaging agents in nanomedicine.

Effect of Coulomb blockade, gold resistance, and thermal expansion on the electrical resistance of ultrathin gold films

Controlling the electrical resistance of granular thin films is of great importance for many applications, yet a full understanding of electron transport in such films remains a major challenge. We have studied experimentally and by model calculations the temperature dependence of the electrical resistance of ultrathin gold films at temperatures between 2 K and 300 K. Using sputter deposition, the film morphology was varied from a discontinuous film of weakly coupled meandering islands to a continuous film of strongly coupled coalesced islands. In the weak-coupling regime, we compare the regular island array model, the cotunneling model, and the conduction percolation model with our experimental data. We show that the tunnel barriers and the Coulomb blockade energies are important at low temperatures and that the thermal expansion of the substrate and the island resistance affect the resistance at high temperatures. At low temperatures our experimental data show evidence for a transition from electron cotunneling to sequential tunneling but the data can also be interpreted in terms of conduction percolation. The resistivity and temperature coefficient of resistance of the meandering gold islands are found to resemble those of gold nanowires. We derive a simple expression for the temperature at which the resistance changes from non-metal-like behavior into metal-like behavior. In the case of strong island coupling, the total resistance is solely determined by the Ohmic island resistance.

Structure-mediated thermal transport of monolayer graphene allotropes nanoribbons

We report the study of the thermal transport management of monolayer graphene allotrope nanoribbons (size ∼20 × 4 nm2) by the modulation of their structures via molecular dynamics simulations. The thermal conductivity of graphyne (GY)-like geometries is observed to decrease monotonously with increasing number of acetylenic linkages between adjacent hexagons. Strikingly, by incorporating those GY or GY-like structures, the thermal performance of graphene can be effectively engineered. The resulting hetero-junctions possess a sharp local temperature jump at the interface, and show a much lower effective thermal conductivity due to the enhanced phonon–phonon scattering. More importantly, by controlling the percentage, type and distribution pattern of the GY or GY-like structures, the hetero-junctions are found to exhibit tunable thermal transport properties (including the effective thermal conductivity, interfacial thermal resistance and rectification). This study provides a heuristic guideline to manipulate the thermal properties of 2D carbon networks, ideal for application in thermoelectric devices with strongly suppressed thermal conductivity.

Signet-ring cell carcinoma of colorectum : current perspectives and molecular biology

PURPOSE Colorectal signet-ring cell carcinoma (SRCC) is rare, and very little detailed information on the molecular biology of the disease is available. METHODS The literature on the clinical, pathological and, in particular, the molecular biology of this rare entity was critically reviewed. The reviewed articles take into account a total of 1,817 cases of SRCC, but only 143 cases have molecular data available. The characteristics of two patients with colorectal SRCC were also discussed. RESULTS Colorectal SRCC mostly occurs in younger patients, is larger and has different site predilection compared with conventional colorectal adenocarcinoma. It can occur as one of the synchronous cancers in the colorectum. The cancer is usually diagnosed at advanced stages because of the late manifestation of symptoms, and aggressive treatment strategy is required. Limited reports in the literature have shown that the variant of colorectal cancer demonstrated a different pattern of genetic alterations of common growth kinase-related oncogenes (K-ras, BRAF), tumour suppressor genes (p53, p16), gene methylation and cell adhesion-related genes related to the Wingless signalling pathway (E-cadherin and beta-catenin) from conventional colorectal adenocarcinoma. Colorectal SRCC also showed high expression of mucin-related genes and genes related to the gastrointestinal system. There was also a higher prevalence of microsatellite instability-high tumours and low Cox-2 expression in colorectal SRCC as opposed to conventional adenocarcinoma. CONCLUSIONS Colorectal SRCC has unique molecular pathological features. The unique molecular profiles in SRCC may provide molecular-based improvements to patient management in colorectal SRCC.

Mechanisms of mono- and poly-ubiquitination : ubiquitination specificity depends on compatibility between the E2 catalytic core and amino acid residues proximal to the lysine

Ubiquitination involves the attachment of ubiquitin to lysine residues on substrate proteins or itself, which can result in protein monoubiquitination or polyubiquitination. Ubiquitin attachment to different lysine residues can generate diverse substrate-ubiquitin structures, targeting proteins to different fates. The mechanisms of lysine selection are not well understood. Ubiquitination by the largest group of E3 ligases, the RING-family E3 s, is catalyzed through co-operation between the non-catalytic ubiquitin-ligase (E3) and the ubiquitin-conjugating enzyme (E2), where the RING E3 binds the substrate and the E2 catalyzes ubiquitin transfer. Previous studies suggest that ubiquitination sites are selected by E3-mediated positioning of the lysine toward the E2 active site. Ultimately, at a catalytic level, ubiquitination of lysine residues within the substrate or ubiquitin occurs by nucleophilic attack of the lysine residue on the thioester bond linking the E2 catalytic cysteine to ubiquitin. One of the best studied RING E3/ E2 complexes is the Skp1/Cul1/F box protein complex, SCFCdc4, and its cognate E2, Cdc34, which target the CDK inhibitor Sic1 for K48-linked polyubiquitination, leading to its proteasomal degradation. Our recent studies of this model system demonstrated that residues surrounding Sic1 lysines or lysine 48 in ubiquitin are critical for ubiquitination. This sequence-dependence is linked to evolutionarily conserved key residues in the catalytic region of Cdc34 and can determine if Sic1 is mono- or poly-ubiquitinated. Our studies indicate that amino acid determinants in the Cdc34 catalytic region and their compatibility to those surrounding acceptor lysine residues play important roles in lysine selection. This may represent a general mechanism in directing the mode of ubiquitination in E2 s.

Molecular basis for lysine specificity in the yeast ubiquitin-conjugating enzyme Cdc34

Ubiquitin (Ub)-conjugating enzymes (E2s) and ubiquitin ligases (E3s) catalyze the attachment of Ub to lysine residues in substrates and Ub during monoubiquitination and polyubiquitination. Lysine selection is important for the generation of diverse substrate-Ub structures, which provides versatility to this pathway in the targeting of proteins to different fates. The mechanisms of lysine selection remain poorly understood, with previous studies suggesting that the ubiquitination site(s) is selected by the E2/E3-mediated positioning of a lysine(s) toward the E2/E3 active site. By studying the polyubiquitination of Sic1 by the E2 protein Cdc34 and the RING E3 Skp1/Cul1/F-box (SCF) protein, we now demonstrate that in addition to E2/E3-mediated positioning, proximal amino acids surrounding the lysine residues in Sic1 and Ub are critical for ubiquitination. This mechanism is linked to key residues composing the catalytic core of Cdc34 and independent of SCF. Changes to these core residues altered the lysine preference of Cdc34 and specified whether this enzyme monoubiquitinated or polyubiquitinated Sic1. These new findings indicate that compatibility between amino acids surrounding acceptor lysine residues and key amino acids in the catalytic core of ubiquitin-conjugating enzymes is an important mechanism for lysine selection during ubiquitination.

Prediction of thermal expansion properties of carbon nanotubes using molecular dynamics simulations

The axial coefficients of thermal expansion (CTE) of various carbon nanotubes (CNTs), i.e., single-wall carbon nanotubes (SWCNTs), and some multi-wall carbon nanotubes (MWCNTs), were predicted using molecular dynamics (MDs) simulations. The effects of two parameters, i.e., temperature and the CNT diameter, on CTE were investigated extensively. For all SWCNTs and MWCNTs, the obtained results clearly revealed that within a wide low temperature range, their axial CTEs are negative. As the diameter of CNTs decreases, this temperature range for negative axial CTEs becomes narrow, and positive axial CTEs appear in high temperature range. It was found that the axial CTEs vary nonlinearly with the temperature, however, they decrease linearly as the CNT diameter increases. Moreover, within a wide temperature range, a set of empirical formulations was proposed for evaluating the axial CTEs of armchair and zigzag SWCNTs using the above two parameters. Finally, it was found that the absolute value of the negative axial CTE of any MWCNT is much smaller than those of its constituent SWCNTs, and the average value of the CTEs of its constituent SWCNTs. The present fundamental study is very important for understanding the thermal behaviors of CNTs in such as nanocomposite temperature sensors, or nanoelectronics devices using CNTs.

A case study of the Iranian national railway and its absolute capacity expansion using analytical models

Identifying railway capacity is an important task that can identify "in principal" whether the network can handle an intended traffic flow, and whether there is any free capacity left for additional train services. Capacity determination techniques can also be used to identify how best to improve an existing network, and at least cost. In this article an optimization approach has been applied to a case study of the Iran national railway, in order to identify its current capacity and to optimally expand it given a variety of technical conditions. This railway is very important in Iran and will be upgraded extensively in the coming years. Hence the conclusions in this article may help in that endeavor. A sensitivity analysis is recommended to evaluate a wider range of possible scenarios. Hence more useful lower and upper bounds can be provided for the performance of the system

Integrin-linked kinase as a target for ERG-mediated invasive properties in prostate cancer models

Approximately half of prostate cancers (PCa) carry TMPRSS2-ERG translocations; however, the clinical impact of this genomic alteration remains enigmatic. Expression of v-ets erythroblastosis virus E26 oncogene like (avian) gene (ERG) promotes prostatic epithelial dysplasia in transgenic mice and acquisition of epithelial-to-mesenchymal transition (EMT) characteristics in human prostatic epithelial cells (PrECs). To explore whether ERG-induced EMT in PrECs was associated with therapeutically targetable transformation characteristics, we established stable populations of BPH-1, PNT1B and RWPE-1 immortalized human PrEC lines that constitutively express flag-tagged ERG3 (fERG). All fERG-expressing populations exhibited characteristics of in vitro and in vivo transformation. Microarray analysis revealed >2000 commonly dysregulated genes in the fERG-PrEC lines. Functional analysis revealed evidence that fERG cells underwent EMT and acquired invasive characteristics. The fERG-induced EMT transcript signature was exemplified by suppressed expression of E-cadherin and keratins 5, 8, 14 and 18; elevated expression of N-cadherin, N-cadherin 2 and vimentin, and of the EMT transcriptional regulators Snail, Zeb1 and Zeb2, and lymphoid enhancer-binding factor-1 (LEF-1). In BPH-1 and RWPE-1-fERG cells, fERG expression is correlated with increased expression of integrin-linked kinase (ILK) and its downstream effectors Snail and LEF-1. Interfering RNA suppression of ERG decreased expression of ILK, Snail and LEF-1, whereas small interfering RNA suppression of ILK did not alter fERG expression. Interfering RNA suppression of ERG or ILK impaired fERG-PrEC Matrigel invasion. Treating fERG-BPH-1 cells with the small molecule ILK inhibitor, QLT-0267, resulted in dose-dependent suppression of Snail and LEF-1 expression, Matrigel invasion and reversion of anchorage-independent growth. These results suggest that ILK is a therapeutically targetable mediator of ERG-induced EMT and transformation in PCa.

Web mediated market interactions and enterprise software

With increased consolidation and a few large vendors dominating the market, how can software vendors distinguish themselves in order to maintain profitability and gain market share? Increasingly customers are becoming more proactive in selecting a vendor and a product, drawing upon various publications, market surveys, mailing lists, and, of course, other users. In particular, though, a company's Web site is the obvious place to begin information gathering. In sum, it may seem that the days of the uninformed customer prepared to be "sold to" are potentially all but gone.