Hes1 Increases the Invasion Ability of Colorectal Cancer Cells via the STAT3-MMP14 Pathway

The Notch pathway contributes to self-renewal of tumor-initiating cell and inhibition of normal colonic epithelial cell differentiation. Deregulated expression of Notch1 and Jagged1 is observed in colorectal cancer. Hairy/enhancer of split (HES) family, the most characterized targets of Notch, involved in the development of many cancers. In this study, we explored the role of Hes1 in the tumorigenesis of colorectal cancer. Knocking down Hes1 induced CRC cell senescence and decreased the invasion ability, whereas over-expression of Hes1 increased STAT3 phosphorylation activity and up-regulated MMP14 protein level. We further explored the expression of Hes1 in human colorectal cancer and found high Hes1 mRNA expression is associated with poor prognosis in CRC patients. These findings suggest that Hes1 regulates the invasion ability through the STAT3-MMP14 pathway in CRC cells and high Hes1 expression is a predictor of poor prognosis of CRC.     

Higher-energy collision-activated dissociation without a dedicated collision cell

Beam-type collisional activation dissociation (HCD) offers many advantages over resonant excitation collision-activated dissociation, including improved identification of phosphorylated peptides and compatibility with isobaric tag-based quantitation (e.g. tandem mass tag (TMT) and iTRAQ). However, HCD typically requires specially designed and dedicated collision cells. Here we demonstrate that HCD can be performed in the ion injection pathway of a mass spectrometer with a standard atmospheric inlet (iHCD). Testing this method on complex peptide mixtures revealed similar identification rates to collision-activated dissociation (2883 versus 2730 IDs for iHCD/CAD, respectively) and precursor-product-conversion efficiency comparable to that achieved within a dedicated collision cell. Compared with pulsed-q dissociation, a quadrupole ion trap-based method that retains low-mass isobaric tag reporter ions, iHCD yielded isobaric tag for relative and absolute quantification reporter ions 10-fold more intense. This method involves no additional hardware and can theoretically be implemented on any mass spectrometer with an atmospheric inlet.     

Grouper as a natural biocontrol of invasive lionfish

Lionfish (Pterois volitans/miles) have invaded the majority of the Caribbean region within five years. As voracious predators of native fishes with a broad habitat distribution, lionfish are poised to cause an unprecedented disruption to coral reef diversity and function. Controls of lionfish densities within its native range are poorly understood, but they have been recorded in the stomachs of large-bodied Caribbean groupers. Whether grouper predation of lionfish is sufficient to act as a biocontrol of the invasive species is unknown, but pest biocontrol by predatory fishes has been reported in other ecosystems. Groupers were surveyed along a chain of Bahamian reefs, including one of the region's most successful marine reserves which supports the top one percentile of Caribbean grouper biomass. Lionfish biomass exhibited a 7-fold and non-linear reduction in relation to the biomass of grouper. While Caribbean grouper appear to be a biocontrol of invasive lionfish, the overexploitation of their populations by fishers, means that their median biomass on Caribbean reefs is an order of magnitude less than in our study. Thus, chronic overfishing will probably prevent natural biocontrol of lionfishes in the Caribbean.     

Connectivity, sustainability, and yield: bridging the gap between conventional fisheries management and marine protected areas

A substantial shift toward use of marine protected areas (MPAs) for conservation and fisheries management is currently underway. This shift to explicit spatial management presents new challenges and uncertainties for ecologists and resource managers. In particular, the potential for MPAs to change population sustainability, fishery yield, and ecosystem properties depends on the poorly understood consequences of three critical forms of connectivity over space: larval dispersal, juvenile and adult swimming, and movement of fishermen. Conventional fishery management describes the dynamics and current status of fish populations, with increasing recent emphasis on sustainability, often through reference points that reflect individual replacement. These compare lifetime egg production (LEP) to a critical replacement threshold (CRT) whose value is uncertain. Sustainability of spatially distributed populations also depends on individual replacement, but through all possible paths created by larval dispersal and LEP at each location. Model calculations of spatial replacement considering larval connectivity alone indicate sustainability and yield depend on species dispersal distance and the distribution of LEP created by species habitat distribution and fishing mortality. Adding MPAs creates areas with high LEP, increasing sustainability, but not necessarily yield. Generally, short distance dispersers will persist in almost all MPAs, while sustainability of long distance dispersers requires a specific density of MPAs along the coast. The value of that density also depends on the uncertain CRT, as well as fishing rate. MPAs can increase yield in areas with previously low LEP but for short distance dispersers, high yields will require many small MPAs. The paucity of information on larval dispersal distances, especially in cases with strong advection, renders these projections uncertain. Adding juvenile and adult movement to these calculations reduces LEP near the edges in MPAs, if movement is within a home-range, but more broadly over space if movement is diffusive. Adding movement of fishermen shifts effort on the basis of anticipated revenues and fishing costs, leading to lower LEP near ports, for example. Our evolving understanding of connectivity in spatial management could form the basis for a new, spatially oriented replacement reference point for sustainability, with associated new uncertainties.     

Nudt21 Controls Cell Fate by Connecting Alternative Polyadenylation to Chromatin Signaling

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C pigment locus mutants of the fowl produce enzymatically inactive tyrosinase-like molecules

Three albino mutants of the fowl were tested for tyrosinase activity. Two of these mutants (c and ca) are alleles at the autosomal C locus, while the third mutant (sal) is sex-linked. Both the standard type, E, and sal are tyrosinase positive whereas the two C mutants are tyrosinase negative. Anti-chicken tyrosinase mouse serum was produced and all four genotypes were found to have cross-reacting material to this antiserum. Tyrosinase from the standard type was isolated and its location on denaturing two-dimensional gels determined. A co-migrating series of spots was found within the protein pattern of both the standard type and the tyrosinase positive albino, sal. The same pattern of spots was also observed for c and ca with no apparent change in either the pI or the molecular weight. Transmembrane blots also showed spots that reacted with anti-tyrosinase serum in all four genotypes and that migrated to the same location as that of standard tyrosinase. It is proposed that both c and ca are CRM+ mutants which produce tyrosinase-like molecules that are inactive due to a change that is electrophoretically and antigenically "silent".     

Studies on the binding site of the galactose-specific agglutinin PA-IL from Pseudomonas aeruginosa

The binding properties of Pseudomonas aeruginosa agglutinin-I (PA-IL) with glycoproteins (gps) and polysaccharides were studied by both the biotin/avidin-mediated microtiter plate lectin-binding assay and the inhibition of agglutinin-glycan interaction with sugar ligands. Among 36 glycans tested for binding, PA-IL reacted best with two glycoproteins containing Galalpha1-->4Gal determinants and a human blood group ABO precursor equivalent gp, but this lectin reacted weakly or not at all with A and H active gps or sialylated gps. Among the mammalian disaccharides tested by the inhibition assay, the human blood group Pkactive Galalpha1-->4Gal, was the best. It was 7.4-fold less active than melibiose (Galalpha1-->6Glc). PA-IL has a preference for the alpha-anomer in decreasing order as follows: Galalpha1-->6 >Galalpha1-->4 >Galalpha1-->3. Of the monosaccharides studied, the phenylbeta derivatives of Gal were much better inhibitors than the methylbeta derivative, while only an insignificant difference was found between the Galalpha anomer of methyl- and p -NO2-phenyl derivatives. From these results, it can be concluded that the combining size of the agglutinin is as large as a disaccharide of the alpha-anomer of Gal at nonreducing end and most complementary to Galalpha1-->6Glc. As for the combining site of PA-IL toward the beta-anomer, the size is assumed to be less than that of Gal; carbon-6 in the pyranose form is essential, and hydrophobic interaction is important for binding.      

Release of spectrin-free vesicles from human erythrocytes during ATP depletion: 1. characterization of spectrin-free vesicles

Human erythrocytes incubated without glucose at 37 degrees C (in vitro aging) release spectrin-free vesicles after 12 or more hours. The release of vesicles is dependent upon ATP depletion. If the endogenous level of ATP is maintained, vesicle release is completely inhibited up to 54 h. Vesicle release is independent of hemolysis because in vitro aged cells and cells that maintain their ATP levels lose identical amounts of hemoglobin up to 45 h. 93 percent of all membrane particles released constitute a uniform population of spheres with a diameter of 185 +/- 23nm. These vesicles are of slightly varying densities due to varying contents of hemoglobin. Vesicles contain half the amount of membrane protein that is found in intact membranes when referred to the content of phospholipids phosphorus. This is primarily due to the absence of spectrin. However, their content of protein component III, glycophorin, and cholesterol remains the same as in intact membranes. Thus, the major integral membrane proteins are present in vesicles in similar quantities were surface area as in cells except for the enzyme acetylcholinesterase that is enriched up to twofold. The phospholipids composition of these vesicles is representative of the intact membrane except that the amount of phosphatidic acid is 10-fold higher and the amount of phosphatidylethanolamine is slightly lower than in erythrocytes. These results suggest a selective release of membrane domains that lack peripheral membrane proteins and are enriched in acetylcholinesterase. This release of spectrin-free vesicles from cells aged in vitro could represent an acceleration of the physiological aging process.     

Specific binding of 1alpha,25-dihydroxycholecalciferol to nuclear components of chick intestine.

Specific binding of 1alpha,25-dihydroxycholecalciferol to macromolecular components of small intestinal mucosa nuclei is demonstrated in vitamin D-deficient chicks. The nuclear 1alpha,25-dihydroxycholecalciferol-macromolecule complex was isolated on sucrose density gradients and sediments at 3.7 S in the presence of 0.3 M KCl. Agarose gel filtration of the nuclear component indicated an apparent molecular weight of 47,000. The nuclear receptor complexes could not be distinguished from previously described cytoplasmic 1alpha,25-dihydroxycholecalciferol-binding components by the ultracentrifugation and chromatographic procedures employed. The association of the 3-H-sterol with the nuclear component is thermolabile and is destroyed by treatment with pronase, but not by nucleases; the receptor component is therefore presumed to be a protein. The macromolecular-1alpha,25-dihydroxycholecalciferol complex formed in vivo or in vitro at 25 degrees can be extracted from intestinal nuclei by 0.3 M KCl, but not by low salt buffers. Smaller amounts of the 3.7 S binding component can be detected in isolated purified chromatin or after incubation of 1alpha,25-dihydroxy[3-H]cholecalciferol with reconstituted cytosol-chromatin at 0 degrees. Following incubation of the labeled hormone with reconstituted cytosol-chromatin at 0 degrees, 1alpha,25-dihydroxy[3-H]cholecalciferol is primarily associated with the cytoplasmic receptor, After shifting the incubation temperature to 25 degrees, a progressive increase in the concentration of the nuclear receptor complex and a concomitant decrease in the concentration of the cytoplasmic binding component occur. Thus the 1alpha,25-dihydroxycholecalciferol binding molecules appear to exist primarily in the cytoplasm, where they presumably function to transport the hormone into the nucleus. Experiments employing incubation of 1alpha,25-dihydroxy[3-H]cholecalciferol with reconstituted cytosol-chromatin from nontarget tissues indicate a requirement for both intestinal cytosol and chromatin for maximal formation of the nuclear hormone-receptor complex. These results suggest that the nuclear-binding component arises from hormone-dependent transfer of the cytoplasmic 1alpha,25-dihydroxycholecalciferol receptor to intestinal chromatin acceptor sites.