Retinoic acid inhibits beta-catenin through suppression of Cox-2: a role for truncated adenomatous polyposis coli

Mutations in adenomatous polyposis coli (APC) underlie the earliest stages of colorectal carcinogenesis. Consequences of APC mutation include stabilization of beta-catenin, dysregulation of cyclooxygenase-2 (COX-2) expression, and loss of retinoic acid production, events with poorly defined interactions. Here we showed that treatment of zebrafish expressing a truncated form of Apc with either retinoic acid or a selective COX-2 inhibitor decreased beta-catenin protein levels and downstream signaling events. Interestingly, the destruction of beta-catenin in apc mutant embryos following Cox-2 inhibition required the presence of truncated Apc. These findings support roles for retinoic acid and Cox-2 in regulating the stability of beta-catenin following Apc loss. Furthermore, truncated Apc appears to retain the ability to target beta-catenin for destruction, but only in the absence of Cox-2 activity. This novel function of truncated Apc may provide a molecular basis for the efficacy of COX-2 inhibitors in the treatment of colon cancer.     

NO and CO differentially activate soluble guanylyl cyclase via a heme pivot-bend mechanism

Diatomic ligand discrimination by soluble guanylyl cyclase (sGC) is paramount to cardiovascular homeostasis and neuronal signaling. Nitric oxide (NO) stimulates sGC activity 200-fold compared with only four-fold by carbon monoxide (CO). The molecular details of ligand discrimination and differential response to NO and CO are not well understood. These ligands are sensed by the heme domain of sGC, which belongs to the heme nitric oxide oxygen (H-NOX) domain family, also evolutionarily conserved in prokaryotes. Here we report crystal structures of the free, NO-bound, and CO-bound H-NOX domains of a cyanobacterial homolog. These structures and complementary mutational analysis in sGC reveal a molecular ruler mechanism that allows sGC to favor NO over CO while excluding oxygen, concomitant to signaling that exploits differential heme pivoting and heme bending. The heme thereby serves as a flexing wedge, allowing the N-terminal subdomain of H-NOX to shift concurrent with the transition of the six- to five-coordinated NO-bound state upon sGC activation. This transition can be modulated by mutations at sGC residues 74 and 145 and corresponding residues in the cyanobacterial H-NOX homolog.     

Synthesis, characterization, and magnetic properties of new homotrinuclear bis(oxamato) copper(II) complexes with an asymmetric central N,N′-bridge

The new mononuclear bis(oxamato) complex [n-Bu₄N]₂[Cu(obbo)] (1) (obbo=o-benzyl-bis(oxamato)) has been synthesized as a precursor for trinuclear oxamato-bridged transition metal complexes. Starting from 1 the homotrinuclear complexes [Cu₃(obbo)(pmdta)₂(NO₃)](NO₃)·CH₂Cl₂·H₂O (2) and [Cu₃(obbo)(tmeda)₂(NO₃)₂(dmf)] (3) have been prepared, where pmdta = N,N,N′,N″,N″-pentamethyldiethylenetriamine, tmeda = N,N,N′,N′-tetramethylethylenediamine and dmf = dimethylformamide. The crystal structures of 1-3 were solved. The magnetic properties of 2 and 3 were studied by susceptibility measurements versus temperature. For the intramolecular J parameter values of −111 cm⁻¹ (2) and −363 cm⁻¹ (3) were obtained.     

Role of RNA structure and susceptibility to RNase E in regulation of a cold shock mRNA, cspA mRNA

Degradation of the cspA mRNA in vivo is very rapid at temperatures greater than 30 degrees C and is moderately dependent on RNase E. Investigations in vitro show that degradosomes prepared from normal or cold-shocked cultures cleave the cspA mRNA preferentially at a single site in vitro between two stem-loops approximately 24 residues 3' to the termination codon and approximately 31 residues from the 3' end. The site of cleavage is independent of the temperature and largely independent of the phosphorylation status of the 5' end of cspA mRNA. A 5' stem-loop, potential occlusion of the initiation and termination codons, temperature-dependent translational efficiency, and the position of the RNase E cleavage site can explain the differential stability of the cspA mRNA.     

Dual electrochemical determination of glucose and insulin using enzyme and ferrocene microcapsules

Dual electrochemical determination of glucose and insulin has been developed, based on enzymatic reaction and immunoassay with utilization of ferrocene microcapsules, respectively. Glucose was determined through electrochemical oxidation of formed product, hydrogen peroxide, by the action of glucose oxidase (GOx). The layer-by-layer (LbL) films on the ferrocene microcrystal followed by anti-insulin antibody sensitization were employed for the biolabled ferrocene microcapsules production. The antibody sensitized ferrocene microcapsules worked as a probe in the proposed system. The microcapsules provided a higher signal generating molecule to antibody (S/P) ratio of 4.52x10(6) to 12.4x10(6). Microcapsules with different antibody loads (388-1070 antibody molecules per capsule) were subjected to a solid-phase immunoassay for the detection of insulin. The microcapsule having 1030 anti-insulin antibody molecules per capsule demonstrated good performance for insulin determination. The calibration curve for insulin had a linear range of 10(-10) to 10(-7) g mL(-1) with R(2)=0.990, 3.9% R.S.D. The limit of detection for insulin was 10 pg mL(-1) of 100 microL sample (equivalent to 10(-12)g of insulin). The determination range for the glucose was 0.5 and 40 mM with R(2)=0.996 and 4.1% R.S.D.     

A barcode of life database for the Cephalopoda? Considerations and concerns

The concept of a Barcode of Life Database (BoLD) for the Class Cephalopoda (Phylum Mollusca) was introduced at the Cephalopod International Advisory Council (CIAC) symposium in Hobart, Australia, February 2006. This suggestion was met with significant interest, concern and debate. This review attempts to describe the concept of the BoLD initiative and to outline considerations and concerns specific to a cephalopod BoLD. Jan M. Strugnell and Annie R. Lindgren contributed equally to this work.     

Autoregulation of the rsc4 tandem bromodomain by gcn5 acetylation

An important issue for chromatin remodeling complexes is how their bromodomains recognize particular acetylated lysine residues in histones. The Rsc4 subunit of the yeast remodeler RSC contains an essential tandem bromodomain (TBD) that binds acetylated K14 of histone H3 (H3K14ac). We report a series of crystal structures that reveal a compact TBD that binds H3K14ac in the second bromodomain and, remarkably, binds acetylated K25 of Rsc4 itself in the first bromodomain. Endogenous Rsc4 is acetylated only at K25, and Gcn5 is identified as necessary and sufficient for Rsc4 K25 acetylation in vivo and in vitro. Rsc4 K25 acetylation inhibits binding to H3K14ac, and mutation of Rsc4 K25 results in altered growth rates. These data suggest an autoregulatory mechanism in which Gcn5 performs both the activating (H3K14ac) and inhibitory (Rsc4 K25ac) modifications, perhaps to provide temporal regulation. Additional regulatory mechanisms are indicated as H3S10 phosphorylation inhibits Rsc4 binding to H3K14ac peptides.