A conserved sequence in caveolin-1 is both necessary and sufficient for caveolin polarity and cell directional migration

Caveolin-1 (Cav-1) plays an important role in the organization of signaling molecules involved in a variety of signaling pathways, including those mediating cell motility. Here we show that amino acids K47-K57 of Cav-1 are a highly conserved sequence in Cav-1 and Cav-3 proteins, and that expression of either K47-K57 deletion Cav-1 mutant or wild-type Cav-2 that lacks this sequence exhibits a non-polarized distribution pattern. Expression of K47-K57 in Cav-2 leads to Cav-2 polarity, suggesting that expression of K47-K57 is sufficient to direct caveolin polarity. Importantly, we show that expression of this sequence is both necessary and sufficient to promote cell directional migration. Thus, our results support the conclusion that Cav-1 polarity is critical for cell directional migration.     

Ghrelin O-acyltransferase (GOAT) has a preference for n-hexanoyl-CoA over n-octanoyl-CoA as an acyl donor

Ghrelin is a peptide hormone in which serine 3 is modified by n-octanoic acid through GOAT (ghrelin O-acyltransferase). However, the enzymological properties of GOAT remain to be elucidated. We analyzed the in vitro activity of GOAT using the recombinant enzyme. Unexpectedly, although the main active form of ghrelin is modified by n-octanoic acid, GOAT had a strong preference for n-hexanoyl-CoA over n-octanoyl-CoA as an acyl donor. Moreover, a four-amino acid peptide derived from the N-terminal sequence of ghrelin can be modified by GOAT, indicating that these four amino acids constitute the core motif for substrate recognition by the enzyme.     

Synthesis,biological studies and molecular dynamics of new anticancer RGD-based peptide conjugates for targeted drug delivery

New cyclic RGD peptide-anticancer agent conjugates, with different chemical functionalities attached to the parent peptide were synthesized in order to evaluate their biological activities and to provide a comparative study of their drug release profiles. The Integrin binding c(RGDfK) penta-peptide was used for the synthesis of Camptothecin (CPT) carbamate and Chlorambucil (CLB) amide conjugates. Substitution of the amino acid Lys with Ser resulted in a modified c(RGDfS) with a new attachment site, which enabled the synthesis of an ester CLB conjugate. Functional versatility of the conjugates was reflected in the variability of their drug release profiles, while the conserved RGD sequence of a selective binding to the α_v integrin family, likely preserved their recognition by the Integrin and consequently their favorable toxicity towards targeted cancer cells. This hypothesis was supported by a computational analysis suggesting that all conjugates occupy conformational spaces similar to that of the Integrin bound bio-active parent peptide.     

Structural-Thermodynamic Relationships of Interactions in the N-Terminal ATP-Binding Domain of Hsp90

Despite its importance as a target in anti-cancer therapeutics and the numerous rational-based inhibitor design efforts aimed at it, there are only limited data available on structural-thermodynamic relationships of interactions of the N-terminal ATP-binding domain of Hsp90 (N-Hsp90). Here, we redress this by presenting an investigation of binding of nucleotides and ansamycin compounds to this domain. Interactions of nucleotides with N-Hsp90 are relatively weak (> 10 μM) and are strongly enthalpy driven over the temperature range 10-25 °C. Geldanamycin (GA) and its analogues 17-AAG [17-(allylamino)-17-demethoxy-GA] and 17-DMAG (17-N,N-dimethylaminoethylamino-17-demethoxy-GA) bind more strongly and have a dominant favourable enthalpic contribution over the temperature range investigated. We investigated the temperature dependence of the enthalpic contribution to binding. We found that while the ansamycin compounds have the commonly observed negative value, the nucleotides show a negligible or even a positive ΔC_p of binding. These data represent the first observation of a single binding site for which interactions with different ligands result in both negative and positive ΔC_p values. By addressing the likely impact of the potential contributions from protein-ligand interactions, we are able to attribute the anomalous ΔC_p for the nucleotides largely to a change in the conformation of the domain structure and local motion in the lid region of N-Hsp90 with the concomitant exposure of hydrophobic amino acid side chains.     

Survival of Escherichia coli in the environment: fundamental and public health aspects

In this review, our current understanding of the species Escherichia coli and its persistence in the open environment is examined. E. coli consists of six different subgroups, which are separable by genomic analyses. Strains within each subgroup occupy various ecological niches, and can be broadly characterized by either commensalistic or different pathogenic behaviour. In relevant cases, genomic islands can be pinpointed that underpin the behaviour. Thus, genomic islands of, on the one hand, broad environmental significance, and, on the other hand, virulence, are highlighted in the context of E. coli survival in its niches. A focus is further placed on experimental studies on the survival of the different types of E. coli in soil, manure and water. Overall, the data suggest that E. coli can persist, for varying periods of time, in such terrestrial and aquatic habitats. In particular, the considerable persistence of the pathogenic E. coli O157:H7 is of importance, as its acid tolerance may be expected to confer a fitness asset in the more acidic environments. In this context, the extent to which E. coli interacts with its human/animal host and the organism''s survivability in natural environments are compared. In addition, the effect of the diversity and community structure of the indigenous microbiota on the fate of invading E. coli populations in the open environment is discussed. Such a relationship is of importance to our knowledge of both public and environmental health.     

Quelques données ultrastructurales sur un myoépithélium: le pharynx d'un Bryozoaire

Résumé L'épithélium pharyngien d'Alcyonidium polyoum possède des cellules pourvues d'une très grande vacuole. L'incompressibilité du liquide vacuolaire permet un élargissement brusque de l'organe lors de la contraction du manchon musculaire strié qui enserre cette vacuole. Les fibres musculaires sont insérées sur le plasmalemme apical par des filaments unitifs. Le point d'attache est relié à la lame amorphe du cell-coat qui entoure les microvillosités par des fibrilles, réalisant probablement une liaison mécamique plus efficace. Le reticulum sarcoplasmique porte des ribosomes. Le cytoplasme apical renferme des vésicules de diverses catégories.

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Some ultrastructural data about a myoepithelium: The pharynx of a bryozoan

Summary Pharyngeal cells of Alcyonidium polyoum (Bryozoa) are provided with very large vacuoles. Each vacuole is enveloped by a thin layer of striated muscle, whose contraction enlarges the organ. Filaments join the muscular elements to the apical plasmalemma. This point of muscular insertion is connected by fibrils with the amorphic lamina of cell-coat which surrounds the microvilli. Ribosomes are often found on dyads. Various vesicles are located in the apical cytoplasm.

     

Water and solute transport by the Malpighian tubules of the stick insect,Carausius morosus

Summary Structural features of the principal, urine-secreting cells (type 1 cells) of the Malpighian tubules of Carausius are de scribedquantitatively and discussed in relation to possible mechanisms of water and solute transport. Mitochondria are arranged in two bands of about equal volume near to the basal and apical surfaces, suggesting active processes occur at both surfaces. Basal infoldings and apical microvilli which greatly amplify the cell surface are probably primarily devices to increase the passive permeability of the tissue to solutes. They do not provide functionally significant standing-osmotic-gradients. The extensive endoplasmic reticulum is locally differentiated into several components and ramifies between the infoldings and along microvilli but probably is not an intracellular conduit for the majority of urinary constituents. Vesicles and stages in their formation or liberation are observed both basally and apically although they probably do not contribute significantly to transcellular transport. At present it remains a problem to satisfactorily account for observations that the urine of Carausius can be hypotonic.This investigation formed part of a dissertation for the degree of Ph. D. in the University of Newcastle upon Tyne. It is a pleasure to thank Prof. J. Shaw for his advice and encouragement and the Science Research Council for financial support.     

Degradation of cholecystokinin octapeptide,related fragments and analogs by human and rat plasma in vitro

The rate of degradation of cholecystokinin octapeptide, related fragments and analogs by human and rat plasma was investigated, using high pressure liquid chromatography for the separation and identification of the degradation products.CCK tetrapeptide showed a half-life of 13 min in human plasma while its cleavage in rat plasma occurred at a very high rate (half-life of less than 1 min).The kinetics of disappearance of both sulphated and unsulphated CCK-8 indicated more than a single rate of degradation; the faster degrading system showed a half-life of 18 min for unsulphated CCK-8 and of 50 min for the sulphated peptide in human plasma as compared respectively with 5 and 17 min in rat plasma. The cleavage of CCK-8 was inhibited by bestatin and puromycin, suggesting that aminopeptidases play a major role in the breakdown of this peptide.CCK-9 analogs were rapidly converted into their corresponding octapeptides (half-life of 2.7 min in human plasma). This conversion was inhibited by puromycin and bestatin, suggesting the participation of aminopeptidase(s) probably specific for basic amino acids.CCK decapeptide exhibited a greater stability than the nonapeptides (half-life of 30 and 45 min in human and rat plasma respectively) and also gave rise to CCK-8 as degradation product. This cleavage was not affected by aminopeptidase inhibitors but was decreased by aprotinin (Trasylol®), suggesting that trypsin-like and/or kallikrein-like enzyme(s) were involved in the plasma metabolism of this peptide.     

Alpha‐fetoprotein accelerates the progression of hepatocellular carcinoma by promoting Bcl‐2 gene expression through an RA‐RAR signalling pathway

Previous studies have found that alpha‐fetoprotein (AFP) can promote the proliferation of hepatoma cells and accelerate the progression of hepatocellular carcinoma (HCC). However, the exact mechanism of action remains unclear. Recent bioinformatics studies have predicted the possible interaction between AFP and retinoic acid receptors (RARs). Thus, the purpose of this study was to investigate the molecular mechanism through which AFP promotes tumour cell proliferation by interfering with the RA‐RAR signal pathway. Our data indicated that AFP could significantly promote the proliferation and weaken ATRA‐induced apoptosis of hepatoma cells. Besides, cytoplasmic AFP interacts with RAR, disrupting its entrance into the nucleus, which in turn affects the expression of the Bcl‐2 gene. In addition, knockdown of AFP in HepG2 cells was synchronously associated with an incremental increase of RAR binding to DNA, as well as down‐regulation of Bcl‐2; the opposite effect was observed in AFP gene‐transfected HLE cells. Moreover, a similar effect of AFP was detected in tumour tissues with high serum AFP, but not in adjacent non‐cancerous liver tissues, or HCC tissues with low serum AFP levels. These results indicate that AFP acts as signalling molecule and prevents RAR from entering into the nucleus by interacting with RAR, thereby promoting the expression of Bcl‐2. Our data reveal a novel mechanism through which AFP regulates Bcl‐2 expression and further suggest that AFP may be used as a novel target for treating HCC.