Isolation and1H-NMR spectroscopic identification of the glucose-containing lipid-linked precursor oligosaccharide ofN-linked carbohydrate chains

The lipid-linked precursor ofN-type glycoprotein oligosaccharides was isolated from porcine thyroid microsomes after in cubation with UDP[³H] Glucose. The carbohydrate was released from dolichol pyrophosphate by mild acid hydrolysis, purified by gel filtration and characterized by 500-MHz¹H-NMR spectroscopy in combination with enzymatic degradation. The parent oligosaccharide was found to be Glc₃Man₉Glc-NAc₂. The three glucose residues are present in the linear sequence Glcα1-2Glα1-3 Glc, the latter being α(1-3)-linked to one of the mannose residues. In order to establish the branch location of the triglucosyl unit, the parent compound was digested with jack-bean α-mannosidase. The oligosaccharide product was purified by gel filtration, and identified by¹H-NMR as Glc₃Man₅GlcNAc₂ lacking the mannose residues A, D₂, B and D₃. Therefore, the structure of the precursor oligosaccharide is as follows: $$\begin{gathered} c b a D_1 C 4 \hfill \\ Glc\alpha 1 - 2Glc\alpha 1 - 3Glc\alpha 1 - 3Man\alpha 1 - 2Man\alpha 1 - 2Man\alpha 1 \hfill \\ 3 \swarrow 3 2 1 \hfill \\ Man\alpha 1 - 2Man\alpha 1 Man\beta 1 - 4GlcNAc\beta 1 - 4GlcNAc \hfill \\ D_{2 } A 3 6 \hfill \\ Man\alpha 1 \hfill \\ 6 \hfill \\ Man\alpha 1 - 2Man\alpha 1 \nwarrow 4 \hfill \\ D_3 B \hfill \\ \end{gathered} $$      

Protein synthetic activity and adenylate energy charge in Rhein-treated cultured human glioma cells.

The effect of Rhein (RH) on the protein synthetic activity and adenylate energy charge in human glioma cells cultured in vitro has been investigated. The results demonstrate that in RH-treated cells, the protein synthesis is strongly decreased, but no modifications in the qualitative pattern occur. The extent of inhibition is a function of the drug concentration as well as of the time of exposure. Such an inhibition must be ascribed mainly to a reduction of adenylate energy charge brought about by RH because of its effect on respiration and glycolysis. The correlation between the adenylate energy charge and cell viability, as well as the possibility of using rhein as a biochemical modulator to reduce or to reverse multidrug resistance, are also discussed.     

Morphometric stepwise discriminant analysis of three genetically identified species within Pseudoterranova decipiens (Krabbe, 1878) (Nematoda: Ascaridida)

Univariate and multivariate statistical analyses have been performed on 19 morphometric variables of adult male specimens belonging to three genetically identified species within Pseudoterranova decipiens (Nematoda: Ascaridida) parasitic in the digestive tract of seals. Two morphometric keys are proposed for the identification of the three species. One key, which uses two variables, determines a frequency of error of 3.8% (3/79). The second key, which uses two canonical discriminant functions based on seven variables previously selected with a stepwise procedure, gives 100% (76/76) accurate classification.     

Parasites of Salmo trutta L. from the Tirino River. Part IV. Intermediate hosts of Cyathocephalus truncatus (Pallas, 1781) and Dentitruncus truttae Sinzar, 1955

During studies on helminth parasites of brown trout (Salmo trutta L.) from the river Tirino (L'Aquila - Italy), the intermediate hosts of Cyathocephalus truncatus (Pallas, 1781) and Dentitruncus truttae Sinzar, 1955 have been searched for. A total of 15,444 amphipoda belonging to the following 3 species: Echinogammarus roco Karman, 1973, Echinogammarus tibaldii Pinkster and Stock, 1970, and Gammarus italicus Goedmakers and Pinkster, 1977, have been dissected. E. roco, E. tibaldii and G. italicus were found infected with procercoid of Cyathocephalus truncatus (new hosts record and first procercoid record in Italy). E. roco, E. tibaldii and G. italicus were found infected with acanthella and cystacanth of Dentitruncus truttae (first record of intermediate hosts). Accordingly the ecology of the cestod C. truncatus and of the acanthocephalan D. truttae in their intermediate hosts has been studied considering principally the incidence and the seasonal occurence of the parasites.     

The small molecule SI113 hinders epithelial-to-mesenchymal transition and subverts cytoskeletal organization in human cancer cells

The small molecule SI113 is an inhibitor of the kinase activity of SGK1, a key biological regulator acting on the PI3K/mTOR signal transduction pathway. Several studies demonstrate that this compound is able to strongly restrain cancer growth in vitro and in vivo, alone or in associative antineoplastic treatments, being able to elicit an autophagic response, either cytotoxic or cytoprotective. To elucidate more exhaustively the molecular mechanisms targeted by SI113, we performed activity-based protein profiling (ABPP) proteomic analysis using a kinase enrichment procedure. This technique allowed the identification via mass spectrometry of novel targets of this compound, most of them involved in functions concerning cell motility and cytoskeletal architecture. Using a glioblastoma multiforme, hepatocarcinoma and colorectal carcinoma cell line, we recognized an inhibitory effect of SI113 on cell migration, invading, and epithelial-to-mesenchymal transition. In addition, these cancer cells, when exposed to this compound, showed a remarkable subversion of the cytoskeletal architecture characterized by F-actin destabilization, phospho-FAK delocalization, and tubulin depolimerization. These results were definitely concordant in attributing to SI113 a key role in hindering cancer cell malignancy and, due to its negligible in vivo toxicity, can sustain performing a Phase I clinical trial to employ this drug in associative cancer therapy.     

Cyclin T: three forms for different roles in physiological and pathological functions

Cyclins are members of family of proteins involved in the cell cycle regulation. They are regulatory subunits of complexes with proteins called cyclin-dependent kinases (CDKs). There are three forms of cyclin T: cyclin T1, cyclin T2a, and T2b. All cyclin T contain an N-terminal "cyclin homology box," the most conserved region among different members of the cyclin family that serves to bind CDK9. In addition to the N-terminal cyclin domain, cyclin T contains a putative coiled-coil motif, a His-rich motif, and a C-terminal PEST sequence. The CDK9/cyclin T complex is able to activate gene expression in a catalytic-dependent manner, phosphorylating the carboxy-terminal domain (CTD) of RNA polymerase II. In addition, only cyclin T1 supports interactions between Tat and TAR. The interaction of Tat with cyclin T1 alters the conformation of Tat to enhance the affinity and specificity of the Tat:TAR interaction. On the other hand, CDK9/cyclin T2 complexes are involved in the regulation of terminal differentiation in muscle cells.     

cDNA array technology in melanoma: an overview

Genetic aberrations, mostly resulting in changes in gene expression, are critical events in cancer onset and progression. The advent of the cDNA array technology allows the screening and the efficient measurement of expression of thousands genes simultaneously in a wide spectrum of experimental and clinical models. This genomic scale approach is being currently used to obtain global views of human cancer gene expression and to identify genetic markers that might be important for diagnosis, prognosis, and therapy. This review discusses some recent findings obtained by means of cDNA arrays investigating the human melanoma.     

Detection of quorum sensing signals in the haloalkaliphilic archaeon Natronococcus occultus

Bacteria communicate at high cell density through quorum sensing, however, there are no reports about this mechanism in archaea. The archaeon Natronococcus occultus produces an extracellular protease at the end of growth. Early production of protease activity was observed when a low density culture was incubated with late exponential conditioned medium suggesting the presence of factor(s) inducing this activity. Conditioned medium and ethyl acetate extracts corresponding to the transition from exponential to stationary phase showed a positive signal in Agrobacterium biosensor. We report the detection of potential autoinducer molecules of the acylated homoserine lactone type in the archaeon N. occultus. These molecules may be responsible for the production/activation of extracellular protease.