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1.  NO Metabolites Levels in Human Red Blood Cells are Affected by Palytoxin, an Inhibitor of Na+/K+-ATPase Pump 
Palytoxin (PTX), a marine toxin, represents an increasing hazard for human health. Despite its high toxicity for biological systems, the mechanisms triggered by PTX, are not well understood. The high affinity of PTX for erythrocyte Na+/K+-ATPase pump is largely known, and it indicates PTX as a sensitive tool to characterize the signal transducer role for Na+/K+-ATPase pump. Previously, it has been reported that in red blood cells (RBC), probably via a signal transduction generated by the formation of a PTX-Na+/K+-ATPase complex, PTX alters band 3 functions and glucose metabolism. The present study addresses the question of which other signaling pathways are regulated by Na+/K+-ATPase in RBC. Here it has been evidenced that PTX following its interaction with Na+/K+-ATPase pump, alters RBC morphology and this event is correlated to decreases by 30% in nitrites and nitrates levels, known as markers of plasma membrane eNOS activity. Orthovanadate (OV), an antagonist of PTX binding to Na+/K+-ATPase pump, was able to reverse the effects elicited by PTX. Finally, current investigation firstly suggests that Na+/K+-ATPase pump, following its interaction with PTX, triggers a signal transduction involved in NO metabolism regulation.
doi:10.2174/1874091X01408010068
PMCID: PMC4157343  PMID: 25246985
Metabolism; Na+/K+-ATPase pump; Nitric oxide; Palytoxin; Red blood cells.
2.  Insertion of a myc-tag within α-dystroglycan domains improves its biochemical and microscopic detection 
BMC Biochemistry  2012;13:14.
Background
Epitope tags and fluorescent fusion proteins have become indispensable molecular tools for studies in the fields of biochemistry and cell biology. The knowledge collected on the subdomain organization of the two subunits of the adhesion complex dystroglycan (DG) enabled us to insert the 10 amino acids myc-tag at different locations along the α-subunit, in order to better visualize and investigate the DG complex in eukaryotic cells.
Results
We have generated two forms of DG polypeptides via the insertion of the myc-tag 1) within a flexible loop (between a.a. 170 and 171) that separates two autonomous subdomains, and 2) within the C-terminal domain in position 500. Their analysis showed that double-tagging (the β-subunit is linked to GFP) does not significantly interfere with the correct processing of the DG precursor (pre-DG) and confirmed that the α-DG N-terminal domain is processed in the cell before α-DG reaches its plasma membrane localization. In addition, myc insertion in position 500, right before the second Ig-like domain of α-DG, proved to be an efficient tool for the detection and pulling-down of glycosylated α-DG molecules targeted at the membrane.
Conclusions
Further characterization of these and other myc-permissive site(s) will represent a valid support for the study of the maturation process of pre-DG and could result in the creation of a new class of intrinsic doubly-fluorescent DG molecules that would allow the monitoring of the two DG subunits, or of pre-DG, in cells without the need of antibodies.
doi:10.1186/1471-2091-13-14
PMCID: PMC3432625  PMID: 22835149
3.  Duplication of the dystroglycan gene in most branches of teleost fish 
Background
The dystroglycan (DG) complex is a major non-integrin cell adhesion system whose multiple biological roles involve, among others, skeletal muscle stability, embryonic development and synapse maturation. DG is composed of two subunits: α-DG, extracellular and highly glycosylated, and the transmembrane β-DG, linking the cytoskeleton to the surrounding basement membrane in a wide variety of tissues. A single copy of the DG gene (DAG1) has been identified so far in humans and other mammals, encoding for a precursor protein which is post-translationally cleaved to liberate the two DG subunits. Similarly, D. rerio (zebrafish) seems to have a single copy of DAG1, whose removal was shown to cause a severe dystrophic phenotype in adult animals, although it is known that during evolution, due to a whole genome duplication (WGD) event, many teleost fish acquired multiple copies of several genes (paralogues).
Results
Data mining of pufferfish (T. nigroviridis and T. rubripes) and other teleost fish (O. latipes and G. aculeatus) available nucleotide sequences revealed the presence of two functional paralogous DG sequences. RT-PCR analysis proved that both the DG sequences are transcribed in T. nigroviridis. One of the two DG sequences harbours an additional mini-intronic sequence, 137 bp long, interrupting the uncomplicated exon-intron-exon pattern displayed by DAG1 in mammals and D. rerio. A similar scenario emerged also in D. labrax (sea bass), from whose genome we have cloned and sequenced a new DG sequence that also harbours a shorter additional intronic sequence of 116 bp. Western blot analysis confirmed the presence of DG protein products in all the species analysed including two teleost Antarctic species (T. bernacchii and C. hamatus).
Conclusion
Our evolutionary analysis has shown that the whole-genome duplication event in the Class Actinopterygii (ray-finned fish) involved also DAG1. We unravelled new important molecular genetic details about fish orthologous DGs, which might help to increase the current knowledge on DG expression, maturation and targeting and on its physiopathological role in higher organisms.
doi:10.1186/1471-2199-8-34
PMCID: PMC1885269  PMID: 17509131

Results 1-3 (3)