Biomedical subjects

Igor Andreev

Publications and source records attributed to Igor Andreev.

3 recordsLinked to original sources

PubMed2006

Evidence for direct amelogenin-target cell interactions using dynamic force spectroscopy.

Increasing evidence suggests that amelogenin, long held to be a structural protein of developing enamel matrix, may also have cell signaling functions. However, a mechanism for amelogenin cell signaling has yet to be described. The aim of the present study was to use dynamic chemical force spectroscopy to measure amelogenin interactions with possible target cells. Full-length amelogenin (rM179) was covalently attached to silicon nitride AFM tips. Synthetic RGD peptides and unmodified AFM tips were used as controls. Amelogenin-RGD cell binding force measurements were carried out using human periodontal ligament fibroblasts (HPDF) from primary explants and a commercially available osteoblast-like human sarcoma cell line as the targets. Results indicated a linear logarithmic dependence between loading rate and unbinding force for amelogenin-RGD target cells across the range of loading rates used. For RGD controls, binding events measured at 5.5 nN s-1 force loading rate resulted in a mean force of 60 pN. Values for amelogenin-fibroblast and amelogenin-osteoblast-like cell unbinding forces, measured at similar loading rates, were 50 and 55 pN, respectively. These data suggest that amelogenin interacts with potential target cells with forces characteristic of specific ligand-receptor binding, suggesting a direct effect for amelogenin at target cell membranes.

Amelogenin↗

PubMed2004

The C-terminal 33 amino acids of the cucumber mosaic virus 3a protein affect virus movement, RNA binding and inhibition of infection and translation.

The capsid protein (CP) of Cucumber mosaic virus (CMV) is required for cell-to-cell movement, mediated by the 3a movement protein (MP). Deletion of the C-terminal 33 amino acids of the CMV 3a MP (in the mutant designated 3aDeltaC33 MP) resulted in CP-independent cell-to-cell movement, but not long-distance movement. RNA-binding studies done in vitro using isolated bacterially expressed MP showed that the 3aDeltaC33 MP bound RNA more strongly, with fewer regions sensitive to RNase and formed cooperatively bound complexes at lower ratios of protein : RNA than the wild-type (wt) 3a MP. Analysis of the architecture of the complexes by atomic force microscopy showed that the wt 3a MP formed a single type of complex with RNA, resembling beads on a string. By contrast, the 3aDeltaC33 MP formed several types of complexes, including complexes with virtually no MP bound or thicker layers of MP bound to the RNA. Assays showed that protein-RNA complexes containing high levels of either MP inhibited the infectivity and in vitro translatability of viral RNAs. The 3aDeltaC33 MP inhibited these processes at lower ratios of protein : RNA than the wt 3a MP, consistent with its stronger binding properties. The apparent contradiction between these inhibition data and the CP-independent cell-to-cell movement of CMV expressing the 3aDeltaC33 MP is discussed.

Chenopodium↗

PubMed2001

Umbravirus-encoded movement protein induces tubule formation on the surface of protoplasts and binds RNA incompletely and non-cooperatively.

Various functions of the cell-to-cell movement protein (MP) of Groundnut rosette virus (GRV) were analysed. The GRV ORF4-encoded protein was shown by immunofluorescence microscopy to generate tubular structures that protrude from the surface of the protoplast. The protein encoded by ORF4 was assessed also for RNA-binding properties. This protein was tagged at its C terminus with six histidine residues, produced in Escherichia coli using an expression vector and purified by affinity chromatography. Gel retardation analysis demonstrated that, in contrast to many other viral MPs, including the 3a MP of Cucumber mosaic virus (CMV), the ORF4-encoded protein bound non-cooperatively to viral ssRNA and formed complexes of low protein:RNA ratios. Competition binding experiments showed that the ORF4-encoded protein bound to both ssRNA and ssDNA without sequence specificity, but did not bind to dsDNA. UV cross-linking and nitrocellulose membrane-retention assays confirmed that both the GRV and the CMV MPs formed complexes with ssRNA and that these complexes showed similar stability in NaCl. Probing the MP-RNA complexes by atomic force microscopy demonstrated that the ORF4-encoded protein bound RNA incompletely, leaving protein-free RNA segments of varying length, while the CMV 3a protein formed highly packed complexes. The significance of the two properties of limited RNA binding and tubule formation of the umbraviral MP is discussed.

Arachis↗