The identity of the PARylated proteins was tested by affinity precipitation with PAR-binding reagent followed by western blots. Plasma membrane-associated PAR was tested by immunocytofluorescence (ICF) and subcellular fractionation in Vero cells while TNKS role in this structure and cell junction assembly was evaluated using specific inhibitors. Tankyrase-binding motif (TBM) sequences in hVCL gene were identified and VCL sequences from various vertebrates, Drosophila melanogaster and Caenorhabditis elegans were aligned and compared. TNKS poly(ADP-ribosylates) (PARylates) epithelial belt VCL, affecting its functions in AJ, including cell shape maintenance. TNKS has been formerly involved in the assembly of epithelial cell junctions. Vinculin (VCL) participates in the anchorage of F-actin to the AJ, regulating its functions, and colocalized with the PAR belt. We wondered which PARP synthesizes the belt and which is the PARylation target protein. F-actin depolymerization alters the AJ and PAR belts while PARP inhibitors prevent the assembly of the AJ belt and cortical actin. In the epithelial belt, AJ are accompanied by a PAR belt and a subcortical F-actin ring. Four PARPs synthesize PAR, namely PARP-1/2 and Tankyrase-1/2 (TNKS). Poly(ADP-ribose) polymerases (PARPs) catalyze the synthesis of poly(ADP-ribose) (PAR) as a posttranslational modification.
Still, their composition has not been completely disclosed. PeerJ 9: e11442 Īdherens junctions (AJ) are involved in cancer, infections and neurodegeneration. First body of evidence suggesting a role of a tankyrase-binding motif (TBM) of vinculin (VCL) in epithelial cells. Cite this article Vilchez Larrea S, Valsecchi WM, Fernández Villamil SH, Lafon Hughes LI. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited. Licence This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. 5 Departamento de Genética, Instituto de Investigaciones Biológicas Clemente Estable, Ministerio de Educación y Cultura, Montevideo, Uruguay DOI 10.7717/peerj.11442 Published Accepted Received Academic Editor Gwyn Gould Subject Areas Biochemistry, Cell Biology, Genetics, Molecular Biology Keywords Poly(ADP)ribose, PARP, Tankyrase, Epithelia, Adherens junctions, Vinculin, Vero, MCF-7, Tankyrase-binding motif (TBM), Epithelial to mesenchymal transition (EMT) Copyright © 2021 Vilchez Larrea et al.
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