Post-translational allosteric activation of the P2X7 receptor through glycosaminoglycan chains of CD44 proteoglycans

Abstract

Here, we present evidence for the positive allosteric modulation of the P2X7 receptor through glycosaminoglycans (GAGs) in CHO (cell line derived from the ovary of the Chinese hamster) cells. The marked potentiation of P2X7 activity through GAGs in the presence of non-saturating agonists concentrations was evident with the endogenous expression of the receptor in CHO cells. The presence of GAGs on the surface of CHO cells greatly increased the sensitivity to adenosine 5′-triphosphate and changed the main P2X7 receptor kinetic parameters EC50, Hill coefficient and Emax. GAGs decreased the allosteric inhibition of P2X7 receptor through Mg2+. GAGs activated P2X7 receptor-mediated cytoplasmic Ca2+ influx and pore formation. Consequently, wild-type CHO-K1 cells were 2.5-fold more sensitive to cell death induced through P2X7 agonists than mutant CHO-745 cells defective in GAGs biosynthesis. In the present study, we provide the first evidence that the P2X7 receptor interacts with CD44 on the CHO-K1 cell surface. Thus, these data demonstrated that GAGs positively modulate the P2X7 receptor, and sCD44 is a part of a regulatory positive feedback loop linking P2X7 receptor activation for the intracellular response mediated through P2X7 receptor stimulation.

Heparan sulfate proteoglycan deficiency up‐regulates the intracellular production of nitric oxide in Chinese hamster ovary cell lines

“…It maintains the previously described associations with elements of the matrix or cytoskeletal elements, and it is also involved in the antioxidant system in gastrointestinal cancer cells (Ishimoto et al, ). CD44 proteoglycan staining was detected in large amounts in CHO‐K1 cells and very low amounts in CHO‐745 cells (Figure ), confirming the results of the gene expression observed in a previous work from our group (Moura et al, ). These data indicate that the difference in the rate of formation of NO in the CHO cells can be related to the presence of proteoglycans/GAGs related to CD44 in CHO cells.…”

“…Next, we used heparin mimicking endogenous GAGs chains of HSPG on the cell surface (Moura et al, ). We examined the effects of depleting or loading GAGs chain on the basal NO production in both CHO cell lines.…”

Antimicrobial endotoxin‐neutralizing peptides promote keratinocyte migration via P2X7 receptor activation and accelerate wound healing in vivo

“…Recent studies indicate that calcium influx triggered by ATP-induced P2X7 receptor activation leads to ADAM10-mediated ectodomain shedding of CD44 resulting in formation of soluble CD44 (sCD44) (Stamenkovic and Yu, 2009). Thus, the peptide-increased calcium influx could further induce sCD44-mediated allosteric activation of the P2X7 receptor through a positive feed-back loop (Moura et al, 2015).…”

PAR-1 and PAR-2 Expression Is Enhanced in Inflamed Odontoblast Cells

“…The resulting fluorescence was measured using a Flex Station 3 microplate reader (Molecular Devices). The Fluo-4 was excited at 490 nm and the emission was detected at 525 nm (Moura et al 2015).…”