Neurite outgrowth inhibitory levels of organophosphates induce tissue transglutaminase activity in differentiating N2a c
- PDF / 1,313,559 Bytes
- 15 Pages / 595.276 x 790.866 pts Page_size
- 0 Downloads / 165 Views
ORGAN TOXICITY AND MECHANISMS
Neurite outgrowth inhibitory levels of organophosphates induce tissue transglutaminase activity in differentiating N2a cells: evidence for covalent adduct formation Ibtesam S. Almami1,2 · Maha A. Aldubayan1,3 · Shatha G. Felemban1,4 · Najiah Alyamani1,5 · Richard Howden1 · Alexander J. Robinson1,6 · Tom D. Z. Pearson1 · David Boocock1 · Alanood S. Algarni1,7 · A. Christopher Garner1 · Martin Griffin8 · Philip L. R. Bonner1 · Alan J. Hargreaves1 Received: 19 January 2020 / Accepted: 14 July 2020 © The Author(s) 2020
Abstract Organophosphate compounds (OPs) induce both acute and delayed neurotoxic effects, the latter of which is believed to involve their interaction with proteins other than acetylcholinesterase. However, few OP-binding proteins have been identified that may have a direct role in OP-induced delayed neurotoxicity. Given their ability to disrupt Ca2+ homeostasis, a key aim of the current work was to investigate the effects of sub-lethal neurite outgrowth inhibitory levels of OPs on the Ca2+-dependent enzyme tissue transglutaminase (TG2). At 1–10 µM, the OPs phenyl saligenin phosphate (PSP) and chlorpyrifos oxon (CPO) had no effect cell viability but induced concentration-dependent decreases in neurite outgrowth in differentiating N2a neuroblastoma cells. The activity of TG2 increased in cell lysates of differentiating cells exposed for 24 h to PSP and chlorpyrifos oxon CPO (10 µM), as determined by biotin-cadaverine incorporation assays. Exposure to both OPs (3 and/or 10 µM) also enhanced in situ incorporation of the membrane permeable substrate biotin-X-cadaverine, as indicated by Western blot analysis of treated cell lysates probed with ExtrAvidin peroxidase and fluorescence microscopy of cell monolayers incubated with FITC-streptavidin. Both OPs (10 µM) stimulated the activity of human and mouse recombinant TG2 and covalent labelling of TG2 with dansylamine-labelled PSP was demonstrated by fluorescence imaging following SDS-PAGE. A number of TG2 substrates were tentatively identified by mass spectrometry, including cytoskeletal proteins, chaperones and proteins involved protein synthesis and gene regulation. We propose that the elevated TG2 activity observed is due to the formation of a novel covalent adduct between TG2 and OPs. Keywords Organophosphate toxicity · Neurite outgrowth · Covalent adduct · Tissue transglutaminase
* Alan J. Hargreaves [email protected] 1
School of Science and Technology, Nottingham Trent University, Nottingham NG11 8NS, UK
2
Department of Biology, College of Science, Qassim University, Al‑Qassim, Saudi Arabia
3
Department of Pharmacology and Toxicology, College of Pharmacy, Qassim University, Al‑Qassim, Saudi Arabia
4
Department of Medical Laboratory Science, Fakeeh College for Medical Science, Jeddah, Saudi Arabia
5
Department of Biology, Faculty of Science, University of Jeddah, Jeddah, Kingdom of Saudi Arabia
6
Department of Life Sciences, School of Health Sciences, Birmingham City University, City S
Data Loading...
