Peptidylarginine Deiminases Post-Translationally Deiminate Prohibitin and Modulate Extracellular Vesicle Release and MicroRNAs in Glioblastoma Multiforme

Abstract

Glioblastoma multiforme (GBM) is the most aggressive form of adult primary malignant brain tumour with poor prognosis. Extracellular vesicles (EVs) are a key-mediator through which GBM cells promote a pro-oncogenic microenvironment. Peptidylarginine deiminases (PADs), which catalyze the post-translational protein deimination of target proteins, are implicated in cancer, including via EV modulation. Pan-PAD inhibitor Cl-amidine affected EV release from GBM cells, and EV related microRNA cargo, with reduced pro-oncogenic microRNA21 and increased anti-oncogenic microRNA126, also in combinatory treatment with the chemotherapeutic agent temozolomide (TMZ). The GBM cell lines under study, LN18 and LN229, differed in PAD2, PAD3 and PAD4 isozyme expression. Various cytoskeletal, nuclear and mitochondrial proteins were identified to be deiminated in GBM, including prohibitin (PHB), a key protein in mitochondrial integrity and also involved in chemo-resistance. Post-translational deimination of PHB, and PHB protein levels, were reduced after 1 h treatment with pan-PAD inhibitor Cl-amidine in GBM cells. Histone H3 deimination was also reduced following Cl-amidine treatment. Multifaceted roles for PADs on EV-mediated pathways, as well as deimination of mitochondrial, nuclear and invadopodia related proteins, highlight PADs as novel targets for modulating GBM tumour communication.

Deiminated proteins in extracellular vesicles and serum of llama (Lama glama)—Novel insights into camelid immunity

“…PADs have been identified throughout phylogeny from bacteria to mammals, with 5 tissue specific PAD isozymes in mammals, 3 in chicken, 1 in bony fish and arginine deiminase homologues in bacteria (Vossenaar et al, 2003;Rebl et al, 2010;Magnadóttir et al, 2018a, a;Kosgodage et al, 2019). While studies on PADs in relation to human pathophysiology, including cancer, autoimmune and neurodegenerative diseases (Wang and Wang, 2013;Witalison et al, 2015;Lange et al, 2017;Kosgodage et al, 2017Kosgodage et al, & 2018 and CNS regeneration (Lange et al, 2011 and2014) exist, relatively little phylogenetic research has been carried out on PADs in relation to normal physiology and evolutionary acquired adaptions of the immune system. Recent comparative studies focussing on roles for PADs in teleost fish have identified post-translational deimination in key proteins of innate, adaptive and mucosal immunity (Magnadóttir et al, 2018a, b;Magnadottir et al, 2019a;Magnadóttir et al, 2019b).…”

“…As PADs have been identified to be a key regulator of extracellular (EV)-release, a mechanism that has been found to be phylogenetically conserved from bacteria to mammals (Kholia et al, 2015;Kosgodage et al, 2017Kosgodage et al, , 2018Gavinho et al, 2019;Kosgodage et al, 2019), the characterisation of EVs in camelids is of further interest. Extracellular vesicles (EVs) are found in most body fluids and participate in cellular communication via transfer of cargo proteins and genetic material (Inal et al, 2013;Colombo et al, 2014;Lange et al, 2017;Turchinovich et al, 2019;Vagner et al, 2019).…”

“…In addition to acetylation, phosphorylation and ubiquitination, histones are known to undergo deimination, including H2B (Sohn et al, 2015) and H2A (Hagiwara et al, 2005), as identified in this study in llama. Other histones that are known to undergo deimination include H3 and H4 (Chen et al, 2014;Kosgodage et al, 2018).…”

Peptidylarginine Deiminase inhibition abolishes the production of large extracellular vesicles fromGiardia intestinalis, affecting host-pathogen interactions by hindering adhesion to host cells

“…While it is known that EVs are released by multiple mechanisms, some advances in understanding of their biogenesis has been elucidated via studies on the peptidylarginine deiminase (PAD)-mediated pathway of EV release (Kholia et al, 2015; Kosgodage et al, 2017; Lange et al, 2017; Kosgodage et al, 2018a). PADs are phylogenetically conserved enzymes from bacteria to mammals (Vossenaar et al, 2003; Magnadottir et al, 2018), including in Giardia (arginine deiminase GiADI; Trejo-Soto et al, 2016).…”

“…While exact roles for PADs in EV biogenesis and release remain to be fully elucidated, effects on cytoskeletal, nuclear and mitochondrial proteins have been reported (Kholia et al, 2015; Kosgodage et al, 2018). As pharmacological PAD-inhibitors have previously been shown to be potent inhibitors of EV release in various cancer cells, as well as to modulate EV cargo (Kholia et al, 2015; Kosgodage et al, 2017; Kosgodage et al, 2018a), we sought to investigate a phylogenetically conserved influence of such PAD-inhibitors on the EV production of our protozoa model; putatively targeting GiADI and therefore also reveal a role for GiADI in EV release.…”