Modulation of calcium-induced cell death in human neural stem cells by the novel peptidylarginine deiminase–AIF pathway

Abstract

PADs (peptidylarginine deiminases) are calcium-dependent enzymes that change protein-bound arginine to citrulline (citrullination/deimination) affecting protein conformation and function. PAD up-regulation following chick spinal cord injury has been linked to extensive tissue damage and loss of regenerative capability. Having found that human neural stem cells (hNSCs) expressed PAD2 and PAD3, we studied PAD function in these cells and investigated PAD3 as a potential target for neuroprotection by mimicking calcium-induced secondary injury responses. We show that PAD3, rather than PAD2 is a modulator of cell growth/death and that PAD activity is not associated with caspase-3-dependent cell death, but is required for AIF (apoptosis inducing factor)-mediated apoptosis. PAD inhibition prevents association of PAD3 with AIF and AIF cleavage required for its translocation to the nucleus. Finally, PAD inhibition also hinders calcium-induced cytoskeleton disassembly and association of PAD3 with vimentin, that we show to be associated also with AIF; together this suggests that PAD-dependent cytoskeleton disassembly may play a role in AIF translocation to the nucleus. This is the first study highlighting a role of PAD activity in balancing hNSC survival/death, identifying PAD3 as an important upstream regulator of calcium-induced apoptosis, which could be targeted to reduce neural loss, and shedding light on the mechanisms involved.

Inhibition of peptidylarginine deiminase attenuates inflammation and improves survival in a rat model of hemorrhagic shock

“…PADs have been shown to be upstream regulators of Ca 2+ -induced cell death in human neural cells, and PAD inhibition protects cells from apoptosis by hindering calcium-induced cytoskeleton disassembly [15]. The present in vitro findings are in accordance with a previous study in a murine model of ischemic insult where pan-PAD inhibition resulted in significantly reduced apoptotic cell death [16].…”

Protein Arginine Deiminases and Associated Citrullination: Physiological Functions and Diseases Associated with Dysregulation

“…PAD1 targets keratin and filaggrin [7]; PAD2 targets myelin basic protein (MBP) [30], vimentin [42], actin, and histones [39]; PAD 3 targets filaggrin, trichohyalin [7], apoptosis-inducing factor (AIF), and vimentin [43]; PAD4 targets multiple proteins involved in gene regulation (histones [24, 44], inhibitor of growth 4 (ING4) [45], p300 [46], p21 [47]) and apoptosis (nucleophosmin [48], nuclear lamin C [49]) ; and there are no known protein substrates of PAD6 (Table 1) [35]. Several of these protein targets of PAD citrullination will be described throughout the next sections that have a high arginine content (~10%) and/or have particular arginine-rich regions that are critical for their structure and function.…”

“…Additionally, Mizoguchi et al (1998) reported the citrullination of a 70kDa nuclear protein localized on the periphery of the nucleus was linked to dissociation of the nuclear lamina during apoptosis [58]. Recently, PAD3 was found to be necessary for apoptosis-inducing factor (AIF)-mediated apoptosis [43]. Overall, PAD activation (due to amplified calcium levels during the early stages of apoptosis) can facilitate apoptotic cell death by citrullinating several nuclear and cytoskeletal proteins that can cause structural changes resulting in the disintegration of secondary and tertiary protein structures [6, 18, 19].…”

Expression of peptidylarginine deiminase 4 in an alkali injury model of retinal gliosis

“…PAD2 is expressed in the retina and optic nerve of patients with glaucoma [22] and increased in age-related macular degeneration [24]. PAD3 is expressed in human neural stem cells [25], and PAD4 is found in the brains of Alzheimer’s [26] and multiple sclerosis [27] patients. However, it remained unknown which PAD enzyme is overactive in injury-induced gliosis.…”