Esophageal Cancer Related Gene-4 Is a Choroid Plexus-Derived Injury Response Gene: Evidence for a Biphasic Response in Early and Late Brain Injury

Abstract

By virtue of its ability to regulate the composition of cerebrospinal fluid (CSF), the choroid plexus (CP) is ideally suited to instigate a rapid response to traumatic brain injury (TBI) by producing growth regulatory proteins. For example, Esophageal Cancer Related Gene-4 (Ecrg4) is a tumor suppressor gene that encodes a hormone-like peptide called augurin that is present in large concentrations in CP epithelia (CPe). Because augurin is thought to regulate senescence, neuroprogenitor cell growth and differentiation in the CNS, we evaluated the kinetics of Ecrg4 expression and augurin immunoreactivity in CPe after CNS injury. Adult rats were injured with a penetrating cortical lesion and alterations in augurin immunoreactivity were examined by immunohistochemistry. Ecrg4 gene expression was characterized by in situ hybridization. Cell surface augurin was identified histologically by confocal microscopy and biochemically by sub-cellular fractionation. Both Ecrg4 gene expression and augurin protein levels were decreased 24–72 hrs post-injury but restored to uninjured levels by day 7 post-injury. Protein staining in the supraoptic nucleus of the hypothalamus, used as a control brain region, did not show a decrease of auguin immunoreactivity. Ecrg4 gene expression localized to CPe cells, and augurin protein to the CPe ventricular face. Extracellular cell surface tethering of 14 kDa augurin was confirmed by cell surface fractionation of primary human CPe cells in vitro while a 6–8 kDa fragment of augurin was detected in conditioned media, indicating release from the cell surface by proteolytic processing. In rat CSF however, 14 kDa augurin was detected. We hypothesize the initial release and proteolytic processing of augurin participates in the activation phase of injury while sustained Ecrg4 down-regulation is dysinhibitory during the proliferative phase. Accordingly, augurin would play a constitutive inhibitory function in normal CNS while down regulation of Ecrg4 gene expression in injury, like in cancer, dysinhibits proliferation.

Pulmonary preconditioning, injury, and inflammation modulate expression of the candidate tumor suppressor geneECRG4in lung

“…Likewise, we performed qRT-PCR and demonstrated elevated ECRG4 lung expression (lane 1) in comparison to heart (lane 2) and skin (lane 3) (Figure 1B). Immunoblotting of mouse lung with the anti-ECRG4 antibody [27, 46] revealed the presence of a single 14 kDa peptide that corresponded to the molecular weight of the predicted precursor product, amino acids 31–148, of ECRG4 (Figure 1C). …”

“…Several recent studies suggest that ECRG4 plays a role in injury homeostasis because its downregulation after injury/inflammation allows for dysinhibition of cell proliferation in vivo [27, 29, 46, 50, 51]. Its epigenetic regulation, however, suggests that the return of basal gene expression after injury could depend on epigenetic factors determined by environment, diet, and age.…”

Soluble purified recombinant C2ORF40 protein inhibits tumor cell growth in vivo by decreasing telomerase activity in esophageal squamous cell carcinoma

“…The C2ORF40 gene is important in processes associated with physiological functional regulation, including inflammation, injury, senescence, the neuroendocrines environment, differentiation and apoptosis (5)(6)(7)(8)(9)(10)(11)(12)(13). Notably, previous studies have indicated that C2ORF40 is a candidate tumor suppressor gene associated with prognosis in a variety of tumors (4,(14)(15)(16)(17)(18)(19)(20)(21)(22).…”

Monitoring Neutrophil-Expressed Cell Surface Esophageal Cancer Related Gene-4 after Severe Burn Injury

“…We have focused on the potential for esophageal cancer related gene-4 (ECRG4) to serve as a sentinel factor on immune cells that may regulate the immune set point based on its expression on the surface of leukocytes, its release from the cell surface after inflammatory stimuli, and its epigenetic regulation that may explain differences in Ecrg4 expression between individuals [6][7][8][9]. Esophageal cancer related gene-4 encodes a 148 amino acid precursor protein that is tethered to the cell surface but shed on activation [6,10].…”

“…Esophageal cancer related gene-4 encodes a 148 amino acid precursor protein that is tethered to the cell surface but shed on activation [6,10]. We have demonstrated previously that ECRG4 is present on the surface of polymorphonuclear leukocytes (PMNs) and, on ex vivo activation by agonists such as lipopolysaccharide (LPS), ECRG4 is released and can be detected by immunoblotting and proteomic analyses of biologic fluids including conditioned media, cerebrospinal fluid, and serum [6,7,[10][11][12]. We have also demonstrated that ECRG4 interacts with the human innate immunity receptor complex (TLR4-CD14-MD2) supporting further an important role in responding to inflammatory insults such as severe injury [13].…”