Mor_2008_Chem.Biol.Interact_175_11

Tumorogenic transformation is a multifaceted cellular process involving combinatorial protein-protein interactions that modulate different cellular functions. Here, we report apparent involvement in two independent tumorogenic processes by distinct partner protein interactions of the stress-induced acetylcholinesterase AChE-R and N-AChE-R variants. Human testicular tumors showed elevated levels of N-terminally extended N-AChE-R compared with healthy tissue, indicating alternate promoter usage in the transformed cells. Two-hybrid screens demonstrate that the C-terminus common to both N-AChE-R and AChE-R interacts either with the glycolytic enzyme enolase or with the scaffold protein RACK1. In vitro, the AChE-R C-terminal peptide ARP elevated enolase's activity by 12%, suggesting physiological relevance for this interaction. Correspondingly, CHO cells expressing either human AChE-R or N-AChE-R but not AChE-S showed a 25% increase in cellular ATP levels, indicating metabolic significance for this upregulation of enolase activity. ATP levels could be reduced by AChE-targeted siRNA in CHO cells expressing AChE-R but not AChE-S, attributing this elevation to the AChE-R C-terminus. Additionally, transfected CHO cells expressing AChE-R but not N-AChE-R showed resistance to up to 60 microM of the common chemotherapeutic agent, cis-platinum, indicating AChE-R involvement in another molecular pathway. cis-Platinum elevates the expression of the apoptosis-regulator p53-like protein, p73, which is inactivated by interaction with the scaffold protein RACK1. In co-transfected cells, AChE-R competed with endogenous RACK1 for p73 interaction. Moreover, AChE-R-transfected CHO cells presented higher levels than control cells of the pro-apoptotic TAp73 as well as the anti-apoptotic dominant negative DeltaNp73 protein, leading to an overall decrease in the proportion of pro-apoptotic p73. Together, these findings are compatible with the hypothesis that in cancer cells, both AChE-R and N-AChE-R elevate cellular ATP levels and that AChE-R modifies p73 gene expression by facilitating two independent cellular pathways, thus conferring both a selective metabolic advantage and a genotoxic resistance.