Inhibition of Caspases Protects Mice from Radiation-induced Oral Mucositis and Abolishes the Cleavage of RNA-binding Protein HuR

Abstract

The oral mucosal epithelium is typically insulted during chemotherapy and ionizing radiation (IR) therapy and disposed to mucositis, which creates painful inflammation and ulceration in the oral cavity. Oral mucositis alters gene expression patterns, inhibits cellular growth, and initiates cell death in the oral epithelial compartments. Such alterations are governed by several different factors, including transcription factors, RNA-binding proteins, and microRNAs. IR-induced post-transcriptional regulation of RNA-binding proteins exists but is poorly studied in clinically relevant settings. We herein report that the RNA-binding protein human antigen R (HuR) undergoes cleavage modification by caspase-3 following IR-induced oral mucositis and subsequently promotes the expression of the pro-apoptotic factor BAX (Bcl-2-associated X protein), as well as cell death. Further analyses revealed that the HuR cleavage product-1 (HuR-CP1) directly associates and stabilizes the BAX mRNA and concurrently activates the apoptotic pathway. On the other hand, a noncleavable isoform of HuR promotes the clonogenic capacity of primary oral keratinocytes and decreases the effect of IR-induced cell death. Additionally, specific inhibition of caspase-3 by a compound, NSC321205, increases the clonogenic capacity of primary oral keratinocytes and causes increased basal layer cellularity, thickened mucosa, and elevated epithelial cell growth in the tongues of mice with oral mucositis. This protective effect of NSC321205 is mediated by a decrease in caspase-3 activity and the consequent inhibition of HuR cleavage, which reduces the expression of BAX in mice with IR-induced oral mucositis. Thus, we have identified a new molecular mechanism of HuR in the regulation of mRNA turnover and apoptosis in oral mucositis, and our data suggest that blocking the cleavage of HuR enhances cellular growth in the oral epithelial compartment.