Role of Calpain in the Formation of Human Papillomavirus Type 16 E1∧E4 Amyloid Fibers and Reorganization of the Keratin Network

The human papillomavirus (HPV) type 16 E1 boolean AND E4 (16E1 boolean AND E4) protein is expressed in the middle to upper layers of infected epithelium and has several roles within the virus life cycle. It is apparent that within the epithelium there are multiple species of 16E1 boolean AND E4 that differ in length and/or degree of phosphorylation and that some or all of these can associate with the cellular keratin networks, leading to network disruption. We show here that the cellular cysteine protease calpain cleaves the 16E1 boolean AND E4 protein after amino acid 17 to generate species that lack the N terminus. These C-terminal fragments are able to multimerize and form amyloid-like fibers. This can lead to accumulation of 16E1 boolean AND E4 and disruption of the normal dynamics of the keratin networks. The cleavage of E1 boolean AND E4 proteins by calpain may be a common strategy used by alpha-group viruses, since we show that cleavage of type 18 E1 boolean AND E4 in raft culture is also dependent on calpain. Interestingly, the cleavage of 16E1 boolean AND E4 by calpain appears to be highly regulated as differentiation of HPV genome-containing cells by methylcellulose is insufficient to induce cleavage. We hypothesize that this is important since it ensures that the formation of the amyloid fibers is not prematurely triggered in the lower layers and is restricted to the upper layers, where calpain is active and where disruption of the keratin networks may aid virus release.