Nuclear import pathways of human papillomavirus *E6 proteins
The E6 oncoprotein is of vital importance in the pathogenesis of human papillomavirus (HPV) induced genital disease. The E6 transforming protein is a key player in the development for both benign disease such as condylomata and malignant disease such as cervical carcinoma in those infected with HPV. Certain sub-types of HPV infections have been stratified as low-risk and high-risk by epidemiologic studies, correlating with benign and malignant disease respectively. Our work characterizes the nuclear import of high-risk HPV16 E6 and explores differences in the nuclear import of E6 in high and low risk sub-types of HPVs.
The HPV16 E6 oncoprotein is imported into the nucleus via multiple pathways. Nuclear import in digitonin-permeabilized HeLa cells occurred when HPV16 E6 interacted with adapter karyopherin (Kap) α2 and entered the nucleus via the classical Kap α2β1 mediated pathway. Moreover, HPV16 E6 also localized to the nucleus in the presence of Kap β1 by itself or with Kap β2 (transportin). Binding of RanGTP to these Kap βs inhibited their interactions with the HPV16 E6 NLS (Nuclear Localization Signal).
Significantly, the NLS mutant E6R124G had reduced nuclear import activity while the E6 NLS deletion mutant, lacking 121KKQR 124, did not import at all. Thus, the HPV16 E6 oncoprotein interacts via its C-terminal NLS with several import receptors and enters the nucleus using multiple pathways.
The key difference in the nuclear import of the E6 protein associated with high and low risk HPV is active transport in high-risk types versus passive diffusion in low-risk types. Both high-risk HPV18 and 16 E6 are actively imported into the nucleus in the presence of either Kap β1 or Kap β2, in contrast with HPV11 E6 which does not interact with any of these import receptors. In the presence of Wheat Germ Agglutinin (WGA, inhibits active import) both high-risk HPV18 and 16 E6 remain cytoplasmic and show clear nuclear rim staining, indicating docking at the nuclear pore complex, but arrest of further transport into the nucleus. In contrast, low-risk HPV11 E6 showed similar low levels of diffusion mediated nuclear accumulation in both the presence and absence of WGA.
0379: Cellular biology