Abstract

Kaposi's Sarcoma Associated Herpesvirus (KSHV) or Human Herpesvirus-8 (HHV-8) is the most recently identified human gamma-2 herpesvirus and has been implicated in Kaposi's Sarcoma (KS) and primary effusion lymphoma (PEL). At the right end of the genome KSHV encodes the complex kaposin locus, which consists of two distinct sets of 23 amino acid direct repeats, DR2 and DR1, followed by a short domain originally referred to as open reading frame (ORF) K12. Translational initiation at multiple alternative CUG and one AUG start codons causes expression of a gradient of kaposin molecules with varying length and targeting motifs from one single transcript. The aim of the present study was to investigate in detail the expression pattern of the kaposin locus and the cellular localization and function of kaposin protein isoforms expressed in the KSHV+ PEL cell line BCBL-1. The multitude of translational products from all three reading frames could be resolved and different isoforms assigned to distinct cellular compartments. Depending on the alternative start codon used, the DR1 repeats representing a functional effector domain are fused either to the DR2 repeats harboring a nuclear localization sequence (NLS), or to K12, which encodes a transmembrane domain. Expression of kaposin in the nucleus (kaposin B) causes an activation of the AP-1 transcription factor and cellular promoters. The observed AP-1 induction is dependent on nuclear localization of both DR2 and DR1 repeats, since substitution of DR2 with a SV-40 NLS was not sufficient to restore activation. Other kaposin isoforms which are found in the cytosol (kaposin E) or membrane-associated (kaposin D) failed to activate AP-1. If co-expressed, however, kaposin D and E were able to modulate the kaposin B-caused induction, presumably mediated by a direct interaction between DR2 and DR1. The results presented in this study indicate a novel autoregulatory mechanism based on bidirectional targeting of a viral protein to distinct subcellular compartments by expression from different start codons and reading frames. Supported by the complexity of the translational program and the conservation of the repeat regions, these findings imply that kaposin isoforms have important functions in the viral life cycle.