Complex Alternative Cytoplasmic Protein Isoforms of the Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen 1 Generated through Noncanonical Translation Initiation

Abstract

bKaposi's sarcoma-associated herpesvirus (KSHV) latency associated-nuclear antigen 1 (LANA1) protein is constitutively expressed in all KSHV-infected cells, as well as in all forms of KSHV-associated malignancies. LANA1 is a multifunctional KSHV oncoprotein containing multiple repeat sequences that is important for viral episome maintenance and the regulation of cellular and viral gene expression. We characterize here multiple LANA1 isoforms and show that ϳ50% of LANA1 is naturally generated as N-terminally truncated shoulder proteins that are detected on SDS-PAGE as faster-migrating shoulder bands designated LANA1 S . Higher-molecular-weight LANA1 S isoforms initiate downstream at noncanonical sites within the N-terminal region, whereas lower-molecular-weight LANA1 S isoforms initiate downstream within the central repeat 1 domain. LANA1 S proteins lack an N-terminal nuclear localization signal motif, and some isoforms differ from full-length, canonical LANA1 by localizing to perinuclear and cytoplasmic sites. Although LANA1 has until now been assumed to be solely active in the nucleus, this finding indicates that this major KSHV oncoprotein may have cytoplasmic activities as well. KSHV overcomes its limited genetic coding capacity by generating alternatively initiated protein isoforms that may have distinct biological functions.

Cytoplasmic isoforms of Kaposi sarcoma herpesvirus LANA recruit and antagonize the innate immune DNA sensor cGAS

“…In contrast, the function of the recently discovered cytoplasmic isoforms of LANA, generated by noncanonical translation initiation within the LANA NTD (35,56) or possibly by caspase cleavage (24), has not been explored to date. In this study, we report that LANA, especially its cytoplasmic isoforms, antagonizes the function of cGAS, hereby promoting lytic replication after lytic cycle induction.…”

“…LANA is known to be a nuclear protein forming nuclear speckles in KSHV-infected cells. A recent study reported the existence of cytoplasmic isoforms of LANA, which are mainly generated by noncanonical internal translation initiation sites localized within the LANA N-terminal domain (35). Another recent report suggested the existence of N-terminally truncated LANA forms resulting from caspase cleavage (24).…”

“…In addition, a recent publication reported that lower-molecular-weight LANA isoforms can be generated by the use of noncanonical internal translation initiation sites within the N-terminal domain and are localized to the cytoplasm, because they lack a nuclear localization signal (35). The generation of LANA isoforms lacking part of the N-terminal domain by caspase cleavage has also been recently reported (24).…”

Human DNA tumor viruses generate alternative reading frame proteins through repeat sequence recoding

“…In fact, the names of both proteins specify, and restrictively imply, a nuclear localization. Recently, we have shown that cytoplasmic forms of truncated LANA1 exist (16). In this report, our data demonstrate that the major EBV and KSHV oncogenes generate additional, highly divergent frameshifted protein products.…”

“…The shorter form of the doublet is due to an alternative C-terminal polyadenylation site (15). More recently, even faster migrating isoforms have been characterized to result from in-frame, internal translation initiation at sites in the N-terminal and CR1 regions (16). All of these known isoforms have the same amino acid sequence as canonical LANA1 and differ only in being N-or C-terminally truncated.…”