Characterization of the Kaposi's sarcoma-associated herpesvirus terminase complex component ORF29

Abstract

Kaposis sarcoma-associated herpesvirus (KSHV) belongs to the gammaherpesvirinae subfamily. In the lytic phase of herpesviruses, viral capsids are formed in the nucleus of the host cell and the replicated viral genome is packaged into capsids. The herpesviral genome is replicated as a precursor head-to-tail concatemer consisting of tandemly repeated genomic units, and each genomic unit is flanked by terminal repeats (TRs). The herpesvirus terminase complex packages one genomic unit into a capsid, by cleavage of the TRs in a precursor genome. Although the terminase complexes of alpha- and beta-herpesviruses are well characterized, in KSHV, the terminase complex is poorly understood. KSHV ORF7, ORF67.5, and ORF29 are thought to be components of the terminase complex. We previously reported that ORF7- or ORF67.5-deficient KSHV formed immature soccer ball-like capsids and failed to cleave the TRs, resulting in decreased virion production. Moreover, ORF7 interacted with ORF29 and ORF67.5. However, ORF29 and ORF67.5 did not interact with each other. Thus, although ORF7 and ORF67.5 are important for KSHV terminase function, the function of ORF29 remains largely unknown. Here, we constructed ORF29-deficient BAC16 and analyzed its virological properties. ORF29 was essential for virion production and TR cleavage. Numerous immature soccer ball-like capsids were observed in ORF29-deficient KSHV-harboring cells. The N-terminal region of ORF29 was important for its interaction with ORF7, although full-length ORF29 was required for effective complexation of the KSHV terminase. Furthermore, ORF29 preferentially interacted with itself rather than with ORF7. Thus, our data shows that ORF29 functions as a fundamental terminase component. IMPORTANCESince the role of ORF29 in the KSHV terminase complex remains unknown, we constructed ORF29-deficient KSHV. Our results demonstrated that ORF29 functions as a component of the KSHV terminase and is critical for capsid formation, TR cleavage, and terminase complexation. Moreover, ORF29 robustly interacted with itself. In HSV-1, a terminase complex containing UL15, UL28, and UL33 forms a trimer, and six trimers assemble into a hexameric ring. The HSV-1 genome passes through this ring and undergoes TR cleavage and genome packaging into a capsid. The self-interaction of ORF29 may be involved in the multimerization of a terminase complex or formation of the KSHV terminase ring. In addition, a novel KSHV protein, which we refer to as ORF29.5, was expressed during the lytic state. ORF29.5 mRNA shares the same stop codon with the ORF29 mRNA. Moreover, a part of the ORF29.5 coding region may overlap with the C-terminal ORF29 coding region.