KSHV uses viral IL6 to expand infected immunosuppressive macrophages (RNA-seq)

Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic double-stranded DNA virus and the etiologic agent of Kaposi's sarcoma and hyperinflammatory lymphoproliferative disorders. Understanding the mechanism by which KSHV increases infected cell populations is crucial for curing KSHV-associated diseases. Here, we demonstrate that KSHV preferentially infects CD14+ monocytes and sustains viral replication through the viral interleukin-6 (vIL-6)-mediated activation of STAT1 and 3. Using vIL6-sufficient and vIL6-deficient recombinant KSHV, we demonstrated that vIL6 plays a critical role in promoting the proliferation and differentiation of KSHV-infected monocytes into macrophages. The macrophages derived from vIL6-sufficient (wild type) KSHV infection showed a distinct transcriptional profile of elevated IFN-pathway activation with immune suppression and were compromised in T-cell stimulation function compared to those from vIL6-deficient KSHV infection or uninfected control. These results highlight a clever strategy, in which KSHV utilizes vIL6 to secure its initial viral pool by expanding infected dysfunctional macrophages. This mechanism also facilitates KSHV to escape from host immune surveillance and to establish a lifelong infection. Overall design: The goal of this study was to define the role of KSHV vIL6 in monocyte activation and differentiation by using de novo infection of PBMCs with vIL6-sufficient and vIL6-deficient recombinant KSHV followed by transcription profiling with RNA-seq. To this end, recombinant KSHV lacking vIL6 expression was prepared by inserting stop codons into the vIL6 gene (vIL6STOP; K2_neg) using the KSHV bacterial artificial chromosome (BAC) clone BAC16. A revertant KSHV with intact vIL6 expression (vIL6REV; K2_pos) was also generated by changing the sequence back to the original wild type sequence with BAC recombination. Duplicate PBMC cultures were each infected with vIL6REV, vIL6STOP, or mock-infected (PBS) and cells were harvested at 7 days post-infection for isolation of total RNA and RNA-seq analysis. Comparative gene expression profiling analysis of the RNA-seq data was then performed to identify the differentially expressed genes (DEGs) in monocytes infected with vIL6-sufficient or vIL6-deficient KSHV.