Kaposi's Sarcoma-Associated Herpesvirus Infection Induces the Expression of Neuroendocrine Genes in Endothelial Cells

Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically associated with endothelial Kaposi's sarcoma (KS) in immunocompromised individuals. KS lesion cells exhibit many similarities to neuroendocrine (NE) cancers, such as highly vascular and red/purple tumor lesions, spindle-shaped cells, an insignificant role for classic oncogenes in tumor development, the release of bioactive amines, and indolent growth of the tumors. However, the mechanistic basis for the similarity of KS lesion endothelial cells to neuroendocrine tumors remains unknown. Next-generation sequencing and bioinformatics analysis in the present study demonstrate that endothelial cells latently infected with KSHV express several neuronal and NE genes. De novo infection of primary dermal endothelial cells with live and UV-inactivated KSHV demonstrated that viral gene expression is responsible for the upregulation of five selected NE genes (adrenomedullin 2 [ADM2], histamine receptor H1 [HRH1], neuron-specific enolase [NSE] [ENO2], neuronal protein gene product 9.5 [PGP9.5], and somatostatin receptor 1 [SSTR1]). Immunofluorescence and immunohistochemistry examinations demonstrated the robust expression of the NE genes HRH1 and NSE/ENO2 in KSHV-infected KS tissue samples and KS visceral tissue microarrays. Further analysis demonstrated that KSHV latent open reading frame K12 (ORFK12) gene (kaposin A)-mediated decreased host REST/NRSF (RE1-silencing transcription factor/neuron-restrictive silencer factor) protein, a neuronal gene transcription repressor protein, is responsible for NE gene expression in infected endothelial cells. The NE gene expression observed in KSHV-infected cells was recapitulated in uninfected endothelial cells by the exogenous expression of ORFK12 and by the treatment of cells with the REST inhibitor X5050. When the neuroactive ligand-activating receptor HRH1 and inhibitory SSTR1 were knocked out by CRISPR, HRH1 knockout (KO) significantly inhibited cell proliferation, while SSTR1 KO induced cell proliferation, thus suggesting that HRH1 and SSTR1 probably counteract each other in regulating KSHV-infected endothelial cell proliferation. These results demonstrate that the similarity of KS lesion cells to neuroendocrine tumors is probably a result of KSHV infection-induced transformation of nonneuronal endothelial cells into cells with neuroendocrine features. These studies suggest a potential role of neuroendocrine pathway genes in the pathobiological characteristics of KSHV-infected endothelial cells, including a potential mechanism of escape from the host immune system by the expression of immunologically privileged neuronal-site NE genes, and NE genes could potentially serve as markers for KSHV-infected KS lesion endothelial cells as well as novel therapeutic targets to control KS lesions.IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) manipulates several cellular pathways for its survival advantage during its latency in the infected human host. Here, we demonstrate that KSHV infection upregulates the expression of genes related to neuronal and neuroendocrine (NE) functions that are characteristic of NE tumors, both in vitro and in KS patient tissues and the heterogeneity of neuroendocrine receptors having opposing roles in KSHV-infected cell proliferation. Induction of NE genes by KSHV could also provide a potential survival advantage, as the expression of proteins at immunologically privileged sites such as neurons on endothelial cells may be an avenue to escape host immune surveillance functions. The NE gene products identified here could serve as markers for KSHV-infected cells and could potentially serve as therapeutic targets to combat KSHV-associated KS.