Domain Interaction Studies of Herpes Simplex Virus 1 Tegument Protein UL16 Reveal Its Interaction with Mitochondria

Abstract

The UL16 tegument protein of herpes simplex virus 1 (HSV-1) is conserved among all herpesviruses and plays many roles during replication. This protein has an N-terminal domain (NTD) that has been shown to bind to several viral proteins, including UL11, VP22, and glycoprotein E, and these interactions are negatively regulated by a C-terminal domain (CTD). Thus, in pairwise transfections, UL16 binding is enabled only when the CTD is absent or altered. Based on these results, we hypothesized that direct interactions occur between the NTD and the CTD. Here we report that the separated and coexpressed functional domains of UL16 are mutually responsive to each other in transfected cells and form complexes that are stable enough to be captured in coimmunoprecipitation assays. Moreover, we found that the CTD can associate with itself. To our surprise, the CTD was also found to contain a novel and intrinsic ability to localize to specific spots on mitochondria in transfected cells. Subsequent analyses of HSV-infected cells by immunogold electron microscopy and live-cell confocal imaging revealed a population of UL16 that does not merely accumulate on mitochondria but in fact makes dynamic contacts with these organelles in a time-dependent manner. These findings suggest that the domain interactions of UL16 serve to regulate not just the interaction of this tegument protein with its viral binding partners but also its interactions with mitochondria. The purpose of this novel interaction remains to be determined. IMPORTANCEThe HSV-1-encoded tegument protein UL16 is involved in multiple events of the virus replication cycle, ranging from virus assembly to cell-cell spread of the virus, and hence it can serve as an important drug target. Unfortunately, a lack of both structural and functional information limits our understanding of this protein. The discovery of domain interactions within UL16 and the novel ability of UL16 to interact with mitochondria in HSV-infected cells lays a foundational framework for future investigations aimed at deciphering the structure and function of not just UL16 of HSV-1 but also its homologs in other herpesviruses.KEYWORDS domains, HSV-1, herpes simplex virus, mitochondria, tegument, UL16 H erpes simplex virus 1 (HSV-1) contains a 373-amino-acid tegument protein, UL16 (Fig. 1), which is conserved among all families of herpesviruses (1-15). Encoded by the late-expressing 16th open reading frame in the unique long segment of the viral genome, UL16 initially localizes to both the nucleus and the cytoplasm of the infected cell but later accumulates in the cytoplasm (4, 16). There, a population of UL16 is found to be stably associated with an unknown partner on cytoplasmic capsids (16,17), where it plays a role in envelopment by providing bridging functions via its interaction with at least three viral membrane proteins: UL11, glycoprotein E (gE), and VP22 (12, 18-43). Thus, UL16-null mutants produce only 1/10 the number of wild-type virions (1, 12), and

The longest mitochondrial protein in metazoans is encoded by the male-transmitted mitogenome of the bivalve Scrobicularia plana

“…Such ORFs were not found in S. plana, but a conserved domain was identified in the insertion, i.e. PHA03247 large tegument protein UL36, which is conserved among herpesviruses and plays many roles in viral replication [45]. Interestingly, this result is in line with a previous hypothesis that viral selfish elements may have colonized the male mitogenome in bivalves promoting its segregation into primordial germ cells and allowing its transmission to next generations [8,46].…”

Characteristics of herpes simplex virus infection and pathogenesis suggest a strategy for vaccine development

“…Other studies have also suggested that VP22, which is encoded by the ul49 gene, is capable of interacting with cellular cGAS and inhibiting its enzymatic activity, as VP22 functions in the viral structural network . The tegument proteins encoded by ul16 and ul46 also show capacities to interact with cellular mitochondria and p85, growth factor receptor bound protein 2 (Grb2), and shc of the Src‐family kinases, respectively, as part of their roles in the viral structural network . These data describe a specific context for HSV infection, in which the viral strategy is to present only surface glycoproteins to the immune system, as most of the pathogenic viral molecules work within infected cells and can avoid monitoring by the innate and adaptive immune systems through various forms of immune evasion.…”

Differential Requirements for gE, gI, and UL16 among Herpes Simplex Virus 1 Syncytial Variants Suggest Unique Modes of Dysregulating the Mechanism of Cell-to-Cell Spread

“…Although the explanation for this unexpected phenotype remains to be elucidated, there are two properties of UL16 that raise the possibility that the innate cellular immune response might be enhanced when this viral protein is absent. First, UL16 has been shown to traffic to mitochondria during infection (70), and this is a known site for innate immune signaling (71). Second, in the absence of tegument protein UL37, packaging of UL16 into virions is reduced, suggesting a direct or indirect interaction between these two proteins.…”