Novel transcription regulatory sequences and factors of the immune evasion protein ICP47 (US12) of herpes simplex viruses

Abstract

Background: Herpes simplex virus (HSV) can cause encephalitis. Its infected cell polypeptide 47 (ICP47), encoded by immediate-early gene US12, promotes immune escape. ICP47 was modified in the clinically approved oncolytic HSV (oHSV) T-Vec. However, transcription regulatory sequence (TRS) and transcription regulatory factor (TRF) of HSV US12 are seldom reported.

Methods: Previously, our laboratory isolated a new HSV strain named HSV-1-LXMW from a male patient with oral herpes in Beijing, China. Firstly, the genetic tree was used to analyze its genetic relationship. The US12 TRS and TRF in HSV-1-LXMW were found by using predictive software. Secondly, the further verification by the multi-sequence comparative analysis shown that the upstream DNA sequence of HSV US12 gene contained the conserved region. Finally, the results of literature search shown that the expression of transcription factors was related to the tissue affinity of HSV-1 and HSV-2, so as to increase the new understanding of the transcriptional regulation of HSV biology and oncolytic virus (OVs) therapy.

Results: Here we reported the transcriptional regulation region sequence of our new HSV-1-LXMW, and its close relationship with HSV-1-CR38 and HSV-1-17. Importantly we identified eight different kinds of novel TRSs and TRFs of HSV US12 for the first time, and found they are conserved among HSV-1 (c-Rel, Elk-1, Pax-4), HSV-2 (Oct-1, CF2-II, E74A, StuAp) or both HSVs (HNF-4). The TRFs c-Rel and Oct-1 are biologically functional respectively in immune escape and viral replication during HSV infection.

Conclusions: Our findings have important implication to HSV biology, infection, immunity and oHSVs.

Keywords: HSV-1; HSV-2; ICP47; Transcriptional regulation factor (TRF); Transcriptional regulation sequence (TRS); US12.

Fig. 1

Transcription regulatory regions of US12. a. Schematic of the HSV-1 genome showing the regions of US12. The HSV-1 genome consists of long and short unique regions (UL and US) each bounded by terminal (T) and internal (I) repeat regions (RL and RS). b. The DNA sequence of the US12 gene is marked in green and the transcription regulatory regions of US12 is marked in blue

Fig. 2

Phylogenetic analysis of HSV-1-LXMW together with 11 other HSV strains. Evolutionary analyses were conducted in MEGA7. The evolutionary history was inferred by using the Maximum Likelihood method based on the Tamura-Nei model. The bootstrap consensus tree inferred from 1000 replicates is taken to represent the evolutionary history of the taxa analyzed. Branches corresponding to partitions reproduced in less than 50% bootstrap replicates are collapsed

Fig. 3

The US12 TRSs and TRFs in HSVs. TRFs are represented in different colors, and the number represents the specific location of their binding TRFs

Fig. 4

a. The US12 TRSs and TRFs are conserved among HSV-1 strains. Red lines indicate the conserved regions 1–14. The TRSs and TRFs are shown in colored boxes. b The US12 TRSs and TRFs are identical among HSV-2 strains. Red lines indicate the conserved regions 1–17. The TRSs and TRFs are shown in colored boxes

Fig. 5

ICP47 function during HSV infection. a Intracellular antigenic peptides, mainly generated by the proteasome, are transported into the endoplasmic reticulum by the TAP and then loaded onto the nascent MHC I molecules, which are exported to the cell surface and present peptides to the immune cells. Cytotoxic T lymphocytes recognize and kill the infected cells by granzyme and perforin. b ICP47 can preclude peptide binding and traps TAP in an inward-facing conformation. Binding of ICP47 stabilizes the inward-facing conformation, and thus prevents TAP from transitioning to an outward-facing state, resulting in the emergence of empty carrier MHC I molecules. Therefore, CD8⁺ T cells could not recognize them, and HSVs could avoid the immune responses. We hypothesize that c-Rel bind to its US12 TRS, and enhance US12 (ICP47) expression, leading to HSV-1 immune evasion and HSV-1 encephalitis