The HSV-1 ICP4 Transcriptional Auto-Repression Circuit Functions as a Transcriptional "Accelerator" Circuit.

Sonali Chaturvedi,Ruth Engel,Leor Weinberger

doi:10.3389/fcimb.2020.00265

Abstract

Herpes simplex virus-1 (HSV-1) is a significant human pathogen. Upon infection, HSV-1 expresses its immediate early (IE) genes, and the IE transcription factor ICP4 (infectious cell protein-4) plays a pivotal role in initiating the downstream gene-expression cascade. Using live-cell time-lapse fluorescence microscopy, flow cytometry, qPCR, and chromatin immunoprecipitation, we quantitatively monitored the expression of ICP4 in individual cells after infection. We find that extrinsic stimuli can accelerate ICP4 kinetics without increasing ICP4 protein or mRNA levels. The accelerated ICP4 kinetics—despite unchanged steady-state ICP4 protein or mRNA level—correlate with increased HSV-1 replicative fitness. Hence, the kinetics of ICP4 functionally mirror the kinetics of the human herpesvirus cytomegalovirus IE2 “accelerator” circuit, indicating that IE accelerator circuitry is shared among the alpha and beta herpesviruses. We speculate that this circuit motif is a common evolutionary countermeasure to throttle IE expression and thereby minimize the inherent cytotoxicity of these obligate viral transactivators.

Highlights

As one of the largest families of DNA viruses, Herpesviridae infections are prevalent among the world’s population
To determine whether acceleration without amplification is a conserved regulatory mechanism across herpesviruses, we examined if immediate early (IE) genes in Herpes simplex virus-1 (HSV-1) exhibit similar property as IE2 in Human cytomegalovirus (HCMV)
For RNA extraction followed by qPCR, cells were pretreated with hexamethylene bisacetamide (HMBA) or DMSO for 24 h followed by infection with HSV-1 [MOI = 0.05], harvested 5, 9, 13, and 17 h post infection from three biological replicates, and reversetranscription qPCR was performed as described previously (Vardi et al, 2018)

Summary

INTRODUCTION

As one of the largest families of DNA viruses, Herpesviridae infections are prevalent among the world’s population. We previously reported that HCMV encodes a transcriptional “accelerator” circuit that autoregulates expression of its master transcriptional regulator IE2 to prevent IE protein levels from reaching a cytotoxic threshold (Teng et al, 2012; Vardi et al, 2018). This accelerator circuit is in essence a highly nonlinear, self-cooperative transcriptional negative-feedback circuit wherein IE2 protein binds as a high-order homomultimer to a specific site on its own promoter to repress transcription (Figure 1A). Despite these similarities to HCMV IE2, the mechanism and kinetics of ICP4 autorepression were undetermined

MATERIALS AND METHODS

RESULTS AND DISCUSSION

DATA AVAILABILITY STATEMENT