Abstract
Human herpesvirus 6A and 6B frequently acquires latency. HHV-6 activation has been associated with various human diseases. Germ line inheritance of chromosomally integrated HHV-6 makes viral DNA-based analysis difficult for determination of early stages of viral activation. We characterized early stages of HHV-6 activation using high throughput transcriptomics studies and applied the results to understand virus activation under clinical conditions. Using a latent HHV-6A cell culture model in U2OS cells, we identified an early stage of viral reactivation, which we define as transactivation that is marked by transcription of several viral small non-coding RNAs (sncRNAs) in the absence of detectable increase in viral replication and proteome. Using deep sequencing approaches, we detected previously known as well as a new viral sncRNAs that characterized viral transactivation and differentiated it from latency. Here we show changes in human transcriptome upon viral transactivation that reflect multiple alterations in mitochondria-associated pathways, which was supported by observation of increased mitochondrial fragmentation in virus reactivated cells. Furthermore, we present here a unique clinical case of DIHS/DRESS associated death where HHV-6 sncRNA-U14 was abundantly detected throughout the body of the patient in the presence of low viral DNA. In this study, we have identified a unique and early stage of viral activation that is characterized by abundant transcription of viral sncRNAs, which can serve as an ideal biomarker under clinical conditions.
Highlights
Betaherpesviruses like human herpesvirus 6A (HHV-6A), HHV-6B and HHV-7 integrate into subtelomeric ends of human chromosomes and acquire latency.[1,2] It is speculated that homologous recombination between telomeric repeat sequences allow viral integration as genetically altered herpesviruses lacking telomeric repeat sequences fail to integrate efficiently into human telomeres.[3,4] Several HHV-6 latency associated genes have been described but functional significance of such gene products are not well understood
Integrated viral genome is often lost during initial stages of viral reactivation Understanding initial stages of HHV-6 reactivation strongly depends upon a robust cell line carrying homogeneous latent viral infections
To generate such a cell line, we utilized bacterial artificial chromosomes (BAC) derived[19] HHV-6A viral particles carrying coding potentials for either recombinant green fluorescence protein (GFP)[20] or red fluorescence protein (RFP) as these reporter cells are ideal for cell sorting and further selection of
Summary
HHV-6 encoded small non-coding RNAs define an intermediate and early stage in viral reactivation. Using a latent HHV-6A cell culture model in U2OS cells, we identified an early stage of viral reactivation, which we define as transactivation that is marked by transcription of several viral small noncoding RNAs (sncRNAs) in the absence of detectable increase in viral replication and proteome. We show changes in human transcriptome upon viral transactivation that reflect multiple alterations in mitochondria-associated pathways, which was supported by observation of increased mitochondrial fragmentation in virus reactivated cells. We present here a unique clinical case of DIHS/DRESS associated death where HHV-6 sncRNA-U14 was abundantly detected throughout the body of the patient in the presence of low viral DNA. We have identified a unique and early stage of viral activation that is characterized by abundant transcription of viral sncRNAs, which can serve as an ideal biomarker under clinical conditions
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