In-Depth Temporal Transcriptome Profiling of an Alphaherpesvirus Using Nanopore Sequencing

Dóra Tombácz; Balázs Kakuk; Gábor Torma; Zsolt Csabai; Gábor Gulyás; Vivien Tamás; Zoltán Zádori; Victoria A Jefferson; Florencia Meyer; Zsolt Boldogkői

doi:10.3390/v14061289

In-Depth Temporal Transcriptome Profiling of an Alphaherpesvirus Using Nanopore Sequencing

Viruses. 2022 Jun 13;14(6):1289. doi: 10.3390/v14061289.

Affiliations

¹ Department of Medical Biology, Albert Szent-Györgyi Medical School, University of Szeged, Somogyi u. 4, 6720 Szeged, Hungary.
² Institute for Veterinary Medical Research, Centre for Agricultural Research, Hungária krt. 21, 1143 Budapest, Hungary.
³ Department of Biochemistry & Molecular Biology, Entomology & Plant Pathology, Mississippi State University, 408 Dorman P.O. Box 9655, 32 Creelman St., Starkville, MS 39762, USA.

Abstract

In this work, a long-read sequencing (LRS) technique based on the Oxford Nanopore Technology MinION platform was used for quantifying and kinetic characterization of the poly(A) fraction of bovine alphaherpesvirus type 1 (BoHV-1) lytic transcriptome across a 12-h infection period. Amplification-based LRS techniques frequently generate artefactual transcription reads and are biased towards the production of shorter amplicons. To avoid these undesired effects, we applied direct cDNA sequencing, an amplification-free technique. Here, we show that a single promoter can produce multiple transcription start sites whose distribution patterns differ among the viral genes but are similar in the same gene at different timepoints. Our investigations revealed that the circ gene is expressed with immediate-early (IE) kinetics by utilizing a special mechanism based on the use of the promoter of another IE gene (bicp4) for the transcriptional control. Furthermore, we detected an overlap between the initiation of DNA replication and the transcription from the bicp22 gene, which suggests an interaction between the two molecular machineries. This study developed a generally applicable LRS-based method for the time-course characterization of transcriptomes of any organism.

Keywords: bovine alphaherpesvirus type 1; direct cDNA sequencing; herpesviruses; long-read sequencing; nanopore sequencing; transcript isoforms; transcription end site; transcription start site; transcriptome.

Publication types

Research Support, U.S. Gov't, Non-P.H.S.
Research Support, Non-U.S. Gov't

MeSH terms

Gene Expression Profiling / methods
Herpesvirus 1, Bovine* / genetics
High-Throughput Nucleotide Sequencing / methods
Nanopore Sequencing*
Transcription Initiation Site
Transcriptome

Grants and funding

This study was supported by the National Research, Development and Innovation Office grant: K 128247 to ZB and FK 128252 to DT, University of Szeged. The study was also supported by USDA-AFRI award 2020-67016-31345 to FM. The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.