Viral transcription-translation coupling in mammalian cells

Amiyaranjan Das¹, Julia Bartuli², Clemens Grimm², Utz Fischer², Gabriel Demo¹

¹Central European Institute of Technology, Masaryk University, Kamenice 5, 62500, Bohunice, Brno, Czech Republic

²Department of Biochemistry and Cancer Therapy Research Center (CTRC), Theodor Boveri-Institute, University of Würzburg, Am Hubland, 97074 Würzburg, Germany

amiyaranjan.das@ceitec.muni.cz

The coupling of transcription and translation (CTT) controls the gene regulation in bacteria. Recent cryo-electron microscopy (cryo-EM) studies showed the physical coupling of these two processes [1,2]. Interestingly, the double-strand DNA Vaccinia virus (VACV) performs the viral genome replication, transcription, translation, and assembly of virions in infected mammalian cells within discrete cytoplasmic foci called viral factories [3-5]. The initial rounds of viral transcription and translation during the early phase of infection occur inside the host cytoplasm. In intermediate and late-phase of infection the viral gene expression is carried out inside the viral factories in close association with host ribosomes [5].

Here, we seek to uncover the potential regulation mechanism of viral gene expression via viral-host CTT. The primary approach is to reconstitute the CTT in vitro and use single particle cryo-EM to uncover the detailed view of the structural architecture of the viral-host CTT. We aim to employ cryo-electron tomography and correlated light and electron microscopy (CLEM) to directly visualize the viral factories in a near-native state at a sub-nanometer resolution to confirm the existence of viral-host CTT directly in VACV-infected cells.

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2. C. Wang, V. Molodtsov, E. Firlar, JT. Kaelber, G. Blaha, M. Su, RH. Ebright, Structural basis of transcription-translation coupling. Science 369, 1359-1365. (2020).

3. J. A. den Boon, A. Diaz, P. Ahlquist, Cytoplasmic viral replication complexes. Cell Host Microbe 8, 77-85 (2010).

4. B. Moss, Regulation of vaccinia virus transcription. Annu Rev Biochem 59, 661-688 (1990).

5. G. C. Katsafanas, B. Moss, Colocalization of transcription and translation within cytoplasmic poxvirus factories coordinates viral expression and subjugates host functions. Cell Host Microbe 2, 221-228 (2007).

This study was supported by LL2008 project with financial support from MEYS CR as a part of the ERC CZ program (to G.D.).