Adenovirus replication – it's really all about the RNAs

Adenovirus transcription Human adenoviruses are non-enveloped icosahedral viruses with linear double stranded DNA genomes. The genome is transcribed from both strands and it is organized into several transcription units named mainly according to when they are expressed during the virus life cycle. Five early transcription units encode the E1A, E1B, E2, E3 and E4 proteins, two delayed early units encode the IVa2 and pIX proteins and there is one major late transcription unit (MLTU). The major functions of the early gene products are to force the host cell to enter the S phase in order to provide optimal conditions for viral DNA replication and for suppression of the host antiviral response. The major late gene products are the viral structural proteins which package the viral DNA into new virus particles.

Group C adenoviruses also encode two small RNAs, called virus-associated (VA) RNAI and VA RNAII. They are non-coding RNAs and transcribed by RNA polymerase III. Both VA RNAs are about 160 nucleotides long and GC rich. Expression of the VA RNAs begins during the early phase of infection and increases rapidly to a high level during the late phase. Inactivation of VA RNAI results in a 10–20 fold decrease in virus production, whereas deletion of VA RNAII alone has little impact on virus replication. The functional significance of VA RNAI is well documented whereas little is known about the function of VA RNAII. The primary function of VA RNAI appears to be to block the activity of RNA-dependent protein kinase (PKA), a double-strained RNA activated inhibitor of translation. VA RNAI also stabilizes ribosome-associated viral mRNAs resulting in enhanced levels of viral protein synthesis. In addition, VA RNAI binds efficiently to Exportin 5, interfering with the nuclear export of the cellular RNAi and miRNA precursors and Dicer processing. Finally, large amounts of VA RNA-derived small RNAs associate with RNA-induced silencing (RISC) complexes.

During the last decade, increasing numbers of small RNAs have been identified and characterized and it has become evident that the small RNAs are critical regulators of gene function (Bartel, 2004, Seto et al., 2007 and Zaratiegui et al., 2007). There are three main categories: short interfering RNAs (siRNAs which are ∼21 nt in length), microRNAs (miRNAs, ∼22 nt in length) and PIWI-interacting RNAs (piRNAs, ∼24–32 nt in length). Deep sequencing technologies combined with bioinformatic strategies have revolutionized the identification of rare small RNAs. This study examines the expression of adenovirus-encoded small RNAs at different times after infection using deep sequencing. Adenovirus-encoded small RNAs may thus constitute a front-line defense and be crucial for the survival of the virus.

 

Identification of adenovirus-encoded small RNAs by deep RNA sequencing. Virology. 06 May 2013. pii: S0042-6822(13)00200-6. doi: 10.1016/j.virol.2013.04.006
Using deep RNA sequencing, we have studied the expression of adenovirus-encoded small RNAs at different times after infection. Nineteen small RNAs which comprised more than 1% of the total pool of small RNAs at least one time point were identified. These small RNAs were between 25 and 35 nucleotides long and mapped in the region of the VA RNAI and RNAII genes. However, the overlap was incomplete and some contained a few extra nucleotides at the 3′ end. This finding together with the observation that some of the small RNAs were detected before VA RNA expression had started might indicate that they are derived from other precursors than VA RNAI and II. Interestingly, the small RNAs displayed different expression profiles during the course of the infection suggesting that they have different functions. An effort was made to identify their mRNA targets by using computer prediction and deep cDNA sequencing. The most significant targets for the earliest small RNAs were genes involved in signaling pathways.

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