Abstract
ABSTRACTInfection by most DNA viruses activates a cellular DNA damage response (DDR), which may be to the detriment or advantage of the virus. In the case of adenoviruses, they neutralize antiviral effects of DDR activation by targeting a number of proteins for rapid proteasome-mediated degradation. We have now identified a novel DDR protein, tankyrase 1 binding protein 1 (TNKS1BP1) (also known as Tab182), which is degraded during infection by adenovirus serotype 5 and adenovirus serotype 12. In both cases, degradation requires the action of the early region 1B55K (E1B55K) and early region 4 open reading frame 6 (E4orf6) viral proteins and is mediated through the proteasome by the action of cullin-based cellular E3 ligases. The degradation of Tab182 appears to be serotype specific, as the protein remains relatively stable following infection with adenovirus serotypes 4, 7, 9, and 11. We have gone on to confirm that Tab182 is an integral component of the CNOT complex, which has transcriptional regulatory, deadenylation, and E3 ligase activities. The levels of at least 2 other members of the complex (CNOT3 and CNOT7) are also reduced during adenovirus infection, whereas the levels of CNOT4 and CNOT1 remain stable. The depletion of Tab182 with small interfering RNA (siRNA) enhances the expression of early region 1A proteins (E1As) to a limited extent during adenovirus infection, but the depletion of CNOT1 is particularly advantageous to the virus and results in a marked increase in the expression of adenovirus early proteins. In addition, the depletion of Tab182 and CNOT1 results in a limited increase in the viral DNA level during infection. We conclude that the cellular CNOT complex is a previously unidentified major target for adenoviruses during infection.IMPORTANCE Adenoviruses target a number of cellular proteins involved in the DNA damage response for rapid degradation. We have now shown that Tab182, which we have confirmed to be an integral component of the mammalian CNOT complex, is degraded following infection by adenovirus serotypes 5 and 12. This requires the viral E1B55K and E4orf6 proteins and is mediated by cullin-based E3 ligases and the proteasome. In addition to Tab182, the levels of other CNOT proteins are also reduced during adenovirus infection. Thus, CNOT3 and CNOT7, for example, are degraded, whereas CNOT4 and CNOT1 are not. The siRNA-mediated depletion of components of the complex enhances the expression of adenovirus early proteins and increases the concentration of viral DNA produced during infection. This study highlights a novel protein complex, CNOT, which is targeted for adenovirus-mediated protein degradation. To our knowledge, this is the first time that the CNOT complex has been identified as an adenoviral target.
Highlights
Infection by most DNA viruses activates a cellular DNA damage response (DDR), which may be to the detriment or advantage of the virus
It was previously suggested that Tab182 may have a role in the DDR based on the observation that the protein has multiple potential ataxia telangiectasia mutated (ATM)/ATR phosphorylation sites (SQ/TQ) and is phosphorylated following exposure to ionizing radiation (IR) [33] as well as its recently proposed role in double-strand breaks (DSBs) repair [34, 35]
The effects of the viruses on Tab182 levels closely mirror those on MRE11 and, in the case of Ad4, on p53 (Fig. 5). (Ad7, Ad9, and Ad11 all markedly induce the expression of p53, as was reported previously [28 and 56].) We previously reported that Ad4 facilitates the rapid degradation of various DDR proteins [28] perhaps to a lesser extent than Ad5 and Ad12
Summary
Infection by most DNA viruses activates a cellular DNA damage response (DDR), which may be to the detriment or advantage of the virus. We have identified a novel DDR protein, tankyrase 1 binding protein 1 (TNKS1BP1) ( known as Tab182), which is degraded during infection by adenovirus serotype 5 and adenovirus serotype 12 In both cases, degradation requires the action of the early region 1B55K (E1B55K) and early region 4 open reading frame 6 (E4orf6) viral proteins and is mediated through the proteasome by the action of cullin-based cellular E3 ligases. We have shown that Tab182, which we have confirmed to be an integral component of the mammalian CNOT complex, is degraded following infection by adenovirus serotypes 5 and 12. This requires the viral E1B55K and E4orf proteins and is mediated by cullin-based E3 ligases and the proteasome. Further complexes, comprising Rad9-Rad and Hus1 [-1] and Rad17replication factor C2 (RFC2) clamp loader, together with TOPBP1 are recruited to ssDNA, RPA, and ATR, leading to cell cycle arrest and repair [17,18,19]
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