Page 154 - Avian Virology: Current Research and Future Trends

Infectious Bronchitis Virus | 147

Figure 11. Infectious bronchitis virus (IBV) life cycle.
Figure 5.11 Infectious bronchitis virus (IBV) life cycle.

it would be exhibiting a more efficient infection (Tay et al., on the CDS of rep1b (Brierley et al., 1987, 1989). Currently, the
2012). Following fusion of the viral and cellular membranes, incidence of ribosomal frameshifting is still under debate, with
the virus enters the cell via endocytosis. Here, IBV fusion a range from 15% to as high as 60% depending on the in vitro
occurs at a pH-dependent manner, with a half-maximal fusion studies performed (Baranov et al., 2005; Plant et al., 2005; Su et
rate occurring at pH 5.5, indicating that endosomal acidification al., 2005). It is also not known exactly the rationale behind the
may be a fusion trigger for IBV as well as other CoVs (Chu et adoption of ribosomal frameshifting in CoVs. Two schools of
al., 2006a). Several entry routes have been described for CoVs thought have been proposed: one which holds that by adopting
following infection, such as clathrin- and caveolae-independent this translation mechanism, the ratio of pp1a and pp1ab could
and caveolae-dependent pathways (Nomura et al., 2004; van be regulated, while the other believes that the expression of
Hamme et al., 2008). Inhibitors of clathrin-dependent path- rep1b products could be delayed until a suitable cellular envi-
way such as chlorpromazine have been shown to abolish IBV ronment has been created for RNA replication by the products
infection (Chu et al., 2006b). More recent studies suggest that of rep1a (Liu et al., 1994; Fehr and Perlman, 2015).
the cellular sites of the uncoating events may be at the late Next, pp1a and pp1ab are further processed to form 15 nsps
endosomes (White and Whittaker, 2016; Wong et al., 2015). (Fig. 5.12) (Liu et al., 1994, 1997). The size (amino acids)
and cleavage sites of these 15 final products are summarized in
Replicase translation and processing Table 5.3. All CoVs would encode at least two proteases required
Following the release of the viral genomic RNA (gRNA) into for this cleavage. These are the papain-like proteases (PLpro)
the cytoplasm, the next step in the CoV replication cycle is encoded by nsp3 and the main protease (Mpro) encoded by
to translate the replicase genes, rep1a and rep1b, into poly- nsp5. With the exception of the gammacoronaviruses, SARS-
proteins pp1a and pp1ab (Fig. 5.12). Translation of the rep1b CoV and MERS-CoV, most CoVs would encode two PLpros in
gene does not follow the usual rules of translation. It is instead, nsp3 (Woo et al., 2010). These PLpros would cleave nsps 1–4
translated via an alternate mechanism of translation known as at the nsp 1/2, 2/3, 3/4 boundaries, while Mpro is involved in
ribosomal frame-shifting, in which the translating ribosome the downstream cleavage events. Prediction and comparative
shifts, with a fixed probability, in the –1 direction, from rep1a analyses of Mpro cleavage sites in seven CoVs revealed that
reading frame into rep1b reading frame. Two RNA elements are the substrate specificity of Mpro is exclusively occupied by
essential for ribosome repositioning: (1) a slippery sequence glutamine (Gln) at the P1 position, and this is essential for
(5′-UUUAAAC-3′) and (2) an RNA pseudoknot. Usually, the efficient cleavage (Ziebuhr et al., 2000). Amino acid substitu-
ribosome works by unwinding the pseudoknot and carrying on tions of Gln in the P1 position at different cleavage sites display
with translation in rep 1a until it encounters the stop codon. variable degrees of growth defects in IBV, with some sites being
When that occurs, the pseudoknot could sometimes prevent the well-tolerated while others impede virus recovery (Fang et al.,
translating ribosome from further elongation. Under this cir- 2008, 2010). Collectively, the nsps generated would assemble
cumstance, the ribosome would pause on the slippery sequence to form a larger complex known as the replication-transcription
and shifts the reading frame by –1 position before the ribosome complex (RTC), responsible for RNA replication and sub-
can overcome the pseudoknot structure and resume translation genomic RNA transcription.