Biology Reference
In-Depth Information
In permissive cells, the early phase of the infection is associated with the
stimulation of many genes encoding proteins that are required for host cell DNA
synthesis and proliferation (Hirai and Watanabe 1976; Estes and Huang 1977;
Isom 1979; Boldogh et al. 1991; Wade et al. 1992; Browne et al. 2001). Many
of these genes are regulated by the E2F/DP transcription factors, which are
inhibited by complex formation with the Rb family of proteins. A role for IE2-
86 has been suggested by work showing that there is an increase in the steady
state levels of RNA from several E2F-responsive genes in human fibroblasts
infected with an adenovirus expressing IE2-86 (Song and Stinski 2002). The key
question, however, is what are the underlying mechanisms for the activation of
these growth regulatory genes in the context of the infection?
Cyclin E expression is regulated by E2F, and the potential role of IE2-86 in its
accumulation during the infection has been the focus of several studies (Bresnahan
et al. 1998; McElroy et al. 2000; Wiebusch and Hagemeier 2001; Wiebusch et al.
2003). The majority of the experiments have used transient expression assays to
examine the regulation of the cyclin E promoter driving a reporter gene. In one
study, it was shown that IE2-86 could bind to sequences in the cyclin E promoter
in vitro and could activate expression of a cyclin E promoter-driven reporter con-
struct (Bresnahan et al. 1998). Work by Song and Stinski demonstrated that expres-
sion of IE2-86 induces synthesis of endogenous cyclin E mRNA, reinforcing the
notion that IE2-86 expression upregulates cyclin E in infected cells (Song and
Stinski 2002). In contrast, McElroy et al. reported that early viral gene expression,
not IE2-86 expression, was necessary for accumulation of cyclin E protein
(McElroy et al. 2000). These conflicting results suggest that cyclin E accumulation
in infected cells is controlled by several pathways, and that the increase in cyclin E
might be regulated at both the mRNA and protein level.
Besides E2F transcription factors, other proteins regulate the expression of
cyclin genes. The finding that the architectural transcription factor HMGA2 (high
mobility group AT-hook 2) regulates the transcription of cyclin A (Tessari et al.
2003) prompted our lab to determine whether HCMV affects the expression of
this protein. HMGA proteins are referred to as architectural transcription factors
because of their ability to organize the assembly of nucleoprotein structures
(enhanceosomes), resulting in enhancement or repression of transcription. In the
case of cyclin A, HMGA2 activates its expression through derepression of the
promoter. Our studies showed that the transcription of the HMGA2 gene is spe-
cifically inhibited at the RNA level during the infection (Shlapobersky et al.
2006). To determine whether repression of HMGA2 was important for the HCMV
infection, a recombinant virus expressing HMGA2 driven by the MIE promoter
was constructed. High multiplicity infection with the HMGA2-expressing virus
induced the synthesis of cyclin A mRNA and protein and inhibited virus replica-
tion. Furthermore, a role for IE2-86, but not IE1-72, in HMGA2 repression was
suggested by additional experiments with HCMV recombinant viruses defective
in either IE1-72 or IE2-86. We found that the IE1-72 deletion mutant virus CR208
(Greaves and Mocarski 1998) inhibits HMGA2 transcription. In contrast, in cells
infected with an IE2-86 mutant virus lacking aa 136-290 (referred to as delta SX)
