Biology Reference
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and 275 positively or negatively affects viral growth depending on the location of
the residues (Barrasa et al. 2005) . Mutation of residues between 271 and 275 accel-
erates viral growth and mutation of residues between 258 and 264 or 266 and 269
delays viral growth (Table 1 ).
The IE86 protein has two nuclear localization signals (N) that can independ-
ently target the viral protein to the nucleus (Fig. 2a) (Pizzorno et al. 1991). In the
nucleus, the viral protein affects viral promoters through two transcriptional
activation domains, one amino and the other carboxyl (Fig. 2a) (Malone et al.
1990; Pizzorno et al. 1991; Yeung et al. 1993; Stenberg 1996). For transcriptional
regulatory activity, the IE86 protein functions as a homodimer and dimerizes
through the region broadly designated in Fig. 2a (Macias et al. 1996). Within
this region, there is a putative zinc finger between amino acids 428 and 452,
which may be part of a double zinc finger motif between amino acids 428 and
480 (CX
5
CX
11
HX
5
HXDXCX
13
HXH) (Fig. 2a). Double zinc finger motifs are
important for interactions with other viral or cellular proteins such as transcrip-
tion factors that either activate or repress transcription (Bachy et al. 2002;
Moreno et al. 2003).
Figure 2a summarizes the regions of posttranslational modification and the
broadly mapped functional domains of the IE86 protein. Multiple amino acid dele-
tions were made to determine the functional domains of the IE86 protein and these
mutations are summarized in Table 1. Large deletions resulted in nonreplicating
genomes and confirmed that the IE86 protein is essential for HCMV replication.
Smaller deletions affected the efficiency of either early or late viral gene expression.
Frequently more than one viral protein function was affected by these deletions.
Mutations that affected DNA binding and negative autoregulation produced high
levels of the IE86 protein, which also affected cell viability. Mutations that failed to
interact with TBP or TFIIB affected activation of early viral promoters. Mutations
that affected phosphorylation of the viral protein affected the rate of viral growth.
Delayed viral growth was associated with reduced expression of viral tegument
proteins pp65 and pp28 (Sanchez et al. 2002). Many of the mutations made it diffi-
cult to assign a particular function to a specific region of the IE86 protein. The
region between amino acids 450 and 552 was defined as a core domain because the
IE86 regulatory functions of negative autoregulation of the MIE promoter, early
promoter transactivation, and cell cycle arrest were all affected by any deletion
within this region (Asmar et al. 2004).
Figure 2b shows regions of conserved amino acids in the carboxyl end of the
viral protein between primate and nonprimate CMV homologs of the IE86 pro-
tein. The carboxy terminus is more conserved than the amino terminus. There
are conserved stretches of amino acids suggesting critical structural and
functional domains within the protein. Petrik et al. made rationally designed
amino acid substitutions in the core region based on sequence conservation
(Petrik et al. 2006, 2007); these results are also summarized in Table 1. These
mutations separated the transactivation domain from the cell cycle arrest domain
and the autoregulation domain from the transactivation domain (Fig. 2a).
However, mutations in the putative zinc finger motif affected all functions of the
