The 5' Untranslated Region of the Major Immediate Early mRNA Is Necessary for Efficient Human Cytomegalovirus Replication.

The human cytomegalovirus (HCMV) immediate early 1 (IE1) and IE2 proteins are critical regulators of virus replication. Both proteins are needed to efficiently establish lytic infection, and nascent expression of IE1 and IE2 is critical for reactivation from latency. The regulation of IE1 and IE2 protein expression is thus a central event in the outcome of HCMV infection. Transcription of the primary transcript encoding both IE1 and IE2 is well studied, but relatively little is known about the posttranscriptional mechanisms that control IE1 and IE2 protein synthesis. The mRNA 5' untranslated region (5' UTR) plays an important role in regulating mRNA translation. Therefore, to better understand the control of IE1 and IE2 mRNA translation, we examined the role of the shared 5' UTR of the IE1 and IE2 mRNAs (MIE 5' UTR) in regulating translation. In a cell-free system, the MIE 5' UTR repressed translation, as predicted based on its length and sequence composition. However, in transfected cells we found that the MIE 5' UTR increased the expression of a reporter gene and enhanced its association with polysomes, demonstrating that the MIE 5' UTR has a positive role in translation control. We also found that the MIE 5' UTR was necessary for efficient IE1 and IE2 translation during infection. Replacing the MIE 5' UTR with an unstructured sequence of the same length decreased IE1 and IE2 protein expression despite similar levels of IE1 and IE2 mRNA and reduced the association of the IE1 and IE2 mRNAs with polysomes. The wild-type MIE 5'-UTR sequence was also necessary for efficient HCMV replication. Together these data identify the shared 5' UTR of the IE1 and IE2 mRNAs as an important regulator of HCMV lytic replication.