Interactions between human cytomegalovirus IE1-72 and cellular p107: functional domains and mechanisms of up-regulation of cyclin E/cdk2 kinase activity.
Previous work has demonstrated that the human cytomegalovirus IE1-72 protein is able to bind to the N terminus of p107, and IE1-72 alone is sufficient for alleviation of p107-mediated cell growth suppression. However, the mechanism of this alleviation is unclear. Here, we show that IE1-72 can alleviate p107 inhibition of cyclin E/cdk2 kinase activity. We cotransfected various IE1-72 and p107 constructs into C33A cells and demonstrated that IE1-72 could activate the kinase activity of cyclin E/cdk2. Conversely, IE2-86 did not activate this activity, suggesting that the interaction between p107 and IE1-72 and the subsequent kinase activation are specific. By the use of a series of deletion and point mutants of IE1-72 and p107, we observed that a mutation of the loop region of helix-loop-helix-turn-helix in exon 3 of IE1-72 as well as a mutation of the leucine zipper-2 region in exon 4 of IE1-72 abolished binding to p107. In addition, these two IE1-72 mutants did not alleviate p107 inhibition of cyclin E/cdk2 kinase activity and also failed to alleviate p107 inhibition of the E2F-responsive promoter. Meanwhile, deletion of the N-terminal aa 1 to 175 of p107 abolished both p107 binding with IE1-72 and p107 inhibition of cyclin E/cdk2 kinase activity. This result confirms that the N-terminus aa 1 to 175 region of p107 is a common region where both IE1-72 protein and cyclin E/cdk2 bind. We propose a mechanism in which binding of IE1-72 to p107 displaces cyclin E/cdk2 from p107. Once released from p107, cyclin E/cdk2 is able to function as an active kinase.
Zhang
Zhigang
Z
Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7295, USA.
Huong
Shu-Mei
SM
Wang
Xin
X
Huang
David Y
DY
Huang
Eng-Shang
ES
D001483:Base Sequence; D042846:CDC2-CDC28 Kinases / Q000378:metabolism*; D045744:Cell Line, Tumor; D019927:Cyclin E / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D017931:DNA Primers; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism*; D011485:Protein Binding; D015854:Up-Regulation; D014764:Viral Proteins
D019927:Cyclin E; D017931:DNA Primers; C088645:IE1 protein, cytomegalovirus; D017874:Immediate-Early Proteins; D014764:Viral Proteins; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2