SUMO-1 modification of human cytomegalovirus IE1/IE72.
Human cytomegalovirus (HCMV) immediate-early protein IE1/IE72 is involved in undermining many cellular processes including cell cycle regulation, apoptosis, nuclear architecture, and gene expression. The multifunctional nature of IE72 suggests that posttranslational modifications may modulate its activities. IE72 is a phosphoprotein and has intrinsic kinase activity (S. Pajovic, E. L. Wong, A. R. Black, and J. C. Azizkhan, Mol. Cell. Biol. 17:6459-6464, 1997). We now demonstrate that IE72 is covalently conjugated to the small ubiquitin-like modifier (SUMO-1). SUMO-1 is an 11.5-kDa protein that is conjugated to multiple proteins and has been reported to exhibit multiple effects, including modulation of protein stability, subcellular localization, and gene expression. A covalently modified protein migrating at approximately 92 kDa, which is stabilized by a SUMO-1 hydrolase inhibitor, is revealed by Western blotting with anti-IE72 of lysates from cells infected with HCMV or cells expressing IE72. SUMO modification of IE72 was confirmed by immunoprecipitation with anti-IE72 and anti-SUMO-1 followed by Western blotting with anti-SUMO-1 and anti-IE72, respectively. Lysine 450 is within a sumoylation consensus site (I,V,L)KXE; changing lysine 450 to arginine by point mutation abolishes SUMO-1 modification of IE72. Inhibition of protein phosphatase 1 and 2A, which increases the phosphorylation of IE72, suppresses the formation of SUMO-1-IE72 conjugates. Both wild-type IE72 and IE72(K450R) localize to nuclear PML oncogenic domains and disrupt them. Studies of protein stability, transactivation, and complementation of IE72-deficient HCMV (CR208) have revealed no significant differences between wild-type IE72 and IE72(K450R).
Spengler
Mary L
ML
Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, New York, USA.
Kurapatwinski
Karen
K
Black
Adrian R
AR
Azizkhan-Clifford
Jane
J
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002460:Cell Line; D003587:Cytomegalovirus / Q000502:physiology*; D003586:Cytomegalovirus Infections / Q000821:virology*; D015967:Gene Expression Regulation, Viral; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D025842:SUMO-1 Protein / Q000378:metabolism*; D014764:Viral Proteins
C088645:IE1 protein, cytomegalovirus; D017874:Immediate-Early Proteins; D025842:SUMO-1 Protein; D014764:Viral Proteins