Critical role of the ubiquitin ligase activity of UHRF1, a nuclear RING finger protein, in tumor cell growth.
Early cellular events associated with tumorigenesis often include loss of cell cycle checkpoints or alteration in growth signaling pathways. Identification of novel genes involved in cellular proliferation may lead to new classes of cancer therapeutics. By screening a tetracycline-inducible cDNA library in A549 cells for genes that interfere with proliferation, we have identified a fragment of UHRF1 (ubiquitin-like protein containing PHD and RING domains 1), a nuclear RING finger protein, that acts as a dominant negative effector of cell growth. Reduction of UHRF1 levels using an UHRF1-specific shRNA decreased growth rates in several tumor cell lines. In addition, treatment of A549 cells with agents that activated different cell cycle checkpoints resulted in down-regulation of UHRF1. The primary sequence of UHRF1 contains a PHD and a RING motif, both of which are structural hallmarks of ubiquitin E3 ligases. We have confirmed using an in vitro autoubiquitination assay that UHRF1 displays RING-dependent E3 ligase activity. Overexpression of a GFP-fused UHRF1 RING mutant that lacks ligase activity sensitizes cells to treatment with various chemotherapeutics. Taken together, our results suggest a general requirement for UHRF1 in tumor cell proliferation and implicate the RING domain of UHRF1 as a functional determinant of growth regulation.
Jenkins
Yonchu
Y
Rigel Pharmaceuticals, South San Francisco, CA 94080, USA. yjenkins@rigel.com
Markovtsov
Vadim
V
Lang
Wayne
W
Sharma
Poonam
P
Pearsall
Denise
D
Warner
Justin
J
Franci
Christian
C
Huang
Betty
B
Huang
Jianing
J
Yam
George C
GC
Vistan
Joseph P
JP
Pali
Erlina
E
Vialard
Jorge
J
Janicot
Michel
M
Lorens
James B
JB
Payan
Donald G
DG
Hitoshi
Yasumichi
Y
D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology*; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D009369:Neoplasms / Q000201:enzymology*; D016376:Oligonucleotides, Antisense; D011994:Recombinant Proteins / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D014158:Transcription, Genetic; D044767:Ubiquitin-Protein Ligases
D022762:CCAAT-Enhancer-Binding Proteins; D016376:Oligonucleotides, Antisense; D011994:Recombinant Proteins; C406364:UHRF1 protein, human; D044767:Ubiquitin-Protein Ligases