Molecular cloning of a novel mitogen-inducible nuclear protein with a Ran GTPase-activating domain that affects cell cycle progression.
We have cloned a novel cDNA (Spa-1) which is little expressed in the quiescent state but induced in the interleukin 2-stimulated cycling state of an interleukin 2-responsive murine lymphoid cell line by differential hybridization. Spa-1 mRNA (3.5 kb) was induced in normal lymphocytes following various types of mitogenic stimulation. In normal organs it is preferentially expressed in both fetal and adult lymphohematopoietic tissues. A Spa-1-encoded protein of 68 kDa is localized mostly in the nucleus. Its N-terminal domain is highly homologous to a human Rap1 GTPase-activating protein (GAP), and a fusion protein of this domain (SpanN) indeed exhibited GAP activity for Rap1/Rsr1 but not for Ras or Rho in vitro. Unlike the human Rap1 GAP, however, SpanN also exhibited GAP activity for Ran, so far the only known Ras-related GTPase in the nucleus. In the presence of serum, stable Spa-1 cDNA transfectants of NIH 3T3 cells (NIH/Spa-1) hardly overexpressed Spa-1 (p68), and they grew as normally as did the parental cells. When NIH/Spa-1 cells were serum starved to be arrested in the G1/G0 phase of the cell cycle, however, they, unlike the control cells, exhibited progressive Spa-1 p68 accumulation, and following the addition of serum they showed cell death resembling mitotic catastrophes of the S phase during cell cycle progression. The results indicate that the novel nuclear protein Spa-1, with a potentially active Ran GAP domain, severely hampers the mitogen-induced cell cycle progression when abnormally and/or prematurely expressed. Functions of the Spa-1 protein and its regulation are discussed in the context of its possible interaction with the Ran/RCC-1 system, which is involved in the coordinated nuclear functions, including cell division.
Hattori
M
M
Department of Immunology and Cell Biology, Faculty of Medicine, Kyoto University, Japan.
Tsukamoto
N
N
Nur-e-Kamal
M S
MS
Rubinfeld
B
B
Iwai
K
K
Kubota
H
H
Maruta
H
H
Minato
N
N
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002453:Cell Cycle; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D020690:GTPase-Activating Proteins; D015870:Gene Expression; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008934:Mitogens / Q000494:pharmacology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D020931:ran GTP-Binding Protein; D020668:rap GTP-Binding Proteins
D017931:DNA Primers; D020690:GTPase-Activating Proteins; D008934:Mitogens; D009687:Nuclear Proteins; D012333:RNA, Messenger; C490403:SIPA1 protein, human; C091108:Sipa1 protein, mouse; D019204:GTP-Binding Proteins; D020931:ran GTP-Binding Protein; D020668:rap GTP-Binding Proteins