Functional cloning of SPIN-2, a nuclear anti-apoptotic protein with roles in cell cycle progression.
The balance between hematopoietic cell viability and apoptosis is regulated by exogenous growth factors, however, the molecular mechanisms by which these trophic factors exert their effects remain obscure. A functional retroviral cDNA library-based screen was employed to identify genes that prevent growth factor withdrawal-mediated apoptosis in the myeloid progenitor cell 32Dcl3. This approach identified three classes of genes: those with known roles in apoptosis (bcl-X(L) and ornithine decarboxylase); genes previously identified but not linked directly to apoptotic signaling (O-linked N-acetylglucosamine transferase); and a previously uncharacterized gene we termed SPIN-2. In 32Dcl3 cells, expression of exogenous SPIN-2 provides 25% protection from apoptosis following growth factor withdrawal compared to controls which show approximately 1-2% survival. SPIN-2 overexpression slows cell growth rates and increases the percentage of cells in G(2)/M (32% vs control cells at 12%). Immunolocalization studies indicate that myc-epitope tagged SPIN-2 proteins, which retain their anti-apoptotic function, reside in the nucleus, whereas a C-terminal deletion mutant that loses its anti-apoptotic activity is located in the cytoplasm. These studies suggest that SPIN-2 is a novel nuclear protein that functions to regulate cell cycle progression and this activity is related to the inhibition of apoptosis following the removal of essential growth factors.
Fletcher
B S
BS
Department of Pharmacology and Therapeutics, University of Florida, College of Medicine, Gainesville, FL 32610-0267, USA.
Dragstedt
C
C
Notterpek
L
L
Nolan
G P
GP
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D002453:Cell Cycle / Q000235:genetics*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D002460:Cell Line / Q000166:cytology / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D020869:Gene Expression Profiling; D015723:Gene Library; D005822:Genetic Vectors / Q000235:genetics; D018922:HL-60 Cells / Q000737:chemistry; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D019169:Jurkat Cells / Q000166:cytology; D051379:Mice; D008969:Molecular Sequence Data; D022423:Myeloid Cells / Q000166:cytology / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology; D012190:Retroviridae / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014960:X Chromosome / Q000235:genetics
D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D007377:Interleukin-3; D009687:Nuclear Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D011993:Recombinant Fusion Proteins; C463239:SPIN2A protein, human