Functional analysis of wild-type and malignant glioma derived CDKN2Abeta alleles: evidence for an RB-independent growth suppressive pathway.
The tumor suppressor gene CDKN2A (p16/MTS1/INK4A), which encodes the cyclin-dependent kinase inhibitor p16(INK4a), is a target of 9p21 deletions during the malignant progression of human gliomas. This gene also encodes a second protein product (human p16beta, murine p19ARF), which originates from an unrelated exon of CDKN2A (exon 1beta) spliced onto exon 2 in an alternate reading frame. Cell cycle arrest by p16beta is caused by an as yet unidentified pathway. In order to test the candidacy of p16beta as a glioma suppressor, we replaced p16(INK4a), p15(INK4b) and p16beta wild-type as well as a series of seven glioma-derived p16beta alleles (R87H, A112V, R120H, A121V, G125R, A128A and A128V), into glioma cell lines that had either CDKN2A-/RB+ (U-87MG and U-251MG) or CDKN2A+/RB- (LN-319) endogenous backgrounds and demonstrated that p16beta can act as a functional glioma cell growth suppressor. Moreover, p16beta, but not p16(INK4a) or p15(INK4b) inhibited the growth of RB-negative LN-319 cells, indicating that p16beta likely exerts its effects through an RB-independent pathway. In vitro and in vivo assays of pRB phosphorylation were consistent with this interpretation. Since none of the glioma-derived p16beta mutations inactivated their growth suppressive activities, it appears that mutations in CDKN2A exon 2 (which is shared in the coding sequences of p16(INK4a) and p16beta) likely exclusively target p16(INK4a).
Arap
W
W
Ludwig Institute for Cancer Research, University of California-San Diego, La Jolla 92093-0660, USA.
Knudsen
E
E
Sewell
D A
DA
Sidransky
D
D
Wang
J Y
JY
Huang
H J
HJ
Cavenee
W K
WK
D000483:Alleles; D002455:Cell Division / Q000235:genetics; D002872:Chromosome Deletion; D002899:Chromosomes, Human, Pair 9 / Q000235:genetics; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000502:physiology; D016147:Genes, Tumor Suppressor / Q000235:genetics* / Q000502:physiology; D005822:Genetic Vectors / Q000235:genetics; D005910:Glioma / Q000235:genetics* / Q000473:pathology; D006801:Humans; D010766:Phosphorylation; D017354:Point Mutation; D016160:Retinoblastoma Protein / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D019941:Cyclin-Dependent Kinase Inhibitor p16; D016160:Retinoblastoma Protein