C-terminal domain of p16(INK4a) is adequate in inducing cell cycle arrest, growth inhibition and CDK4/6 interaction similar to the full length protein in HT-1080 fibrosarcoma cells.
The tumor suppressor p16(INK4a) has earned widespread attention in cancer studies since its discovery as an inhibitor of cyclin-dependent kinases (CDKs) 4/6. Structurally, it consists of four complete ankyrin repeats, believed to be involved in CDK4 interaction. According to the previous disparities concerning the importance of domains and inactivating mutations in p16, we aimed to search for the domain possessing the functional properties of the full length protein. Upon our in silico screening analyses followed by experimental assessments, we have identified the novel minimum functional domain of p16 to be the C-terminal half including ankyrin repeats III, IV and the C-terminal flanking region accompanied by loops 2 and 3. Transfection of this truncated form into HT-1080 human fibrosarcoma cells, lacking endogenous p16, revealed that it is able to inhibit cell growth and proliferation equivalent to p16(INK4a). The functional analysis showed that this fragment like p16 can interact with CDK4/6, block the entry into S phase of the cell cycle and suppress growth as indicated by colony formation assay. Identification of p16 minimum functional domain can be of benefit to the future peptidomimetic drug design as well as gene transfer for cancer therapy.
Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
Najmeh
N
Fahham
Soroush
S
Sardari
Seyed Nasser
SN
Ostad
Behrouz
B
Vaziri
Mohammad Hossein
MH
Ghahremani
D002453:Cell Cycle / Q000235:genetics / Q000502:physiology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics / Q000502:physiology; D051358:Cyclin-Dependent Kinase 4 / Q000235:genetics / Q000378:metabolism*; D051361:Cyclin-Dependent Kinase 6 / Q000235:genetics / Q000378:metabolism*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism*; D005354:Fibrosarcoma / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D011485:Protein Binding; D020133:Reverse Transcriptase Polymerase Chain Reaction
D019941:Cyclin-Dependent Kinase Inhibitor p16; C495900:CDK4 protein, human; C495911:CDK6 protein, human; D051358:Cyclin-Dependent Kinase 4; D051361:Cyclin-Dependent Kinase 6