Title:Unraveling Potential Candidate Targets Associated with Expression of
p16INK4a or p16 Truncated Fragment by Comparative Proteomics Analysis
Volume: 19
Issue: 2
Author(s): Najmeh Fahham, Fatemeh Zandi, Mohammad Hossein Ghahremani, Seyed Nasser Ostad, Behrouz Vaziri*, Seyed Sadegh Shahraeini and Soroush Sardari*
Affiliation:
- Protein Chemistry and Proteomics Laboratory, Biotechnology Research Center, Pasteur Institute of Iran, Tehran,
Iran
- Drug Design and Bioinformatics Unit, Department of Medical Biotechnology, Biotechnology Research
Centre, Pasteur Institute of Iran, Tehran, Iran
Keywords:
p16INK4A, proteomics, fibrosarcoma, oxidative stress, protein metabolism, cytoskeleton, signal transduction.
Abstract:
Background: p16 is a tumor suppressor protein that is significantly involved in cycle
regulation through the reduction of cell progression from the G1 phase to the S phase via CDK-cyclin
D/p16INK4a/pRb/E2F cascade. The minimum functional domain of p16 has been uncovered that
may function comparable to wild type p16.
Objective: To expand the knowledge on molecules and mechanisms by which p16 or p1666-156 fragment
suppresses human fibrosarcoma cell line growth, differential proteome profiles of fibrosarcoma
cells following p16 full length or the functional domain overexpression, were analyzed.
Methods: Following transfecting HT-1080 fibrosarcoma cells with p16 full length, p1666-156 truncated
form, and pcDNA3.1 empty vector, protein extract of each sample was harvested and clarified
by centrifugation, and then the protein content was determined via Bradford assay. All protein extract
of each sample was analyzed by two-dimensional gel electrophoresis. Immunoblot analysis
was performed as further validation of the expression status of identified proteins.
Results: Expression of p16 or p1666-156 fragment could induce mostly the common alterations (up/-
down-regulation) of proteome profile of HT-1080 cells. Mass spectrometry identification of the differentially
expressed protein spots revealed several proteins that were grouped in functional clusters,
including cell cycle regulation and proliferation, cell migration and structure, oxidative stress,
protein metabolism, epigenetic regulation, and signal transduction.
Conclusion: The minimum functional domain of p16 could act in the same way as p16 full length.
Also, these new findings can significantly enrich the understanding of p16 growth-suppressive
function at the molecular level by the introduction of potential candidate targets for new treatment
strategies. Furthermore, the present study provides strong evidence on the functional efficacy of
the identified fragment of p16 for further attempts toward peptidomimetic drug design or gene
transfer to block cancer cell proliferation.
