Subcellular Localization and Functional Activity of the Cyclin-Dependent Kinase Inhibitor p21 WAF1/CIP1

18 Pages Posted: 1 Jul 2013

See all articles by Bahareh Shafaei

Bahareh Shafaei

Jackson Park Hospital; University of Ottawa

Niloofer Shahid

University of Ottawa

Date Written: December 5, 2009

Abstract

Human p21WAF1/Cip1 gene encodes for a cyclin-dependent kinase (CDK) inhibitor that blocks the activity of A-Cdk2, cyclin E-CdkP, cyclin Dl-Cdk4, and cyclin D2-Cdk4. By inhibiting CDK’s, which are needed for the expression of proteins required in the S phase, p21 elongates the G1 phase and prevents the cells from progressing into the S phase. In this study we labelled p21 with the EGFP using the cloning vector pEGFP-N1, cloned it in prokaryotic cells (Top 10 E. Coli bacteria), and expressed it in eukaryotic HeLa cells in an attempt to study its distribution, localization, and function. Our results showed that the HeLa cells expressed p21; furthermore, we were able to visualize its location using fluorescence microscopy, and it was found to be in the nucleus. Additionally, by using Flow cytometry, we were able to assay the function of p21 and its effect on the cell cycle by monitoring the percentage of cells in each of the mitotic phases (G1, S, and G2-M) and their DNA content. We found that a higher percentage of cells expressing p21 are found in the G0/G1 phase and a lower percentage of those cells found in the S phase and G2 compared to the negative control.

Keywords: DNA, Genome,Cell Cycle, Cyclin-Dependent Kinase, Flow cytometry, p21, p53

JEL Classification: l11

Suggested Citation

Shafaei, Bahareh and Shafaei, Bahareh and Shahid, Niloofer, Subcellular Localization and Functional Activity of the Cyclin-Dependent Kinase Inhibitor p21 WAF1/CIP1 (December 5, 2009). Available at SSRN: https://ssrn.com/abstract=2287636 or http://dx.doi.org/10.2139/ssrn.2287636

Bahareh Shafaei (Contact Author)

Jackson Park Hospital ( email )

7531 Stony Island Avenue
Chicago, IL 60649
United States

University of Ottawa ( email )

2292 Edwin Crescent
Ottawa, Ontario K2C 1H7
Canada

Niloofer Shahid

University of Ottawa ( email )

2292 Edwin Crescent
Ottawa, Ontario K2C 1H7
Canada

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