Friday, March 13, 2015

Understanding Mechanisms of Cancer: Krüppel-Like Factor 4

Recent research brings medical investigators a step closer to unraveling one of the most intractable problems in medicine today, the mechanisms through which DNA damage and uncontrolled cell division leads to the break-down in homeostasis known as cancer. Published online in Molecular Carcinogenesis, a 2014 study by Colgate University and Memorial-Sloan Kettering researchers examined a pathway involving Krüppel-Like Factor 4 (KLF4), a protein in eukaryotic cells that regulates tumorigenesis. Previous research by El-Karim et al. had demonstrated that KLF4-deficient mouse cells are genetically unstable, putting them at increased risk for mutation leading to cancer. So the researcher’s experimental objective was to determine how the KLF4 pathway works to arrest cell growth in pre-cancerous mouse cells.

KLF4 serves a variety of cellular functions, including tumor suppression in some cancers. (An article published in Molecular and Cellular Biology by Zhang et al. corroborates the role of KLF4 as a tumor suppressor.) Studies indicate p53, a protein heralded as the guardian of the genome, is regulated by KLF4 when DNA damage is detected, resulting in the inhibition of cell growth. When KLF4 was deficient in a commonly used cell line, mouse embryonic fibroblasts, genomic instability, as measured by DNA double strand breaks, increased. Double strand breaks are a consequence of normal cellular metabolic pathways producing reactive oxygen species, but in higher concentrations. Double strand breaks in DNA can be problematic because the restorative mechanism can cause reorganization of the genome. However, when KLF4 is re-expressed in those mouse cells, they are able to repair the genomic instability. So the question becomes, why don’t all cells with genomic instability develop into tumors, and what is the regulatory role played by KLF4?

In the current research, Liu et al. analyzed mouse embryonic fibroblasts where the KLF4 gene was either expressed or repressed. Cells without KLF4 were found to have increased levels of DNA damage leading to earlier manifestation of cellular senescence, where the cells lose their ability to divide and remain in a state of arrested development. A western blot is a microbiology technique using antibodies to identify target proteins after being run through gel electrophoresis. In the experiment, a western blot with p21 antibodies revealed p21 presents earlier in KLF4 deficient cells and later, at a similar concentration, in KLF4 expressed cells. The protein p53 was also found in increased quantities, which is expected since studies have shown that p53 regulates p21 via transcription. The increased p53 and p21 response is consistent with the greater DNA damage that characterizes KLF4 deficient cells. Quantitative PCR, a variant of PCR that quantifies the concentration of target DNA in terms of time, quantitatively determined that Gsta4, a gene that encodes an enzyme that corrects for oxidative damage, is controlled by KLF4 to help reduce reactive oxygen species. The experimental results indicated that the pathway begins with KLF4 regulating Gsta4 that counteracts the DNA-damaging effects of reactive oxygen species. If needed, p53 is expressed to regulate p21 to control cellular senescence to evade tumorigenesis. The figure below demonstrates how a mouse cell with repressed KLF4 affects the pathway (Liu et al., Figure 6).

Figure 6. “A working model for how KLF4 deficiency causes ROS accumulation that leads to premature senescence” From Liu et al.

The research presented in the article by Liu et al. was the first look into the specific pathway through which KLF4 mediates cell growth. More research will be required to confirm the findings, as the reported results were only preliminary.

As expansive as the genome seems, the proteome is even greater and considerably more complex. As such, much more basic research will need to be done in order to gain a complete understanding of how genomic mutations can lead to tumorigenesis. However, each study, like Liu et al., published contributes to the ongoing battle against cancer. Through increased comprehension of the mechanisms that contribute to cancer, a system of personalized medicine, superior to the one size fits all model, can be developed to better fight cancer. For example, if a tumor analysis determined that the patient had a mutated KLF4 gene, the doctors would be able to select a therapy based on that mutation and could correct for the KLF4 mutation. This is only possible if the pharmaceutical companies understand the pathway leading to tumorigenesis and can exploit said pathway to create a drug. Basic research translates into applied research, thereby bettering our ability to repress tumor growth and decrease the death rate from cancer.

Works Cited:

El-Karim, E., E. Hagos, A. Ghaleb, B. Yu, and V. Yang. “Kruppel-like factor 4 regulates genetic stability in mouse embryonic fibroblasts.” Molecular Cancer, 6 August 2013; 12 (1):89
Liu, Changchang, Stephen La Rosa, and Engda G. Hagos. “Oxidative DNA Damage Causes Premature Senescence in Mouse Embryonic Fibroblasts Deficient for Krüppel-Like Factor 4.” Molecular Carcinogenesis. April 30, 2014 doi: 10.1002/mc.22161
Zhang, Wen, Xi Chen, Yoichi Kato, Paul M. Evans, Subo Yuan, Jun Yang, Piotr G. Rychahou, Vincent W. Yang, Xi He, B. Mark Evers, and Chunming Liu. “Novel Cross Talk of Krüppel-Like Factor 4 and β-Catenin Regulates Normal Intestinal Homeostasis and Tumor Repression.” Molecular and Cellular Biology. March 15, 2006 26:6 2055-2064; doi:10.1128/MCB.26.6.2055-2064.2006


  1. I would be interested to see information on how common a mutation of the KLF4 gene is in cancer patients. Are there specific cancer types or locations that result from this mutation?

  2. I agree with Jodi that it would be interesting to see if there are specific cancer conditions based on this mutation. This makes me think of the BRCA lab that everyone does in the lab for Molecules, Cells and GEnes (212/182); it would also be interesting to see if there are any commonalities between the two identified cancerous mutations of the genes. Perhaps by exploring/evaluating any relationship between the two could lead to the discovery of new cancer treatments.