Investigating possible involvement of miR-125b in the E2, NF-кB network

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2015

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Yavuz, Esra

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MicroRNAs (miRNAs) play important roles in diverse biological processes and are emerging as key regulators of tumorigenesis and tumor progression. Some miRNAs act as tumor suppressors and are downregulated in cancers such as miR-125b in breast cancer cell lines and tumors. Earlier, we developed a miR-125b expression model system and performed a transcriptomics experiment. Among many indirect targets, ALCAM (activated leukocyte antigen molecule) appeared to be very significantly upregulated in MCF7-125b cells. ALCAM is an NF-кB induced cellcell contact molecule. Given its significant induction in MCF7-125b cells, we aimed to investigate NF-кB and miR-125b interaction. Interestingly, several studies also linked miR-125b expression and Estrogen (E2) which made us consider whether miR-125b may be a player in the E2-NF-кB network. We investigated nuclear p65, subunit of NF-кB TF (Transcription Factor) and TRAF6 levels, an inducer of NF-кB and a known target of miR-125b in MCF7-125b and T47D-125b cells as well as in E2 treated cells. E2 treated cells demonstrated decreased miR-125a and miR-125b levels both in MCF7 and T47D cells. Accordingly, TRAF6 protein levels increased as a result of E2 treatment in T47D cells. However, we failed to detect TRAF6 in vi MCF7 cells. miR- 125b transfected T47D cells also had decreased levels of TRAF6 consistent with miR-125b targeting of TRAF6. NF-кB component nuclear p65 levels also decreased in T47D cells. On the other hand, nuclar localization of p65 also increased in miR-125b transfected MCF7 cells. When MCF7 cells were treated with E2, p65 levels in the nucleus decreased in MCF7 cells. Our data suggests MCF7 and T47D cells are different models of miR-125b expression, possibly due to variations of NF-кB regulatory mechanims. Finally, we detected p65 and TRAF6 levels in miR-125b silenced T47D cells. miR-125b silenced cells had increased TRAF6 and p65 levels, due to decreased miR-125b levels. Our results indicate the complexity of a possible crosstalk between E2, miR-125b and NF-кB and how this complexity may be variable in different cellular backgrounds.

Subject Keywords

RNA., MicroRNAs., Estrogen., Breast

URI

http://etd.lib.metu.edu.tr/upload/12619188/index.pdf
https://hdl.handle.net/11511/24870

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Graduate School of Natural and Applied Sciences, Thesis

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Citation Formats

E. Yavuz, “Investigating possible involvement of miR-125b in the E2, NF-кB network,” M.S. - Master of Science, Middle East Technical University, 2015.