Identifying the Mechanism(s) Responsible for the Translational Regulation of the Stress Signaling Kinase MKK4
Abstract
We recently demonstrated that the endogenous level of MKK4 is regulated post-transcriptionally in prostate cancer cell lines. Using WI-38 lung fibroblasts (WI-38 cells), which increase their MKK4 expression as they undergo senescence, we showed that a cohort of four miRNAs (miRs-15b, -24, -25, -141) decreased as the cells senesce. Treating WI-38 cells with precursor miRNAs (pre-miRs) to all four target miRNAs together but not individually, led to a significant decrease in MKK4 protein levels. Conversely, treatment with all four antisense miRNAs (ASmiRs) significantly increased MKK4 protein levels. To test if this effect is conserved in prostate cancer models, the levels of MKK4 mRNA and protein were quantitated in C4-2, CWR22-Rv1, DuPro, LAPC4, LNCaP, and PC3 cell lines and correlated to the endogenous levels of the cohort miRNAs. We hypothesized that miR-24 and miR-141 would be the best candidates in the cohort to regulate MKK4 protein expression based on the high versus low miRNA levels in the CaP lines and published data showing relevance to prostate cancer progression. However, our preliminary data showed that transfection of CWR22-Rv1 and PC3 cells with pre-miR-24 or pre-miR-141 did not decrease MKK4 protein levels. Moreover, transfection of those same lines with AS-miR-24 or AS-miR-141 did not increase MKK4 expression. Therefore the revised working hypothesis is that the entire cohort of miRs is required to affect the MKK4 protein levels. Current studies include the simultaneous transient transfection of all four miRs into CWR22-Rv1 and PC3 cells. Also, stable transfections of cohort Pre- and AS-miRNAs will allow us to test the effect of those miRs on the cells metastatic potential.
Document Details
- Document Type
- Technical Report
- Publication Date
- Aug 01, 2010
- Accession Number
- ADA552006
Entities
People
- Carrie Rinker-schaeffer
Organizations
- University of Chicago