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Expression profiling of Ral-depleted bladder cancer cells identifies RREB-1 as a novel transcriptional Ral effector
G Oxford1, S C Smith, G Hampton
1Department of Molecular Physiology and Biological Physics, Health Sciences Center, University of Virginia, Charlottesville, VA 22908, USA.
Abstract:
Although the monomeric GTPases RalA and RalB have been shown to regulate a variety of transcription factors, little is known regarding the differences or similarities in transcriptional programs regulated by RalA compared to RalB. Further, the association of these transcriptional pathways to human carcinogenesis and progression remains unclear. Here, we studied the role of RalA and/or RalB in transcriptional regulation by combining short interfering RNA depletion of Ral with gene expression profiling via microarray in the human bladder cancer cell line, UMUC-3. A large number of genes were found to be similarly modulated in cells with RalA and RalB depletion, suggesting that RalA and RalB impinge on overlapping transcriptional signaling pathways. However, smaller sets of genes were modulated by depletion of RalA or RalB, indicating that these closely related proteins also regulate nonoverlapping transcriptional pathways. Computational analysis of upstream sequences of genes modulated by Ral depletion identified Ras-responsive element-binding protein (RREB)-1, as a putative Ral transcriptional target, which we verified experimentally. Importantly, as a group, Ral-regulated probe sets identified here were disproportionally represented among those differentially expressed as a function of human bladder transformation. Taken together, these data strongly suggest that Ral family members mediate both common and specific transcriptional programs that are associated with human cancer and identify RREB-1 as a novel transcriptional effector of Ral.
Insights
RalA and RalB GTPases regulate overlapping and distinct gene expression pathways. These Ral-regulated pathways are linked to human bladder cancer, with RREB-1 identified as a novel target.
Area of Science:
- Molecular Biology
- Cancer Research
- Genomics
Background:
- Monomeric GTPases RalA and RalB regulate transcription factors, but their distinct roles in transcriptional programs and cancer are unclear.
- Understanding RalA and RalB's specific contributions to carcinogenesis is crucial.
Purpose of the Study:
- To investigate the differential and overlapping transcriptional programs regulated by RalA and RalB.
- To explore the association of these transcriptional pathways with human bladder cancer progression.
Main Methods:
- Utilized short interfering RNA (siRNA) to deplete Ral proteins in the human bladder cancer cell line UMUC-3.
- Performed gene expression profiling using microarray analysis to identify modulated genes.
- Employed computational analysis to predict transcriptional targets and experimental verification.
Main Results:
- RalA and RalB depletion modulated a significant number of overlapping genes, indicating shared signaling pathways.
- Distinct sets of genes were affected by RalA or RalB depletion, revealing unique regulatory roles.
- Ras-responsive element-binding protein (RREB)-1 was identified as a novel, experimentally verified transcriptional target of Ral.
- Ral-regulated genes were disproportionately represented in differentially expressed genes in human bladder cancer.
Conclusions:
- RalA and RalB mediate both common and specific transcriptional programs relevant to human cancer.
- Ral proteins play a significant role in transcriptional regulation associated with bladder cancer.
- RREB-1 is identified as a novel transcriptional effector of Ral, contributing to its role in cancer.
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