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Abstract: FR-PO533

MEF2A Is a Novel Osmoregulatory Transcription Factor

Session Information

Category: Fluid, Electrolytes, and Acid-Base Disorders

  • 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

  • Watts, Jason A., National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States
  • Wan, Ma, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States
  • Delker, Don, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States
  • Arroyo Ornelas, Juan Pablo, Vanderbilt University Medical Center, Nashville, Tennessee, United States
  • Soerianto, Winny, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States
Background

Osmoregulation is a complex component of cell physiology, yet the initial transcriptional regulators of the osmotic stress response remain to be fully characterized. Here we studied the regulation of promoter-proximal RNA polymerase pausing after osmotic stress and uncovered MEF2A as a tonicity-responsive transcription factor.

Methods

We identified tonicity responsive genes by expression profiling in skin fibroblasts and inner medulla collecting ducts cells following osmotic stress. To identify genes with paused RNA polymerase we used the Precision nuclear Run-On and Sequencing assay (PRO-seq) to determine the locations of actively transcribing RNA polymerases.

Results

Aldose reductase (AKR1B1) and sodium myoinositol transporter (SLC5A3) are tonicity-responsive genes marked by abundant paused RNA Pol II at baseline. Both genes are induced by hypertonic stress, but induction is abrogated in the presence of a small molecule that stabilizes paused RNA Pol II. While AKR1B1 and SLC5A3 require pause-release for gene induction, not all genes with paused polymerase are induced by hypertonic stress, indicating specificity to pause-release. To identify other pause-dependent tonicity-responsive genes, we performed RNA sequencing and Precision Nuclear Run-On sequencing (PRO-seq) to measure gene expression and nascent transcription. We observed pause-dependent induction of 335 tonicity-responsive genes. To ask how pausing at these genes is regulated, we searched for transcription factor binding sites and identified motifs for MEF2A at 48% of the induced genes. We then performed CUT&Tag in fibroblasts and found tonicity-dependent binding of MEF2A at 1230 sites, including the SLC5A3 promoter. Next, we knocked down MEF2A using siRNA and found SLC5A3 and AKR1B1 expression is significantly reduced following hypertonic stress. Finally, we water restricted mice to test if MEF2A responds to physiologic hypertonic stress. We observed increased MEF2A staining in the nuclei of renal collecting duct cells of water restricted mice compared to controls.

Conclusion

Together, we show MEF2A exhibits osmotic stress-dependent nuclear localization and chromatin binding, and that it regulates expression of SLC5A3 and AKR1B1. We suggest MEF2A mediates RNA Pol II pause release to induce transcription of tonicity-responsive genes.

Funding

  • Other NIH Support