Abstract

Inwardly rectifying potassium (IRK/Kir) channels regulate intracellular K+ concentrations and membrane potential. Disruption of Kir2.1 causes dominantly inherited Andersen Tawil Syndrome (ATS). ATS patients suffer from cardiac arrhythmias, periodic paralysis, and cognitive impairment. These symptoms are consistent with current understanding of the role of ion channels in muscle cells and neurons. However, ATS symptoms also include craniofacial and digital deformities such as cleft palate, dental defects, wide set eyes, low set ears, and crooked or fused digits. These developmental defects were not consistent with current understanding of developmental signaling or previously described roles for ion channels. We found that phenotypes exhibited by the Kir2.1 knockout mouse recapitulate ATS symptoms. The Kir2.1 knockout mouse phenotypes are strikingly similar to those that occur when Transforming Growth Factor β/Bone Morphogenetic Protein (TGFβ/BMP) signaling is disrupted. Based on this observation, we hypothesized that Kir2.1 may play a role in TGFβ/BMP signaling. We tested this hypothesis using Drosophila melanogaster. We reduced a Kir2.1 homologue Irk2 by siRNA, eliminated the Irk2 channel with a deficiency, and abolished heteromeric Irk channel function with a dominant negative Irk2. Reduction of Irk2 function caused wing patterning defects and size reduction that are similar to BMP/Decapentaplegic (DPP) mutant phenotypes. Ubiquitous expression of Irk2DN is lethal. Wing specific Irk2DN expression caused severe defects compared to irk2Df demonstrating that Irk channels are heteromeric. We found that two downstream targets of Dpp were reduced in irk2Df and siRNA expressing wing discs showing that Dpp requires Irk2 activity. We found that wing specific expression of Irk2DN completely prevents Mad phosphorylation and induces apoptosis. Suppression of apoptosis does not rescue MAD phosphorylation showing that apoptosis is caused by lack of an external signal. We systemically tested the components of the Dpp signaling cascade to find at what point in Dpp signaling Irk2 is required. We found that Irk2 is not directly required for the intracellular propagation of the Dpp signal. Irk2DN could not eliminate the phosphorylation of Mad by a constitutively activated Tkv. We showed that Irk2 affects Dpp spread across the disc. We speculate that Irk2 affects the endocytic pathway that transports Dpp from medial cells to lateral cells. We tested the impact of irk2DN on the development of the Drosophila eye where Irk2 is expressed and Dpp is required for normal patterning. We found that abolition of Irk channels causes eye defects that are similar to those that occur with the loss of Dpp signaling. Trachea development also depends on Dpp. Blocking Irk2 in the developing trachea results in severe defects. We conclude that Irk2 plays a global role in Dpp signaling. Kir/Irk channels could be therapeutic targets to treat diseases that are impacted by TGFβ/BMP signaling, such as cancer. Furthermore, the demonstration that Irk2 is a node for BMP-like signaling could be used to control cell fate decisions for regenerative medicine or stem cell therapeutics.

Degree

PhD

College and Department

Physical and Mathematical Sciences; Chemistry and Biochemistry

Rights

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