Changing the Subcellular Location of the Oncoprotein Bcr-Abl Using Rationally Designed Capture Motifs
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Bcr-Abl, the causative agent of chronic myelogenous leukemia (CML), localizes in the cytoplasm where its oncogenic signaling leads to proliferation of cells. If forced into the nucleus Bcr-Abl causes apoptosis. To achieve nuclear translocation, binding domains for capture of Bcr-Abl were generated and attached to proteins with signals destined for the nucleus. These resulting proteins would be capable of binding and translocating endogenous Bcr-Abl to the nucleus.
Bcr-Abl was targeted at 3 distinct domains for capture: by construction of high affinity intracellular antibody domains (iDabs) to regions of Bcr-Abl known to promote cytoplasmic retention, via its coiled coil domain (CC), and through a naturally occurring protein-protein interaction domain (RIN1). These binding domains were then tested for their ability to escort Bcr-Abl into the nucleus using a “protein switch” or attachment of 4 nuclear localization signals (NLSs).
Although RIN1, ABI7-iDab, and CCmut3 constructs all produced similar colocalization with Bcr-Abl, only 4NLS-CCmut3 produced efficient nuclear translocation of Bcr-Abl.
We demonstrate that a small binding domain can be used to control the subcellular localization of Bcr-Abl, which may have implications for CML therapy. Our ultimate future goal is to change the location of critical proteins to alter their function.
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- Changing the Subcellular Location of the Oncoprotein Bcr-Abl Using Rationally Designed Capture Motifs
Volume 29, Issue 4 , pp 1098-1109
- Cover Date
- Print ISSN
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- Springer US
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- chronic myelogenous leukemia
- intracellular domain antibody
- subcellular targeting
- Industry Sectors
- Author Affiliations
- 1. Department of Pharmaceutics and Pharmaceutical Chemistry College of Pharmacy, University of Utah, 421 Wakara Way, Rm. 318, Salt Lake City, Utah, 84108, USA
- 2. Department of Pharmacology and Toxicology, College of Pharmacy, University of Utah, Salt Lake City, Utah, 84108, USA
- 3. Section of Experimental Therapeutics Leeds Institute of Molecular Medicine, St. James’s University Hospital, Leeds, LS9 7TF, UK