Controlling subcellular localization to alter function: Sending oncogenic Bcr-Abl to the nucleus causes apoptosis.
Altering the subcellular localization of signal transducing proteins is a novel approach for therapeutic intervention. Mislocalization of tumor suppressors, oncogenes, or factors involved in apoptosis results in aberrant functioning of these proteins, leading to disease. In the case of chronic myelogenous leukemia (CML), cytoplasmic Bcr-Abl causes oncogenesis/proliferation. On the other hand, nuclear entrapment of endogenous Bcr-Abl (in K562 human leukemia cells) causes apoptosis. The goal of this study was to determine whether ectopically expressed Bcr-Abl could cause apoptosis of K562 cells when specifically directed to the nucleus via strong nuclear localization signals (NLSs). A single NLS from SV40 large T-antigen or four NLSs were subcloned to Bcr-Abl (1NLS-Bcr-Abl or 4NLS-Bcr-Abl). When transfected into K562 cells, only 4NLS-Bcr-Abl translocated to the nucleus. Bcr-Abl alone was found to localize in the cell cytoplasm, colocalizing with actin due to its actin binding domain. 1NLS-Bcr-Abl also localized with actin. Apoptosis induced by 4NLS-Bcr-Abl was evaluated 24h post-transfection by morphologic determination, DNA staining, and caspase-3 assay. This is the first demonstration that altering the location of ectopically expressed Bcr-Abl can kill leukemia cells. Multiple NLSs are required to overcome Bcr-Abl binding to actin, thus driving it into the nucleus and causing apoptosis.
Dixon
Andrew S
AS
Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT 84108, United States.
Kakar
Mudit
M
Schneider
Korbinian M H
KM
Constance
Jonathan E
JE
Paullin
Blake C
BC
Lim
Carol S
CS
D000199:Actins / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism* / Q000502:physiology*; D003001:Cloning, Molecular; D004247:DNA / Q000096:biosynthesis / Q000378:metabolism; D004273:DNA, Neoplasm / Q000235:genetics; D016044:Fusion Proteins, bcr-abl / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D020014:K562 Cells; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000235:genetics; D013347:Subcellular Fractions / Q000378:metabolism* / Q000502:physiology*; D014162:Transfection
D000199:Actins; D004273:DNA, Neoplasm; D004247:DNA; D016044:Fusion Proteins, bcr-abl; D053148:Caspase 3