Controlling subcellular localization to alter function: Sending oncogenic Bcr– Abl to the nucleus causes apoptosis

• Published in: Journal of controlled release 2009-12, Vol.140 (3), p.245-249
• Main Authors: Dixon, Andrew S., Kakar, Mudit, Schneider, Korbinian M.H., Constance, Jonathan E., Paullin, Blake C., Lim, Carol S.
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Apoptosis; Apoptosis - drug effects; Bcr–Abl; Biological and medical sciences; Caspase 3 - metabolism; Cell Nucleus - metabolism; Cell Nucleus - physiology; Cloning, Molecular; CML; DNA - biosynthesis; DNA - metabolism; DNA, Neoplasm - genetics; Fusion Proteins, bcr-abl - administration & dosage; Fusion Proteins, bcr-abl - pharmacology; General pharmacology; Humans; K562 Cells; Medical sciences; Microscopy, Fluorescence; Nuclear localization; Pharmaceutical technology. Pharmaceutical industry; Pharmacology. Drug treatments; Plasmids - genetics; Protein switch; Simian virus 40; Subcellular Fractions - metabolism; Subcellular Fractions - physiology; Transfection
• ISSN: 0168-3659; 1873-4995
• DOI: 10.1016/j.jconrel.2009.06.026

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 24 h 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. Bcr– Abl with 4 nuclear localization signals localizes to the nucleus (middle panel) and causes apoptosis (first and last panels; apoptosis indicated by arrows). [Display omitted]