The function of the ATP-binding cassette (ABC) transporter ABCB1 is not susceptible to actin disruption

Previously we have shown that the activity of the multidrug transporter ABCC1 (multidrug resistance protein 1), and its localization in lipid rafts, depends on cortical actin (Hummel I, Klappe K, Ercan C, Kok JW. Mol. Pharm. 2011 79, 229–40). Here we show that the efflux activity of the ATP-binding cassette (ABC) family member ABCB1 (P-glycoprotein), did not depend on actin, neither in ABCB1 over expressing murine National Institutes of Health (NIH) 3T3 MDR1 G185 cells nor in human SK-N-FI cells, which endogenously express ABCB1. Disruption of the actin cytoskeleton, upon treatment of the cells with latrunculin B or cytochalasin D, caused severe changes in cell and membrane morphology, and concomitant changes in the subcellular distribution of ABCB1, as revealed by confocal laser scanning and electron microscopy. Nevertheless, irrespective of actin perturbation, the cell surface pool of ABCB1 remained unaltered. In NIH 3T3 MDR1 G185 cells, ABCB1 is partly localized in detergent-free lipid rafts, which partitioned in two different density gradient regions, both enriched in cholesterol and sphingolipids. Interestingly, disruption of the actin cytoskeleton did not change the density gradient distribution of ABCB1. Our data demonstrate that the functioning of ABCB1 as an efflux pump does not depend on actin, which is due to its distribution in both cell surface-localized non-raft membrane areas and lipid raft domains, which do not depend on actin stabilization.

Membrane localizations of ABCB1 compared to other ABC transporters and their interactions with the actin cytoskeleton.Five possible membrane pools of an ABC transporter are depicted: 1) non-raft associated and non-actin associated, 2) raft-associated, but non-actin associated, 3) raft-associated and indirectly linked to actin, 4) raft-associated and directly linked to actin, and 5) non-raft associated, but directly linked to actin. PM = plasma membrane and I = integral membrane protein. ABCB1 (P-glycoprotein) appears to be only in pools 1 and 2, while ABCC1 (multidrug resistance protein 1) is also found in pool 3 and ABCC2 (multidrug resistance protein 2) in pool 4; the latter 2 pools are stabilized by actin.Figure optionsDownload high-quality image (69 K)Download as PowerPoint slideHighlights
► Actin disruption causes membrane redistribution of ABCB1 (P-glycoprotein).
► This does not lead to diminished ABCB1 efflux function.
► Actin disruption does not change the cell surface pool of ABCB1.
► Actin disruption does not affect the extent of lipid raft association of ABCB1.
► ABCB1 function remains normal because its membrane environment does not change.