Intracellular trafficking of plasmids during transfection is mediated by microtubules.
Little is known about how plasmids move through the cytoplasm to the nucleus. It has been suggested that the dense latticework of the cytoskeleton impedes free diffusion of large macromolecules, including DNA. However, since transfections do work, there must be mechanisms by which DNA circumvents cytoplasmic obstacles. One possibility is that plasmids become cargo on cytoskeletal motors, much like viruses do, and move to the nucleus in a directed fashion. Using microinjection and electroporation approaches in the presence of drugs that alter the dynamics and organization of the cytoskeleton, we show that microtubules are involved in plasmid trafficking to the nucleus. Further, by co-injecting inhibitory antibodies, we find that dynein likely facilitates this movement. These results were confirmed using an in vitro spin-down assay that demonstrated that plasmids bind to microtubules through adaptor proteins provided by cytoplasmic extracts. Taken together, these results suggest that plasmids, like most viruses, utilize the microtubule network and its associated motor proteins to traffic through the cytoplasm to the nucleus.
Vaughan
Erin E
EE
Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.
Dean
David A
DA
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000235:genetics; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D002522:Chlorocebus aethiops; D015536:Down-Regulation / Q000235:genetics; D004398:Dyneins / Q000037:antagonists & inhibitors / Q000235:genetics; D018274:Electroporation; D006801:Humans; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism*; D008845:Microinjections; D008870:Microtubules / Q000187:drug effects / Q000235:genetics*; D017239:Paclitaxel / Q000494:pharmacology; D010957:Plasmids / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D015854:Up-Regulation / Q000235:genetics
D004398:Dyneins; D017239:Paclitaxel