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First published online 19 March 2009
doi: 10.1242/jcs.043034

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Nucleocytoplasmic shuttling of soluble tubulin in mammalian cells

¹ Department of Biochemistry, University of Crete, School of Medicine, 71 003 Heraklion, Greece
² Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology-Hellas, 71 110 Heraklion, Greece
³ Stem Cell and Chromatin Group, The Laboratory of Biology, University of Ioannina School of Medicine, 45 110 Ioannina, Greece
⁴ The Biomedical Institute of Ioannina, IBE/ITE, 45 110 Ioannina, Greece

^* Author for correspondence (e-mail: takis{at}med.uoc.gr)

Accepted 17 December 2008

We have investigated the subcellular distribution and dynamics of soluble tubulin in unperturbed and transfected HeLa cells. Under normal culture conditions, endogenous /β tubulin is confined to the cytoplasm. However, when the soluble pool of subunits is elevated by combined cold-nocodazole treatment and when constitutive nuclear export is inhibited by leptomycin B, tubulin accumulates in the cell nucleus. Transfection assays and FRAP experiments reveal that GFP-tagged β-tubulin shuttles between the cytoplasm and the cell nucleus. Nuclear import seems to occur by passive diffusion, whereas exit from the nucleus appears to rely on nuclear export signals (NESs). Several such motifs can be identified by sequence criteria along the β-tubulin molecule and mutations in one of these (NES-1) cause a significant accumulation in the nuclear compartment. Under these conditions, the cells are arrested in the G0-G1 phase and eventually die, suggesting that soluble tubulin interferes with important nuclear functions. Consistent with this interpretation, soluble tubulin exhibits stoichiometric binding to recombinant, normally modified and hyper-phosphorylated/acetylated histone H3. Tubulin-bound H3 no longer interacts with heterochromatin protein 1 and lamin B receptor, which are known to form a ternary complex under in vitro conditions. Based on these observations, we suggest that nuclear accumulation of soluble tubulin is part of an intrinsic defense mechanism, which tends to limit cell proliferation under pathological conditions. This readily explains why nuclear tubulin has been detected so far only in cancer or in transformed cells, and why accumulation of this protein in the nucleus increases after treatment with chemotherapeutic agents.

Key words: Nuclear export, Tubulin, Histones, β-tubulin, Subcellular distribution

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