Cold Adaptation of Microtubule Assembly and Dynamics

doi:10.1074/jbc.M005699200

Cold Adaptation of Microtubule Assembly and Dynamics

STRUCTURAL INTERPRETATION OF PRIMARY SEQUENCE CHANGES PRESENT IN THE α- AND β-TUBULINS OF ANTARCTIC FISHES*

From the ^‡Department of Biology, Northeastern University, Boston, Massachusetts 02115, the ^¶Department of Molecular Biology, Vanderbilt University, Nashville, Tennessee 37235, and the ^‖Life Science Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720

Abstract

The microtubules of Antarctic fishes, unlike those of homeotherms, assemble at very low temperatures (−1.8 °C). The adaptations that enhance assembly of these microtubules are intrinsic to the tubulin dimer and reduce its critical concentration for polymerization at 0 °C to ∼0.9 mg/ml (Williams, R. C., Jr., Correia, J. J., and DeVries, A. L. (1985)Biochemistry 24, 2790–2798). Here we demonstrate that microtubules formed by pure brain tubulins of Antarctic fishes exhibit slow dynamics at both low (5 °C) and high (25 °C) temperatures; the rates of polymer growth and shortening and the frequencies of interconversion between these states are small relative to those observed for mammalian microtubules (37 °C). To investigate the contribution of tubulin primary sequence variation to the functional properties of the microtubules of Antarctic fishes, we have sequenced brain cDNAs that encode 9 α-tubulins and 4 β-tubulins from the yellowbelly rockcod Notothenia coriiceps and 4 α-tubulins and 2 β-tubulins from the ocellated icefish Chionodraco rastrospinosus. The tubulins of these fishes were found to contain small sets of unique or rare residue substitutions that mapped to the lateral, interprotofilament surfaces or to the interiors of the α- and β-polypeptides; longitudinal interaction surfaces are not altered in the fish tubulins. Four changes (A278T and S287T in α; S280G and A285S in β) were present in the S7-H9 interprotofilament “M” loops of some monomers and would be expected to increase the flexibility of these regions. A fifth lateral substitution specific to the α-chain (M302L or M302F) may increase the hydrophobicity of the interprotofilament interaction. Two hydrophobic substitutions (α:S187A in helix H5 and β:Y202F in sheet S6) may act to stabilize the monomers in conformations favorable to polymerization. We propose that cold adaptation of microtubule assembly in Antarctic fishes has occurred in part by evolutionary restructuring of the lateral surfaces and the cores of the tubulin monomers.

Footnotes

↵* This work was supported by National Science Foundation Grants DPP-8317724, DPP-8614788, DPP-8919004, OPP-9120311, OPP-9420712, and OPP-9815381 (to H. W. D.), by National Institutes of Health Grant GM25638, the Vanderbilt University Research Council (to R. C. W.), and by National Institutes of Health Grant GM46033 (to K. H. D.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵§ To whom correspondence should be addressed: Dept. of Biology, Northeastern University, 414 Mugar Hall, 360 Huntington Ave., Boston, MA 02115. Tel.: 617-373-4495; Fax: 617-373-3724; E-mail: iceman@neu.edu.
Published, JBC Papers in Press, August 23, 2000, DOI 10.1074/jbc.M005699200
↵2 V. Redeker, A. Frankfurter, J. Rossier, S. Parker, and H. W. Detrich, III, manuscript in preparation.
↵3 Two growth rate classes, presumably corresponding to the plus and minus ends of the fish microtubules, were observed at 25 °C (Figs. 1 B and 2 B), but the use of sea urchin flagellar axonemes, which lack the polarity observed for Chlamydomonas axonemes (54, 55), precluded a definitive assessment of the identity of individual ends.
↵4 Expression of the isotypes NcTbα1 and NcTbα4 could not be examined because we lack 3′-UTRs for their cognate cDNAs.
↵5 Note that side chains shown belong to the vertebrate consensus but have been coded per Fig. 3 to indicate the polarity change introduced by the fish substitutions.
↵6 A. S. Davis, personal communication.
Abbreviations:

MAP

microtubule-associated protein

UTR

untranslated region

PIPES

1,4-piperazinediethanesulfonic acid
- Received June 28, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.