Several microtubule-associated proteins (MAPs) have been shown to bind to microtubules via short sequences with repeated amino acids motifs. A microtubule-binding domain has hitherto not been defined for the adult brain microtubule-associated protein 1A (MAP1A). We have searched for a microtubule-binding domain by expressing different protein regions of MAP1A in cultured cell lines using cDNA constructs. One construct included an area with homology to the microtubule-binding domain of MAP1B (Noble et al. (1989) J. Cell Biol. 109, 437–448), but this did not bind to microtubules in transfected cells. Further investigation of other areas of MAP1A revealed a protein domain, capable of autonomously binding to microtubules, which bears no homology to any previously described microtubule-binding sequence. This MAP1A domain is rich in charged amino acids, as are other mammalian microtubule-binding domains, but unlike them has no identifiable sequence repeats. Whereas all previously described mammalian microtubule-binding domains are basic, this novel microtubule-binding domain of MAP1A is acidic. The expression of this polypeptide in cultured cell lines led to a rearrangement of the microtubules in a pattern distinct from that produced by MAP2 or tau, and increased their resistance to treatment with the microtubule depolymerising agent nocodazole. When the MAP1A microtubule-binding domain was co-expressed in cultured cell lines together with MAP2c, the MAP1A microtubule-binding domain was able to bind to the MAP2c-induced microtubule bundles. These results suggest that different microtubule-binding sequences have a common ability to stabilise microtubules but differ in their influence on microtubule arrangement in the cell. This may be significant in neurons, where microtubule-associated proteins with different microtubule-binding sequences are expressed in different cell compartments and at different times during development.

REFERENCES

Adams
M. D.
,
Kelley
J. M.
,
Gocayne
J. D.
,
Dubnick
M.
,
Polymeropoulos
M. H.
,
Xiao
H.
,
Merril
C. R.
,
Wu
A.
,
Olde
B.
,
Moreno
R. F.
,
Kerlavage
A. R.
,
McCombie
W. R.
,
Venter
J.
(
1991
).
Complementary DNA sequencing: expressed sequence tags and human genome project.
Science
252
,
1651
1656
Ahmad
F. J.
,
Pienkowski
T. P.
,
Baas
P. W.
(
1993
).
Regional differences in microtubule dynamics in the axon.
J. Neurosci
13
,
856
866
Aizawa
H.
,
Emori
Y.
,
Murofushi
H.
,
Kawasaki
H.
,
Sakai
H.
,
Suzuki
K.
(
1990
).
Molecular cloning of a ubiquitously distributed microtubule-associated protein with M r190,000.
J. Biol. Chem
265
,
13849
13855
Aizawa
H.
,
Emori
Y.
,
Mori
A.
,
Murofushi
H.
,
Sakai
H.
,
Suzuki
K.
(
1991
).
Functional analyses of the domain structure of microtubule-associated protein-4 (MAP-U).
J. Biol. Chem
266
,
9841
9846
Baas
P. W.
,
Black
M. M.
(
1990
).
Individual microtubules in the axon consist of domains that differ in both composition and stability.
J. Cell Biol
111
,
495
509
Baas
P. W.
,
Ahmad
F. J.
(
1992
).
The plus ends of stable microtubules are the exclusive nucleating structures for microtubules in the axon.
J. Cell Biol
116
,
1231
1241
Baas
P. W.
,
Ahmad
F. J.
,
Pienkowski
T. P.
,
Brown
A.
,
Black
M. M.
(
1993
).
Sites of microtubule stabilization for the axon.
J. Neurosci
13
,
2177
2185
Bamburg
J. R.
,
Bray
D.
,
Chapman
K.
(
1986
).
Assembly of microtubules at the tip of growing axons.
Nature
321
,
788
790
Chapin
S. J.
,
Bulinski
J. C.
(
1991
).
Non-neuronal 21010(3) M rmicrotubule-associated protein (MAP4) contains a domain homologous to the microtubule-binding domains of neuronal MAP2 and tau.
J. Cell Sci
98
,
27
36
Chen
C.
,
Okayama
H.
(
1987
).
High efficiency transformation of mammalian cells by plasmid DNA.
Mol. Cell. Biol
7
,
2745
2752
Chou
P. Y.
,
Fasman
G. D.
(
1978
).
Prediction in the secondary structure of proteins from their amino acid sequence.
Advan. Enzymol
47
,
45
147
Evan
G. I.
,
Lewis
G. K.
,
Ramsay
G.
,
Bishop
J. M.
(
1985
).
Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product.
Mol. Cell. Biol
5
,
3610
3616
Faivre
C.
,
Legrand
C.
,
Rabie
A.
(
1985
).
The microtubular apparatus of cerebelar Purkinje cells during postnatal development of the rat: The density and cold-stability of microtubules increase with age and are sensitive to thyroid hormone.
Int. J. Dev. Neurosci
3
,
559
565
Fellous
A.
,
Lennon
A. M.
,
Francon
J.
,
Nunez
J.
(
1979
).
Thyroid hormones and neurotubule assembly in vitro during brain development.
Eur. J. Biochem
101
,
365
376
Francon
J.
,
Lennon
A. M.
,
Fellous
A.
,
Mareck
A.
,
Pierre
M.
,
Nunez
J.
(
1982
).
Heterogeneity of microtubule-associated proteins and brain development.
Eur. J. Biochem
129
,
465
472
Friederich
E.
,
Vancompernolle
K.
,
Huet
C.
,
Goethals
M.
,
Finidori
J.
,
Vandekerckhove
J.
,
Louvard
D.
(
1992
).
An actin-binding site containing a conserved motif of charged amino acid residues is essential for the morphogenic effect of villin.
Cell
70
,
81
92
Garner
C. C.
,
Garner
A.
,
Huber
G.
,
Kozak
C.
,
Matus
A.
(
1990
).
Molecular cloning of microtubule-associated protein 1 (MAP1A) and microtubule-associated protein 5 (MAP1B): identification of distinct genes and their differential expression in developing brain.
J. Neurochem
55
,
146
154
Garnier
J.
,
Osguthorpe
D. J.
,
Robson
B.
(
1978
).
Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins.
J. Mol. Biol
120
,
97
120
Glisin
V.
,
Crkvenjakov
R.
,
Byus
C.
(
1974
).
Ribonucleic acid isolated by cesium chloride centrifugation.
Biochemistry
13
,
2633
2643
Goedert
M.
,
Crowther
R. A.
,
Garner
C. C.
(
1991
).
Molecular characterization of microtubule-associated proteins Tau and MAP2.
Trends Neurosci
14
,
193
199
Goldstein
L. S.
,
Laymon
R. A.
,
McIntosh
J. R.
(
1986
).
A microtubule-associated protein in Drosophila melanogaster: identification, characterization, and isolation of coding sequences.
J. Cell Biol
102
,
2076
2087
Gordon-Weeks
P. R.
(
1991
).
Microtubule organization in growth cones.
Biochem. Soc. Trans
19
,
1080
1085
Hammarback
J. A.
,
Obar
R. A.
,
Hughes
S. M.
,
Vallee
R. B.
(
1991
).
MAP1B is encoded as a polyprotein that is processed to form a complex N-terminal microtubule-binding domain.
Neuron
7
,
129
139
Hemphill
A.
,
Seebeck
T.
,
Lawson
D.
(
1991
).
The Trypanosoma brucei cytoskeleton: ultrastructure and localization of microtubule-associated and spectrin-like proteins using quick-freeze, deep-etch, immunogold electron microscopy.
J. Struct. Biol
107
,
211
220
Hemphill
A.
,
Affolter
M.
,
Seebeck
T.
(
1992
).
A novel microtubule-binding motif identified in a high molecular weight microtubule-associated protein from Trypanosoma brucei.
J. Cell Biol
117
,
95
103
Hirokawa
N.
,
Shiomura
Y.
,
Okabe
S.
(
1988
).
Tau proteins: the molecular structure and mode of binding on microtubules.
J. Cell Biol
107
,
1449
1459
Irminger-Finger
I.
,
Laymon
R. A.
,
Goldstein
L. S.
(
1990
).
Analysis of the primary sequence and microtubule-binding region of the Drosophila 205K MAP.
J. Cell Biol
111
,
2563
2572
Jacobs
J. R.
,
Stevens
J. K.
(
1986
).
Experimental modification of PC12 neurite shape with the microtubule-depolymerising drug nocodazole: a serial electron microscopic study of neurite shape control.
J. Cell Biol
103
,
907
915
Kanai
Y.
,
Takemura
R.
,
Oshima
T.
,
Mori
H.
,
Ihara
Y.
,
Yanagisawa
M.
,
Masaki
T.
,
Hirokawa
N.
(
1989
).
Expression of multiple tau isoforms and microtubule bundle formation in fibroblasts transfected with a single tau cDNA.
J. Cell Biol
109
,
1173
1184
Kanai
Y.
,
Chen
J.
,
Hirokawa
N.
(
1992
).
Microtubule bundling by tau proteins in vivo: analysis of functional domains.
EMBO J
11
,
3953
3961
Kuznetsov
S. A.
,
Rodionov
V. I.
,
Nadezhdina
E. S.
,
Murphy
D. B.
,
Gelfand
V. I.
(
1986
).
Identification of a 34-kD polypeptide as a ligth chain of microtubule-assciated protein 1 (MAP1) and its association with a MAP1 peptide that binds to microtubules.
J. Cell Biol
102
,
1060
1066
Langkopf
A.
,
Hammarback
J. A.
,
Muller
R.
,
Vallee
R. B.
,
Garner
C. C.
(
1992
).
Microtubule-associated proteins 1A and LC2. Two proteins encoded in one messenger RNA.
J. Biol. Chem
267
,
16561
16566
Lee
G.
,
Cowan
N.
,
Kirschner
M.
(
1988
).
The primary structure and heterogeneity of tau protein from mouse brain.
Science
239
,
285
288
Lee
G.
,
Rook
S. L.
(
1992
).
Expression of tau protein in non-neuronal cells: microtubule binding and stabilization.
J. Cell Sci
102
,
227
237
Lee
J. C.
,
Field
D. J.
,
Lee
L. L. Y.
(
1980
).
Effects of nocodazole on structures of calf brain tubulin.
Biochemistry
19
,
6209
6215
Lennon
A. M.
,
Francon
J.
,
Fellous
A.
,
Nunez
J.
(
1980
).
Rat, mouse and guinea-pig brain development and microtubule assembly.
J. Neurochem
35
,
804
813
Lewis
S. A.
,
Wang
D. H.
,
Cowan
N. J.
(
1988
).
Microtubule-associated protein MAP2 shares a microtubule binding motif with tau protein.
Science
242
,
936
939
Lewis
S. A.
,
Ivanov
I. E.
,
Lee
G. H.
,
Cowan
N. J.
(
1989
).
Organization of microtubules in dendrites and axons is determined by a short hydrophobic zipper in microtubule-associated proteins MAP2 and tau.
Nature
342
,
498
505
Lewis
S. A.
,
Cowan
N. J.
(
1990
).
Microtubule bundling.
Nature
345
,
674
–.
Littauer
U. Z.
,
Giveon
D.
,
Thierauf
M.
,
Ginzburg
I.
,
Ponstingl
H.
(
1986
).
Common and distinct tubulin binding sites for microtubule-associated proteins.
Proc. Nat. Acad. Sci. USA
83
,
7162
7166
Lundberg
K. S.
,
Shoemaker
D. D.
,
Adams
M. W.
,
Short
J. M.
,
Sorge
J. A.
,
Mathur
E. J.
(
1991
).
High-fidelity amplification using a thermostable DNA polymerase isolated from Pyrococcus furiosus.
Gene
108
,
1
6
Matus
A.
,
Riederer
B.
(
1986
).
Microtubule-associated proteins in the developing brain.
Ann. NY Acad. Sci
466
,
167
179
Matus
A.
(
1988
).
Microtubule-associated proteins: their potential role in determining neuronal morphology.
Annu. Rev. Neurosci
11
,
29
44
Meichsner
M.
,
Doll
T.
,
Reddy
D.
,
Weisshaar
B.
,
Matus
A.
(
1993
).
The low molecular weight form of microtubule-associated protein 2 is transported into both axons and dendrites.
Neuroscience
54
,
873
880
Mitchison
T.
,
Kirschner
M.
(
1988
).
Cytoskeletal dynamics and nerve growth.
Neuron
1
,
761
772
Munro
S.
,
Pelham
H. R.
(
1987
).
A C-terminal signal prevents secretion of luminal ER proteins.
Cell
48
,
899
907
Ngsee
J. K.
,
Scheller
R. H.
(
1989
).
Isolation and characterization of two homologous cDNA clones from Torpedo electromotor neurons.
DNA
8
,
555
561
Noble
M.
,
Lewis
S. A.
,
Cowan
N. J.
(
1989
).
The microtubule binding domain of microtubule-associated protein MAP1B contains a repeated sequence motif unrelated to that of MAP2 and tau.
J. Cell Biol
109
,
437
448
Paschal
B. M.
,
Obor
R. A.
,
Vallee
R. B.
(
1989
).
Interaction of brain cytoplasmic dynein and MAP2 with a comon sequence at the C terminus of tubulin.
Nature
342
,
569
572
Pedrotti
B.
,
Soffientini
A.
,
Islam
K.
(
1993
).
Sulphonate buffers affect the recovery of microtubule-associated proteins MAP1 and MAP2: evidence that MAP1A promotes microtubule assembly.
Cell Motil. Cytoskel
25
,
234
242
Pelham
H. R. B.
(
1993
).
Is epimorphin involved in vesicular transport?.
Cell
73
,
425
426
Riederer
B.
,
Matus
A.
(
1985
).
Differential expression of distinct microtubule-associated proteins during brain development.
Proc. Nat. Acad. Sci. USA
82
,
6006
6009
Safer
D.
,
Elzinga
M.
,
Nachmias
V. T.
(
1991
).
Thymosin4 and Fx, an actin-sequestering peptide, are indistinguishable.
J. Biol. Chem
266
,
4029
4032
Sanger
F.
,
Nicklen
S.
,
Coulson
A. R.
(
1977
).
DNA sequencing with chain-terminating inhibitors.
Proc. Nat. Acad. Sci. USA
74
,
5463
5467
Schneider
A.
,
Hemphill
A.
,
Wyler
T.
,
Seebeck
T.
(
1988
).
Large microtubule-associated protein of T. brucei has tandemly repeated, near-identical sequences.
Science
241
,
459
462
Schoenfeld
T. A.
,
McKerracher
L.
,
Obar
R.
,
Vallee
R. B.
(
1989
).
MAP 1A and MAP 1B are structurally related microtubule associated proteins with distinct developmental patterns in the CNS.
J. Neurosci
9
,
1712
1730
Serrano
L.
,
Avila
J.
,
Maccioni
R. B.
(
1984
).
Controlled proteolysis of tubulin by subtilisin: localization of the site for MAP2 interaction.
Biochemistry
23
,
4675
4681
Serrano
L.
,
de la Torre
J.
,
Maccioni
R. B.
,
Avila
J.
(
1984
).
Involvement of the carboxyl-terminal domain of tubulin in the regulation of its assembly.
Proc. Nat. Acad. Sci. USA
81
,
5989
5993
Schulze
E.
,
Asai
D. J.
,
Bulinski
J. C.
,
Kirschner
M.
(
1987
).
Posttranslational modification and microtubule stability.
J. Cell Biol
105
,
2167
2177
Schulze
E.
,
Kirschner
M.
(
1987
).
Dynamic and stable populations of microtubules in cells.
J. Cell Biol
104
,
277
288
Sherwin
T.
,
Gull
K.
(
1989
).
Visualisation of detyrosination along single microtubules reveals novel mechanisms of assembly during cytoskeletal duplication in trypanosomes.
Cell
57
,
211
221
Shiomura
Y.
,
Hirokawa
N.
(
1987
).
The molecular structure of microtubule-associated protein 1A (MAP1A) in vivo and in vitro. An immunoelectron microscopy and quick-freeze, deep-etch study.
J. Neurosci
7
,
1461
1469
Small
J. V.
(
1988
).
The actin cytoskeleton.
Electron Microsc. Rev.
1
,
155
174
Solomon
F.
,
Magendantz
M.
,
Salzman
A.
(
1979
).
Identification with cellular microtubules of one of the co-assembling microtubule-associated proteins.
Cell
18
,
431
438
Takemura
R.
,
Okabe
S.
,
Umeyama
T.
,
Kanai
Y.
,
Cowan
N. J.
,
Hirokawa
N.
(
1992
).
Increased microtubule stability and alpha tubulin acetylation in cells transfected with microtubule-associated proteins MAP1B, MAP2 or tau.
J. Cell Sci
103
,
953
964
Umeyama
T.
,
Okabe
S.
,
Kanai
Y.
,
Hirokawa
N.
(
1993
).
Dynamics of microtubules bundled by microtubule associated protein 2c (MAP2c).
J. Cell Biol
120
,
451
465
Vallee
R. B.
(
1982
).
A taxol-dependent procedure for the isolation of microtubules and microtubule-associated proteins (MAPs).
J. Cell Biol
92
,
435
442
Vallee
R. B.
,
Davis
S. E.
(
1983
).
Low molecular weight microtubule-associated proteins are light chains of microtubule-associated protein 1 (MAP 1).
Proc. Nat. Acad. Sci. USA
80
,
1342
1346
Weisshaar
B.
,
Doll
T.
,
Matus
A.
(
1992
).
Reorganization of the microtubular cytoskeleton by embryonic microtubule-associated protein 2 (MAP2c).
Development
116
,
1151
1161
West
R. R.
,
Tenbarge
K. M.
,
Olmsted
J. B.
(
1991
).
A model for microtubule-associated protein 4 structure. Domains defined by comparisons of human, mouse, and bovine sequences.
J. Biol. Chem
266
,
21886
21896
Yamamoto
K.
(
1991
).
Identification of the site important for the actin-activated MgATPase activity of myosin subfragment-1.
J. Mol. Biol
217
,
229
233
Yonezawa
N.
,
Nishida
E.
,
Ohba
M.
,
Seki
M.
,
Kumagai
H.
,
Sakai
H.
(
1989
).
An actin-interacting heptapeptide in the cofilin sequence.
Eur. J. Biochem
183
,
235
238
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