Intermediate Filaments: A Review

275 Pages·1985·12.241 MB·English

by  Professor Dr. Peter Traub (auth.)

Peter Traub
Intermediate Filaments
A Review
Springer-¥erlag
Berlin Heidelberg New York Tokyo
Professor Dr. PETER TRAUB
Max-Planck-Institut
fUr Zellbiologie
Rosenhof
6802 Ladenburg, FRG
With 2 Figures
ISBN-13: 978-3-642-70232-7 e-ISBN-13: 978-3-642-70230-3
DOl: 10.1 007/978-3-642-70230-3
Library of Congress Cataloging in Publication Data. Traub, Peter, 1935- , Intermediate ft1aments.
Bibliography: p. . Includes index. 1. Cytoplasmic ft1aments. 2. Intermediate filament proteins.
I. Title. QH603.C95T73 1985 574.87'34 85-2618
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© by Springer-Verlag Berlin Heidelberg 1985
Softcover reprint of the hardcover I st edition 1985
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Preface
Research on cytoskeletal elements of eukaryotic cells has been expand
ing explosively during the past 5 to 10 years. Due largely to the
employment of electron and immunofluorescent microscopy, significant
results have been obtained which have provided interesting new insights
into the dynamics of nucleated cells at the structural, physiological, as
well as developmental levels. While a substantial amount of knowledge
has accumulated on the function of microfilaments and microtubules,
the roles of the third major class of cytoskeletal structures in vertebrate
cells, the intermediate filaments, have largely resisted clarification. The
investigation of cultured cells and of tissues from various developmental
stages has furnished a host of information on the inter-and intracellular
distribution of the different types of intermediate filaments and led to the
contention that they have a structural and organizing function in the
cytoplasm of vertebrate cells. However, the results of recent experimen
tation have shown that vertebrate cells can function perfectly in the
complete absence of cytoplasmically extended intermediate filament
meshworks. It is legitimate to suppose, therefore, that their function in
vertebrate cells is much more subtle and complex than generally
presumed.
Our interest in the structure and function of intermediate filament
proteins was initiated approximately 7 years ago while working on the
regulation of macromolecular synthesis in picornavirus-infected mam
malian cells. In attempts to demonstrate virus-induced changes in the
nuclear protein components of the host cells, the nonionic detergent
extraction method was used to purify nuclei. To our great surprise, those
isolated from infected and uninfected cells were consistently contami
nated with significant amounts of a cytoplasmic protein with an
apparent molecular weight of 58,000. It was eventually found that we
were actually dealing with the intermediate filament subunit protein
vimentin. Our original studies on the purification and biochemical
characterization of this protein yielded results which, after our finding
that the protein was known and could indeed assemble into intermediate
filaments, led us to develop an entirely new concept of the cellular
function of such material. One of the objectives of this review is to briefly
summarize our experimental results and to introduce an alternative,
unifying hypothesis on how intermediate filaments and their subunit
proteins might act in eukaryotic cells.
It is clear that too much emphasis on either the structural or
biochemical aspects of intermediate filament proteins could lead to
biased views of their cellular function. Thus, considerations only of
VI Preface
electron and immunofluorescent microscopic studies will result in the
development of a structural concept of intermediate filament function,
whereas preferential considerations of the biochemical properties of
their subunit proteins will give rise to the opposite view, and neither will
be directed to the central problem. I would like to point out, therefore,
that I am not so presumptuous as to believe that the novel, alternative
hypothesis included in this review represents the only approach to
elucidating the functional role of intermediate filaments and their
subunit proteins. They may indeed playa dual role or even be multi
functional in the life cycle of eukaryotic cells. In this sense, it is my hope
that our finding of their binding to nucleic acids as well as their noted
high susceptibilities to Ca2 + -dependent post-translational modifications
will give rise to a reinterpretation of the results of the more or less
morphological studies carried out in so many other laboratories. Since
knowledge of the structure of a cellular constituent is one of the
prerequisites for the understanding of its function(s), the elucidation of
the structure of intermediate filament proteins energetically pursued at
present by several research groups will be of great assistance in this
respect.
The last decade has witnessed fundamental advances in intermediate
filament research, with thousands of papers published on their various
aspects. In particular the involvement ofintermediate filaments in many
pathological conditions has been the subject of numerous investigations.
The extensive literature, particularly that in peripheral areas, makes it
impossible to present a complete bibliography of the field. My intention
has been to emphasize those reports, occasionally after reinterpretation
of the experimental results, which might contribute to a better under
standing of the cellular function of intermediate filaments. Although I
have attempted to be as thorough as possible in citing the published work
of many investigators up to December 1983, I am certain that in
advertant errors were made and that some contributions were not
appropriately weighed or not considered adequately in certain sections
of the review. I also wish to apologize for the unintentional omissions of
publications which relate to this area of investigation.
This review was undertaken at the suggestion of Dr. H. G. SCHWEI
GER from our institute. I sincerely appreciate his critical reading of the
manuscript and his initiative in negotiating with the Springer-Verlag,
Heidelberg, on its publication as a monograph. It is my particular wish
to express my deep appreciation to my former coworker Dr. W. J.
NELSON, not only for his enthusiastic cooperation during the com
pilation of the literature in the early stages of this review, but also for his
never-ceasing contribution in ideas, criticisms, and work throughout our
collaborations. Finally, I would like to thank Mrs H. KLEMPP for her
care and patience in preparing the manuscript and the subject index and
to Mrs. B. GERNERT for proofreading the manuscript.
Spring 1985 PETER TRAUB
Contents
1 Introduction.. . . . . . . . . . . . . . . . . 1
2 Distribution of Intermediate Filaments . . . . . . 2
2.1 Intercellular Distribution of Intermediate Filaments. 2
2.1.1 Intermediate Filaments in Early Differentiation 3
2.1.1.1 Murine Embryogenesis 3
2.1.1.2 Chick Embryogenesis. . . . 6
2.1.1.3 Teratocarcinoma Cells . . . 7
2.1.1.4 Epithelial Cell Differentiation 8
2.1.1.5 Myogenesis . . . . . . . . 10
2.1.2 Intermediate Filament Proteins in Differentiated Tissues. 12
2.1.2.1 Cytokeratins 12
2.1.2.2 Desmin. . . . . . . . . . . . . . 13
2.1.2.3 Vimentin . . . . . . . . . . . . . 14
2.1.2.4 Glial Fibrillary Acidic Protein (GFAP) 16
2.1.2.5 Neurofilament Proteins. . . . . 18
2.1.3 Intermediate Filaments in Disease 19
2.1.3.1 Tumor Diagnosis 19
2.1.3.2 Mallory Bodies 21
2.1.3.3 Neuropathies . . 23
2.1.3.4 Autoantibodies . 26
2.1.4 Intermediate Filament Proteins in Evolution . 27
2.1.5 Potential Intermediate Filament Subunit Proteins . 34
2.1.5.1 Synemin . . . 34
2.1.5.2 Paranemin . . 37
2.1.5.3 66 kDa Protein 37
2.1.5.4 68 kDa Protein 38
2.1.5.5 95 kDa Protein 38
2.1.5.6 50 kDa Neurofilament Proteins 39
2.1.5.7 60 to 70 kDa Intermediate Filament-Associated Proteins 39
2.1.6 Intermediate Filament Proteins in Cell Culture . 39
2.1.6.1 Vimentin . . . . . . . . . 41
2.1.6.2 Desmin. . . . . . . . . . 42
2.1.6.3 Glial Fibrillary Acidic Protein 43
2.1.6.4 Neurofilament Proteins . . . 46
2.1.6.5 Cytokeratins . . . . . . . 47
2.2 Intracellular Distribution of Intermediate Filaments and
Their Interaction with Organelles and Proteins. . . . . 51
VIII Contents
2.2.1 Microtubules . 51
2.2.2 Microfilaments 57
2.2.3 Mitochondria . 58
2.2.4 Plasma Membrane 60
2.2.5 Nucleus ..... 63
2.2.6 Endoplasmic Reticulum and Golgi Apparatus 65
2.2.7 Other Cellular Organelles . . . . 65
2.2.8 Proteins and Enzymatic Activities . . . 66
2.2.8.1 Filaggrin ............. . 66
2.2.8.2 Microtubule-Associated Proteins (MAPs) 67
2.2.8.3 Enzymes and Other Proteins. . . . . . 68
2.2.9 Intermediate Filaments in Muscle 71
2.3 Intracellular Reorganization of Intermediate Filament
Systems ......... . 73
2.3.l Mitosis ......... . 74
2.3.2 Microinjection of Antibodies 78
2.3.3 Virus Infection. . . . . . . 80
2.3.4 Receptor-Mediated Endocytosis 83
2.3.5 Drugs, Toxins, and Growth Factors 85
2.3.6 Physical Manipulations . . . . . . 89
2.3.7 Cell Spreading. . . . . . . . . . 90
2.4 In Vivo Assembly of Intermediate Filaments . 91
2.5 Isolation and Subunit Composition of Intermediate
Filaments ................. . 93
3 In Vitro Assembly and Structure of Intermediate
Filaments ...... . 98
3.1 In Vitro Reconstitution. . . . 98
3.1.1 Cytokeratin Filaments . . . . 98
3.1.2 Vimentin and Desmin Filaments 100
3.1.3 Glial Filaments .. 101
3.1.4 Neurofilaments . . . . . . . 102
3.2 Helical Substructure . . . . . 105
3.3 Architecture of Protofilaments . 108
3.3.1 Three-Strand Model of Proto filaments 109
3.3.2 Four-Strand Model of Protofilaments . 111
3.4 Function of the N-Terminal Polypeptide 113
3.5 Structure of Intermediate Filament Subunit Proteins 116
3.5.1 Peptide Mapping. . . . . 116
3.5.2 Amino Acid Composition . 119
3.5.3 Amino Acid Sequence 122
3.5.3.1 Desmin. . . . . . . . . 123
3.5.3.2 Vimentin . . . . . . . . 127
3.5.3.3 Glial Fibrillary Acidic Protein 129
3.5.3.4 Neurofilament Proteins 130
3.5.3.5 Cytokeratins 131
3.5.3.6 Wool a-Keratins. . . 134
Contents IX
3.5.3.7 General Remarks on the Structure oflntermediate
Filament Proteins . . . . . . . . . . . . . . 135
4 Synthesis of Intermediate Filament Proteins in Vitro . . 137
5 Posttranslational Modification of Intermediate Filament
Proteins . . . . . . . . . . . . . . . . . . 140
5.1 Phosphorylation of Intermediate Filament Proteins 141
5.1.1 Neurofilament Proteins 141
5.1.2 Desmin and Vimentin. 144
5.1.3 Cytokeratins . . . . 148
5.1.4 Phosphorylation Sites. 148
5.1.5 Functional Role oflntermediate Filament Protein
Phosphorylation . . . . . . . . . . . . . . .. 149
5.2 Ca2+ -Dependent Proteolysis of Intermediate Filament
Proteins . . . . . . . . . 150
5.2.1 Neurofilament Proteins. . . 150
5.2.2 Glial Fibrillary Acidic Protein 153
5.2.3 Vimentin and Desmin. . . . 153
5.2.4 Cytokeratins . . . . . . . .. 157
5.2.5 Relatedness of Different Ca2+ -Activated Proteinases 157
5.2.6 Putative Site of Action of Ca2+ -Activated Proteinases 161
5.3 Modification of Intermediate Filament Proteins by
Transglutaminases . . . . . . . . . 164
5.3.1 Neurofibrillary Tangles . . . . . . . . . . . . 164
5.3.2 Transglutaminases in Nonneural Cells . . . . . 166
5.3.3 Properties and Putative Site of Action ofTransglutaminases 167
5.3.4 Are Transglutaminases and Ca2+ -Activated Proteinases
Jointly Involved in Receptor-Mediated Endocytosis? .. 168
6 Cellular Function(s) of Intermediate Filaments and Their
Subunit Proteins. . . . . . . . . . . . . . . . . . 170
6.1 Interaction in Vitro of Intermediate Filament Proteins with
Nucleic Acids and Histones . . . . . . . . . 172
6.2 Are Intermediate Filament Proteins Involved in
Information Transfer? . . . . . . . . . . . 178
6.3 Possible Function of Intermediate Filament Proteins in
Nerve Cells . . . . . . . . . . . . . . . . . .. 186
6.4 Possible Function of Intermediate Filament Proteins in
Muscle Cells. . . . . . . . . . . 193
7 Summary and Concluding Remarks 196
References. . 199
Subject Index . 257
List of Abbreviations
BHK baby hamster kidney
CHO Chinese hamster ovary
GFAP glial fibrillary acidic protein
HMG high mobility group
MAP microtubule-associated protein
Mr relative molecular weight
MUGB 4-methylumbelliferyl-p-guanidinobenzoate
NFP neurofilament protein
NPGB p-nitrophenyl-p'-guanidinobenzoate
SDS sodium dodecylsulfate
TLCK Ncx-p-tosyl-L-lysine -chloromethyl ketone
TPCK L-l-tosylamide-2-phenylethyl chloromethyl ketone
1 Introduction
During the past decade, our knowledge and understanding of the structure and
organization of the eukaryotic cytoplasm has increased dramatically. With im
provements in the fIxation and preservation of subcellular structures and the in
troduction of high voltage electron microscopy of whole cells, it has become evi
dent that the cytoplasm is innovated by an intricate and complex meshwork of
filaments. Attempts to isolate this structure have revealed that it is resistant to ex
traction by nonionic detergents and high salt and that it is composed of a discrete
and relatively small number of polypeptides. However, only by the use of mono
specillc antibodies raised against individual proteins, in conjunction with indirect
immunofluorescent microscopy, has it been possible to visualize the individual
filament networks in the light microscope. As a result of these investigations, at
least three cytoplasmic filament systems have been distinguished: microtubules,
microfilaments, and intermediate (10 nm) filaments. Each class of filaments has
been shown to have a characteristic morphology, a discrete polypeptide compo
sition, and specifIc physicochemical properties which distinguish it from the
others. How these three filament systems are associated with the microtrabecular
system is still a matter of debate. This review will concentrate on only one of these
systems, the intermediate (10 nm) filaments and their subunit proteins. The other
filament systems will be discussed only with regard to their possible interactions
with these proteins; for a detailed description of microfilaments and micro
tubules, the reader is referred to the relevant literature.
Several recent reviews on intermediate filaments have discussed the classillca
tion, cellular distribution, and morphology of these protein fIbrils (LAZARIDES
1980, 1982a; ANDERTON 1981; ZACKROFF et al. 1981; STEINERT 1981; FRANKE et
al. 1982e; OSBORN et al. 1982a) and have, as a result, concluded that they play
an, as yet undefIned, structural role in the cytoplasm; they have been postulated
to be involved with microfilaments and microtubules in the construction of the
cytoskeleton. However, recent experimentation on the effects of microinjection of
monospecillc antibodies raised against intermediate filament proteins into living
cells has demonstrated that all the cellular activities previously proposed to be
mediated by these proteins are in fact independent of their cytoplasmic distribu
tion as intermediate filaments. Therefore, it is a particularly auspicious time to
reexamine the properties of these proteins in the light of recent developments. Al
though this review will attempt to concisely summarize the cellular distribution
and the morphological aspects of intermediate filaments with particular attention
to recent results, the main thrust will be a detailed discussion of new biochemical
and molecular biological evidence which indicates that intermediate filament pro
teins may have a different cellular function than that proposed to date.