Institut für Biochemie I, Medizinische Fakultät,
Universität zu Köln, Joseph-Stelzmann-Strasse 52, D-50931 Köln,
Germany
* Present address: Abteilung Biophysikalische Chemie, OE 4350, Medizinische
Hochschule Hannover, Carl-Neuberg-Str. 1, D-30625, Hannover, Germany
Author for correspondence (e-mail:
francisco.rivero{at}uni-koeln.de)
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Introduction |
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On the basis of the degree of sequence similarity, the CH domains can be
divided into several groups. Three major classes are represented by the
ABD-forming CH domains (CH1 and CH2) and by the CH3 domains characteristic of
most 1xCH proteins. The CH domains of fimbrins form four further groups,
which we designate as CHf1, CHf2, CHf3 and CHf4, starting with the N-terminal
domain. Using the nomenclature of Gimona et al., they correspond to CH1.1,
CH2.1, CH1.2 and CH2.2, respectively
(Gimona et al., 2002).
Typically, the CH domains that are present as a single copy (CH3) are also
functionally distinct from those found in tandem pairs and in 4xCH
proteins. Likewise, the N-terminal CH domains in tandem pairs (CH1) differ
from the C-terminal domains (CH2) (Banuelos
et al., 1998
; Gimona et al.,
2002
). An isolated CH1 domain is able to bind to actin, but a
tandem pair of CH domains is required for a fully functional ABD. The globular
fold of CH domains is built by four core helices, three of them forming a
loose triple helix bundle, and by one to three short helices present in the
loops between the core helices. Comparison of the structures of the CH domains
of fimbrin, spectrin, dystrophin, utrophin and calponin shows overall
conservation of the tertiary fold, but reveals substantial differences between
them that correlate with the domain classification outlined above
(Bramham et al., 2002
).
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1xCH proteins |
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2xCH proteins |
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The diversity of ABD-containing proteins arose through gene duplication
events followed by shuffling and intragenic multiplication
(Dubreuil, 1991).
-actinin, which is present in lower eukaryotes, is the prototype that
gave rise to spectrin and dystrophin and probably also NUANCE/enaptin and
plakins (Leung et al., 2001
) in
higher eukaryotes by multiplication of spectrin repeats.
Dictyostelium filamin is the prototype that gave rise to filamins of
animal species by multiplication of filamin repeats. Cortexillins and
interaptin appear to be exclusive acquisitions of Dictyostelium, as
are calmin and NUANCE/enaptin in mammals.
Parvin/actopaxin constitutes an exception to the `CH1+CH2' rule for a single ABD. Both CH domains of parvin/actopaxin are more closely related to the CH1 domain than to any other class, and yet they diverge from the CH1 domain of the ABD. Sequence analysis suggests that these proteins arose by a duplication of the CH domain that was independent of that which gave rise to the ABD. This duplication probably took place after the branching of metazoa, because parvins are documented only in animal species. Parvins are also exceptional in that the CH domains do not appear in combination with other known domains.
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3xCH and 4xCH proteins |
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A new combination of CH domains was found in the C. elegans 4xCH protein AAK71387, where the N-terminal CH1+CH2 tandem was followed by two further domains of the CH2 type. These two additional domains probably result from a duplication of the first CH2 domain. Finally, the most intriguing was the finding of the 3xCH proteins. The CH1 and CH2 domains are followed by a highly divergent CH domain in the filamin-related proteins DmAAF46896 and CeAAB37032. No mammalian homologue of the 3xCH proteins has been found so far.
The general role of CH domains is largely unclear. Originally thought to be
an actin-binding unit, the CH3 domain fails to interact with F-actin
(Fu et al., 2000;
Gimona and Mital, 1998
),
although its structure can be fitted into a calponin-decorated F-actin image
reconstruction in an utrophin-like mode
(Bramham et al., 2002
). The
actin-binding property was reported to be an exclusive feature of the CH1
domain and, to a lesser extent, of the CH2 domain of the `CH1+CH2' proteins
(Winder et al., 1995
). It is
not yet known whether the CHa, CHc, CHe, CH2 and CHf2 domains that occur in
single-copy CH proteins retain their actin-binding properties. Various CH
domains have also been found to harbour interaction sites for
phosphatidylinositol (4,5)-bisphosphate, integrin ß4, vimentin,
calmodulin, paxillin, integrin-linked kinase (ILK) and the extracellular
signal regulated kinase (ERK). However, only ERK has been identified as a
common ligand for the CH3 domain of calponin and the ABD of
-actinin so
far (Leinweber et al., 1999
),
which points to a potential role for CH domains as targets in signalling
pathways. A common function for the CH domains of different types remains to
be uncovered.
For the phylogenetic analysis of CH domains we have considered representative organisms whose genomes have been fully sequenced (Arabdidopsis thaliana, Saccharomyces cerevisiae and Schizosaccharomyces pombe, Caenorhabditis elegans, Drosophila melanogaster, Homo sapiens) or where sequencing is well advanced (Dictyostelium discoideum). We initially screened the protein databases of these organisms for proteins containing CH domains through the SMART server (http://smart.embl-heidelberg.de/). A preliminary classification on the basis of domain structure was generated. For proteins annotated as hypothetical or predicted that could not be classified into any of the known families, the corresponding EST database was screened to verify that their existence was supported by cDNA sequences. The species distribution of each subfamily was investigated by extensive screening of the non-redundant and EST databases using the BLAST and TBLASTN tools. Our catalog of proteins carrying CH domains is extensive but probably not complete. Less conserved CH domains might not have been identified and true proteins annotated as hypothetical might have been rejected owing to the absence of supporting cDNA sequences. Primary sequences of the CH domains were aligned using the ClustalW program, and the alignment was manually edited using BioEdit. Usually only one isoform of each protein family in every species was taken for the alignment. Some 2xCH proteins (dystrophin and some members of the plakin family are prominent examples) undergo alternative splicing, yielding isoforms lacking one or both CH domains. To avoid complexity, splice variants have not been considered in this study, and the longest transcribed variant is depicted in the poster. In very few cases a sequence was rejected when it was too divergent for a reliable alignment; an example is C. elegans dystrophin. The phylogenetic tree was constructed using the neighbour-joining method. For unnamed proteins, sequence names are composed of the species initials and NCBI accession number. We have conserved the domain nomenclature of the SMART server, where information about the domains can be retrieved. Exceptions are DH (Dbl-homology domain, equivalent to RhoGEF), CLR (calponin-like repeats), GSR (Gly-Ser-Arg repeats), TM (transmembrane region) and FB3 (FAD-binding domain type 3). Non-standard abbreviations for some proteins are LRNP (leucin-rich neuronal protein) and LMO (LIM only 7).
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References |
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Bramham, J., Hodgkinson, J. L., Smith, B. O., Uhrin, D., Barlow, P. N. and Winder, S. J. (2002). Solution structure of the calponin CH domain and fitting to the 3D-helical reconstruction of F-actin:calponin. Structure 10,249 -258.[CrossRef][Medline]
Dubreuil, R. R. (1991). Structure and evolution of the actin crosslinking proteins. Bioessays 13,219 -226.[Medline]
Fu, Y., Liu, H. W., Forsythe, S. M., Kogut, P., McConville, J.
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Gimona, M. and Mital, R. (1998). The single CH
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