1 Max Planck Institute for Biophysical Chemistry, Department of Biochemistry, Am
Fassberg 11, D-37077 Goettingen, Germany
2 Department of Cell Biology, University of Bremen, Leobener Strasse NW II,
D-28359 Bremen, Germany
* Author for correspondence (e-mail: office.weber{at}mpibpc.gwdg.de)
Accepted 24 February 2003
![]() |
Summary |
---|
![]() ![]() ![]() ![]() ![]() |
---|
Key words: Fugu rubripes, Gene clusters, Intermediate filaments, Keratin, Lamin, Neurofilament, Vertebrate genomes
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() |
---|
Point mutations in a still-growing number of IF genes are connected with
human diseases. Mutations in at least 14 epidermal keratin genes cause
fragility syndromes of the skin (Irvine
and McLean, 1999) and similar mutations in the type III desmin
gene connect to myopathies of heart and skeletal muscle
(Goldfarb et al., 1998
),
whereas mutations in the GFAP gene are found in Alexander's disease
(Brenner et al., 2001
;
Li et al., 2002
). Finally, in
Caenorhabditis elegans, at least four of the 11 IF genes are
essential for nematode development
(Karabinos et al., 2001
). Type
I-III genes are not restricted to vertebrates but have also been documented in
the early chordates, which, however, seem to lack type IV genes (reviewed in
Karabinos et al., 2002
;
Wang et al., 2002
).
Some genes for type I-IV cytoplasmic IF proteins from fish have previously
been documented by cDNA cloning in particular in the goldfish and the rainbow
trout (Markl and Schechter,
1998; Schaffeld et al.,
2002a
,b
),
and nuclear lamins have been analysed in the goldfish
(Yamaguchi et al., 2001
) and
the zebrafish (Hofemeister et al.,
2002
). However, only the emerging genome of the teleost fish
Fugu rubripes (Aparicio et al.,
2002
) allows a detailed comparison of IF gene organization and
complexity in man and a lower vertebrate. Here, we report on some unexpected
differences between IF genes in F. rubripes and mammals.
![]() |
Results and Discussion |
---|
![]() ![]() ![]() ![]() ![]() |
---|
|
|
The F. rubripes database
(Aparicio et al., 2002) is very
reliable and well organized. In a few cases we needed to introduce a frame
shift to keep the obvious reading frame or to explore a major change from the
proposed gene structure (Table
1). Two expected difficulties arose: the occasional presence of
sequence gaps in some genes situated in the interior of a scaffold and the
location of a gene at the end of a scaffold
(Table 1). The first problem
can be solved directly by PCR amplification bridging the gaps between the
known neighboring sequences. The second set of problems, which relates to
eight cytoplasmic IF genes, requires overlaps for the more than 12,000
scaffolds, which should be supplied in the future by the F. rubripes
genome sequencing consortium (Aparicio et
al., 2002
).
Striking excess of type I over type II keratin genes in F.
rubripes
Tables 1 and
2 show the presence of 13
complete and six incomplete F. rubripes type I genes. The total of 19
genes surpasses the 16 human type I genes (not including the nine type I hair
keratin genes thought to be a mammalian specialization). An entirely different
situation is given by the type II keratin genes because we located only four
complete and two nearly complete genes. Thus, there are about three times as
many type I than type II genes in F. rubripes, whereas, in humans,
the numbers of type I and II genes is similar (Tables
1,
2). The large excess of type I
over type II genes could indicate that functional differences between the
obligatory heteropolymeric keratin filaments of different cell types depend
primarily on the type II genes that are expressed, whereas the type I genes
provide additional variability.
Most F. rubripes keratin genes show the intron patterns previously
described for mammalian type I and type II genes. The two complete keratin I
genes on scaffold 2605 have an additional intron between the traditional
introns 5 and 6. The keratin II gene on scaffold 3830 has another novel intron
position situated between introns 6 and 7. A striking case of an unusual
intron pattern is observed in the keratin I gene on scaffold 7354. It has the
normal type I intron pattern but, in addition, has an intron that occurs in
all mammalian type II and in all F. rubripes type II genes (intron 1
of type II genes). The keratin I gene on scaffold 135 shows an unusual
doubling of exon 6 (protein sequence identity 95%), which encodes the
C-terminal end of the rod domain. Possibly, these exons are alternatives. The
type I gene situated at the end of scaffold 8680
(Fig. 1;
Table 1) is incomplete and
lacks the 5' end. Interestingly, it is the only F. rubripes IF
gene that shows several gaps in alignments of the predicted protein sequence
and might be a rare pseudogene. Finally the single keratin I genes on the two
small scaffolds 7320 and 8762 (Table
1) share 96.8% sequence identity on the nucleotide level including
the six introns. This observation is clearly unrelated to the often suggested
partial tetraploidity of fish genomes
(Aparicio et al., 2002) and
instead signals a very recent gene duplication event.
|
Lack of the keratin II gene cluster in F. rubripes
The human genome contains all 25 type I keratin genes except for the
keratin 18 gene in a cluster on chromosome 17q21, where they are arranged in
the same orientation. Similarly, all 24 type II keratin genes are in a similar
cluster on chromosome 12q13, which also harbors the keratin 18 gene next to
the keratin 8 gene (Hesse et al.,
2001). Although the more than 12,000 scaffolds are not yet
arranged as continuous F. rubripes genome sequence, the current
mapping results (Table 1,
Fig. 1) suggest that keratin
genes are differently organized in the F. rubripes and the mammalian
genomes.
Fig. 1 shows that four of the six type II genes locate as either paired (scaffold 214) or single (scaffolds 2158 and 3159) genes next to either one or two type I genes. The other two type II genes map to two separate scaffolds (285 and 3830). Thus, the presence of a single keratin II gene cluster as in humans is excluded. Interestingly, when type I and II genes map together, they show different orientations.
There is already clear evidence for some clustering of keratin I genes in F. rubripes. Fig. 1 shows four groups of two and three directly neighboring keratin I genes, and a pair of keratin I genes separated only by a hypothetical gene. Because another four keratin I genes lie on rather small scaffolds, one could envision a single cluster of many type I genes in F. rubripes. However, it seems not to be possible to build a cluster that collects, as in humans, all type I genes without the simultaneous incorporation of four type II genes and various unrelated genes. Neighboring type I genes can have either the same or opposite orientation.
The emerging differences of keratin gene locations in mammalian and fish
genomes argue that the keratin gene clustering as documented for mammals was
acquired after the fish lineage separated from the lineage leading to higher
vertebrates. Forthcoming genomic data on the amphibian Xenopus
tropicalis and the chicken (as a representative of the birds) will shed
light on the question when keratin gene clustering was acquired during
vertebrate evolution. We also note that a recent phylogenetic analysis of
keratin I and II proteins indicates that fish epidermal keratins diversified
independently from the mammalian epidermal keratin radiation, keratins 8 and
18 of interior epithelia are true orthologs in fish and mammals
(Schaffeld et al., 2002a).
Mammalian keratins 8 and 18 are typical of internal epithelia and represent
the earliest keratin expression pair in embryogenesis. Interestingly the gene
for keratin 18, a type I keratin, is adjacent to the keratin 8 gene in the
type II gene cluster on human chromosome 12q13
(Waseem et al., 1990;
Hesse et al., 2001
). The close
proximity of keratin 8 and 18 genes also seems to hold for F.
rubripes. The type II gene on scaffold 3159 codes for a keratin 8 (92%
and 78% sequence identity with the rod domain of keratins 8 from rainbow trout
and human, respectively). The keratin 8 gene is separated by a very small
hypothetical gene from a type I keratin gene whose sequence is still not
complete (Fig. 1). BLAST
analysis registers the predicted protein as keratin 18 (81% and 64% identity
of the available rod sequence with keratins 18 from rainbow trout and human,
respectively). Fig. 2 gives
some examples of the sequence similarity of F. rubripes and human IF
proteins.
|
Duplication of the type III desmin gene
Previous cDNA cloning studies established in fish the homologs of the four
mammalian type III genes encoding vimentin, desmin, GFAP and peripherin [fish
peripherin is often referred to as plasticin in the literature]
(Markl and Schechter, 1998).
The four complete genes are also present in F. rubripes, which shows
two additional type III genes (Table
1). The fifth type III gene arises by joining scaffolds 3504 and
4275, with the assumption that the scaffold ends protrude into the first
intron of the predicted desmin 2 gene. The two gene fragments predict 70-80%
sequence identity for desmins 1 and 2. The desmin 2 gene has, in addition to
the eight typical type III introns, a further novel intron situated between
the traditional introns 5 and 6. Using PCR amplification on F.
rubripes DNA and sequence analysis, we have verified the proposed
arrangement of the two scaffolds covering the desmin 2 gene.
The sixth gene (scaffold 117) has some unexpected features. It is
intronless and lies in the large tenth intron (3.2 kb) of a gene encoding the
enzyme isoleucine tRNA synthase. The open reading frame predicts a second
vimentin that shares 43% sequence identity with vimentin 1. The canonical
sequence motif LNDR in coil 1a is changed to LNAK in vimentin 2. Currently, we
do not know whether vimentin 2 is an active gene. The lack of a polyA tract
argues against its being a processed pseudogene. The presence of two desmin
and vimentin genes in F. rubripes and the previous finding of at
least two vimentin genes in Xenopus laevis
(Herrmann et al., 1989
), which
is a tetraploid species, open the possibility that only higher vertebrates
have single vimentin and desmin genes.
The beaded filaments of the mammalian and avian eye lens contain the two
special cytoplasmic IF proteins, phakinin (CP49)
(Hess et al., 1996) and
filensin (Masaki and Quinlan,
1997
), which together form the BF subfamily. The corresponding
genes in F. rubripes locate to contig 29247 and scaffold 91,
respectively (Table 1). The
F. rubripes phakinin gene lacks the 5' end and the consensus
sequence at the end of the rod domain of the phakinin protein is, as in other
phakinins, changed from YRKLLEGE to YHGILDGE
(Sandilands et al., 1998
). The
F. rubripes filensin gene has still a small sequence gap.
Surprisingly many type IV genes
The seven mammalian type IV genes
(Lewis and Cowan, 1986) show
an entirely different organization than do type I-III genes
(Tyner et al., 1985
)
(Fig. 3). They have only two
introns (three for NF-H), which occupy unique positions and occur late in the
region encoding the rod domain. To account for this different placement of
introns, it was proposed that the first type IV gene arose by an mRNA-mediated
transposition event and that subsequent events led to the acquisition of the
few new introns (Lewis and Cowan,
1986
).
|
Using the presence of the two intron positions conserved in all mammalian
type IV genes, a total of nine type IV genes can be identified in F.
rubripes (Tables 1,
2). Several of these genes pose
problems in annotation compared with the mammalian genes and so some
assignments are tentative. The gene on scaffold 1912 predicts a protein with
84% sequence identity to goldfish gefiltin, the fish homolog for mammalian
-internexin (Markl and Schechter,
1998
). Indeed, the F. rubripes gefiltin-internexin
protein shares 60% identity with human internexin
(Fig. 2). The gene on scaffold
2208 predicts a gefiltin-like protein (gefiltin-like 1) that shares 74%
sequence identity with gefiltin but has a divergent tail domain (identity with
human internexin 50%). A further gefiltin like protein, gefiltin-like 2, is
coded by the type IV gene on scaffold 1885. Although gefiltin-like 2 shows
nearly the same similarity with gefiltin and vimentin over the rod domain, the
intron pattern identifies the gene as a type IV gene.
The two genes present on scaffolds 137 and 2296 predict proteins that are
related to gefiltin but have unique tail domains. The second halves of the
tail domains are highly acidic owing to the presence of many glutamic acid
residues, which often form polyglutamic acid strings. Because this is a
distinctive feature of mammalian (Lewis
and Cowan, 1986) and Xenopus
(Charnas et al., 1992
)
neurofilament triplet NF-L proteins, we tentatively name these F.
rubripes type IV genes NF-L1 and NF-L2, respectively
(Table 1). The F.
rubripes neurofilament triplet NF-M gene located on scaffold 6593 still
has two sequence gaps that obscure the intron pattern. Because of its
convincing relation to the corresponding goldfish gene
(Glasgow et al., 1994
), we used
this latter gene in the comparison below. The type IV gene located on scaffold
1245 is tentatively called NF-H because it has the additional intron position
of mammalian NF-H genes (Lees et al.,
1988
) and the predicted protein has a tail domain containing many
short degenerate repeats. Depending on the choice between two possible gene
structures, there are 19 or 30 degenerate repeats. Whereas the 21 degenerate
repeats of mammalian NF-H involve essentially the 14 residue motif
KSPEKAKSPVKEEA with two serine phosphorylation sites
(Lees et al., 1988
), the
F. rubripes repeat is based on the 10 residue motif ETKPAAKEEP with
one threonine phosphorylation site.
Gene Y on scaffold 2477 predicts the only F. rubripes type IV
protein with a low sequence similarity with gefiltin. The predicted protein
has a very small head and a very long tail domain. Although this is a
structural feature of mammalian nestin
(Lendahl et al., 1990) and
synemin (Titeux et al., 2001
),
no convincing homology was detected. Finally, gene X located on scaffold 120
predicts again a protein of 43% similarity with gefiltin but its astonishing
intron pattern (see below) makes an annotation very difficult.
Although the F. rubripes collection of type IV genes already
includes nine genes, it lacks obvious homologs encoding the large proteins
nestin (Lendahl et al., 1990)
and synemin (Titeux et al.,
2001
), the protein syncoilin, which is a constituent IF member of
the muscle dystrobrevin complex (Newey et
al., 2001
). Genes coding for non-keratin IF proteins are not
clustered in the human genome (Hesse et
al., 2001
). Similarly, in F. rubripes, there is no
scaffold that harbors more than one type III or one type IV gene.
Surprising intron additions and the problem of the origin of type IV
genes
Although, in general, fish and mammalian genes have the same intron pattern
(Aparicio et al., 2002), some
F. rubripes type IV genes do not
(Fig. 3). The genes for
gefiltin, gefiltin-like 2, NF-L1, NF-M and protein Y have only the conserved
two intron positions of mammalian type IV proteins. An additional intron is
found in the same position in mammalian and F. rubripes NF-H genes.
However, the genes encoding NF-L2 and gefiltin-like 1 have one or two
additional introns situated at novel positions. Even more complex is the
situation in gene X, which has eight introns: the two conserved intron
positions of type IV genes, three novel intron positions and a further three
positions that are characteristically found only in mammalian and F.
rubripes type I-III genes. These include the first intron position of
type II genes, the third intron position of type II genes (which is also
present in type III genes) and the intron position corresponding to the end of
the coil 1b domain, which is found in all type I-III genes. The documentation
of a fish IF gene that combines type I-III intron positions with type IV
intron positions (Fig. 3) is at
first difficult to accommodate in a model assuming that the first type IV gene
arose by translocation of an intronless mRNA into the genome
(Lewis and Cowan, 1986
). One
possibility for the origin of this gene X that stays within this model is the
speculation that it arose as a chimera of a keratin II and a type IV gene
(Fig. 3).
Genes NF-L2, gefiltin-like 1 and X together provide a total of six new intron positions of IF genes that have no counterpart in human IF genes. The number of novel fish IF intron positions is increased to ten by the novel intron positions in two type I keratin genes, one type II gene and the desmin 2 gene (see above). If we consider vimentin 2 as a special case, there are ten intron gains in the F. rubripes IF genes analysed, but no unusual intron loss (except for vimentin 2).
F. rubripes has two A lamins
Previous studies have shown that fish have four nuclear lamins. Whereas
lamins A, B1 and B2 are found in all classes of vertebrates, the additional
lamin LIII is only detected in amphibia and fish
(Döring and Stick, 1990;
Yamaguchi et al., 2001
;
Hofemeister et al., 2002
).
Table 3 shows that the genomic
F. rubripes sequences cover the complete genes for lamins A, B1 and
LIII (with its two alternative last exons, which produce the isoforms LIIIa
and LIIIb). The intron pattern of these three genes is perfectly conserved
between fish and human. The lamin B2 gene bridges scaffolds 6482 and 7682.
Exon 1 is located to scaffold 6482, where it is followed by a long intron
sequence that overlaps extensively with the end of scaffold 7678. This
scaffold carries also the middle part of the gene, but the 3' end is
probably obscured by the following large sequence gap. Interestingly, the
F. rubripes B2 lamin gene has an additional intron inserted in the
region encoding the coil 2a domain. Unexpectedly, a second lamin A is also
indicated (Table 3). Lamin A2
starts in scaffold 2719 (exon 1 plus intron) and continues with scaffold 6631
(exon 2 till end). Using the zebrafish lamin A as reference
(Hofemeister et al., 2002
), we
find sequence identity of 70% for both F. rubripes A lamins, which
share only 63% identity. This is the first report of the presence of two lamin
A genes in a vertebrate genome. It raises the question of whether one of them
belongs to those genes that contribute to the partial tetraploidy of F.
rubripes (Aparicio et al.,
2002
). The comparatively low degree of sequence similarity would
indicate an ancient duplication event.
|
![]() |
Acknowledgments |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() |
---|
Aparicio, S., Chapman, J., Stupka, E., Putnam, N., Chia, J. M.,
Dehal, P., Christoffels, A., Rash, S., Hoon, S., Smit, A. et al.
(2002). Whole-genome shotgun assembly and analysis of the genome
of Fugu rubripes. Science
297,1301
-1310.
Brenner, M., Johnson, A. B., Boespflug-Tanguy, O., Rodriguez, D., Goldman, J. E. and Messing, A. (2001). Mutations in GFAP, encoding glial fibrillary acidic protein, are associated with Alexander disease. Nat. Genet. 27,117 -120.[CrossRef][Medline]
Charnas, L. R., Szaro, B. G. and Gainer, H. (1992). Identification and developmental expression of a novel low molecular weight neuronal intermediate filament protein expressed in Xenopus laevis. J. Neurosci. 12,3010 -3024.[Abstract]
Coulombe, P. A., Ma, L. L., Yamada, S. and Wawersik, M. (2001). Intermediate filaments at a glance. J. Cell Sci. 114,4345 -4347.[Medline]
Döring, V. and Stick, R. (1990). Gene structure of nuclear lamin LIII of Xenopus laevis: a model for the evolution of IF proteins from a lamin-like ancestor. EMBO J. 9,4073 -4081.[Abstract]
Fuchs, E. and Weber, K. (1994). Intermediate filaments: structure, dynamics, function and disease. Annu. Rev. Biochem. 63,345 -382.[CrossRef][Medline]
Glasgow, E., Hall, C. M. and Schechter, N. (1994). Organization, sequence, and expression of a gene encoding goldfish neurofilament medium protein. J. Neurochem. 63, 52-61.[Medline]
Goldfarb, L. G., Park, K. Y., Cervenakova, L., Gorokhova, S., Lee, H. S., Vasconcelos, O., Nagle, J. W., Semino-Mora, C., Sivakumar, K. and Dalakas, M. C. (1998). Missense mutations in desmin associated with familial cardiac and skeletal myopathy. Nat. Genet. 19,402 -403.[CrossRef][Medline]
Herrmann, H. and Aebi, U. (2000). Intermediate filaments and their associates: multi-talented structural elements specifying cytoarchitecture and cytodynamics. Curr. Opin. Cell Biol. 12,79 -90.[CrossRef][Medline]
Herrmann, H., Fouquet, B. and Franke, W. W. (1989). Expression of intermediate filament proteins during development of Xenopus laevis. I. cDNA clones encoding different forms of vimentin. Development 105,279 -298.[Abstract]
Hess, J. F., Casselman, J. T. and FitzGerald, P. G.
(1996). Gene structure and cDNA sequence identify the beaded
filament protein CP49 as a highly divergent type I intermediate filament
protein. J. Biol. Chem.
271,6729
-6735.
Hesse, M., Magin, T. M. and Weber, K. (2001). Genes for intermediate filament proteins and the draft sequence of the human genome: novel keratin genes and a surprisingly high number of pseudogenes related to keratin genes 8 and 18. J. Cell Sci. 114,2569 -2575.[Medline]
Hofemeister, H., Kuhn, C., Franke, W. W., Weber, K. and Stick, R. (2002). Conservation of the gene structure and membrane targeting signals of germ cell specific lamin LIII in amphibians and fish. Eur. J. Cell Biol. 81,51 -60.[Medline]
International Human Genome Sequencing Consortium (2001). Initial sequencing and analysis of the human genome. Nature 409,860 -921.[CrossRef][Medline]
Irvine, A. D. and McLean, W. H. (1999). Human keratin diseases: the increasing spectrum of disease and subtlety of the phenotype-genotype correlation. Br. J. Dermatol. 140,815 -828.[CrossRef][Medline]
Karabinos, A., Schmidt, H., Harborth, J., Schnabel, R. and
Weber, K.(2001). Essential roles for four cytoplasmic
intermediate filament proteins in Caenorhabditis elegans development.
Proc. Natl. Acad. Sci. USA
98,7863
-7868.
Karabinos, A., Schunemann, J., Parry, D. A. and Weber, K. (2002). Tissue-specific co-expression and in vitro heteropolymer formation of the two small branchiostoma intermediate filament proteins A3 and B2. J. Mol. Biol. 316,127 -137.[CrossRef][Medline]
Lees, J. F., Shneidman, P. S., Skuntz, S. F., Carden, M. J. and Lazzarini, R. A. (1988). The structure and organization of the human heavy neurofilament subunit (NF-H) and the gene encoding it. EMBO J. 7,1947 -1955.[Abstract]
Lendahl, U., Zimmerman, L. B. and McKay, R. D. (1990). CNS stem cells express a new class of intermediate filament protein. Cell 60,585 -595.[Medline]
Lewis, S. A. and Cowan, N. J. (1986). Anomalous placement of introns in a member of the intermediate filament multigene family: an evolutionary conundrum. Mol. Cell. Biol. 6,1529 -1534.[Medline]
Li, R., Messing, A., Goldman, J. E. and Brenner, M. (2002). GFAP mutations in Alexander disease. Int. J. Dev. Neurosci. 20,259 -268.[CrossRef][Medline]
Markl, J. and Schechter, N. (1998). Fish intermediate filament proteins in structure, function and evolution. In Intermediate Filaments (ed. H. Hermann and J. R. Harris), pp. 1-33.New York: Plenum Press.
Masaki, S. and Quinlan, R. A. (1997). Gene structure and sequence comparisons of the eye lens specific protein, filensin, from rat and mouse: implications for protein classification and assembly. Gene 201,11 -20.[CrossRef][Medline]
Newey, S. E., Howman, E. V., Ponting, C. P., Benson, M. A.,
Nawrotzki, R., Loh, N. Y., Davies, K. E. and Blake, D. J.
(2001). Syncoilin, a novel member of the intermediate filament
superfamily that interacts with alpha-dystrobrevin in skeletal muscle.
J. Biol. Chem. 276,6645
-6655.
Sandilands, A., Masaki, S. and Quinlan, R. A. (1998). Lens intermediate filament proteins. In Intermediate Filaments (ed. H. Hermann and J. R. Harris), pp. 291-318.New York: Plenum Press.
Schaffeld, M., Höffling, S., Haberkamp, Conrad, M. and Markl, J. (2002a). Type I keratin cDNAs from the rainbow trout: independent radiation of keratins in fish. Differentiation 70,282 -291.[CrossRef][Medline]
Schaffeld, M., Haberkamp, M., Braziulis, E., Lieb, B. and Markl, J. (2002b). Type II keratin cDNAs from the rainbow trout: implications for keratin evolution. Differentiation 70,292 -299.[CrossRef][Medline]
Titeux, M., Brocheriou, V., Xue, Z., Gao, J., Pellissier, J. F.,
Guicheney, P., Paulin, D. and Li, Z. (2001). Human synemin
gene generates splice variants encoding two distinct intermediate filament
proteins. Eur. J. Biochem.
268,6435
-6449.
Tyner, A. L., Eichman, M. J. and Fuchs, E. (1985). The sequence of a type II keratin gene expressed in human skin: conservation of structure among all intermediate filament genes. Proc. Natl. Acad. Sci. USA 82,4683 -4687.[Abstract]
Wang, J., Karabinos, A., Zimek, A., Mayer, M., Riemer, D., Hudson, C., Lemaire, P. and Weber, K. (2002). Cytoplasmic intermediate filament protein expression in tunicate development; a specific marker for the test cells. Eur. J. Cell Biol. 81,302 -311.[Medline]
Waseem, A., Gough, A. C., Spurr, N. K. and Lane, E. B. (1990). Localization of the gene for human simple epithelial keratin 18 to chromosome 12 using polymerase chain reaction. Genomics 7,188 -194.[Medline]
Yamaguchi, A., Yamashita, M., Yoshikuni, M. and Hagahama, Y.
(2001). Identification and molecular cloning of germinal vesicle
lamin B3 in goldfish (Crassius auratus) oocytes. Eur. J.
Biochem. 268,932
-939.