A new dominant cataract mutation was observed among the
F
1 offspring of ENU-treated male mice. This
opacity, preliminarily referred to as
Aey7, was demonstrated
to be caused by a T→A exchange within the third exon of the
Cryaa gene, and the mutant allele was therefore designated
Cryaa Aey7 . The predicted amino acid in the
mutants is Glu instead of Val at position 124, close to the C terminus.
The lens opacity is already visible at eye opening. The cataractous
changes coincide with the expression of the
Cryaa gene,
which is observed first in the lens cup. Later on, αA-crystallin
becomes very abundant in lens fiber cells
8 9 10 11 and is
considered as a major structural protein of the ocular
lens.
2
α-Crystallin participates in the intracellular architecture through
cellular filaments. Together with CP49, a lens-specific cytoskeletal
protein, α-crystallins form the beaded filaments.
26 α
A-Crystallin also interacts with tubulin
27 and
actin.
28 Moreover, there are several lines of biochemical
evidence that α-crystallin may become associated with the plasma
membrane. Because the molecular sites of these interactions are not yet
known in detail,
29 whether the V124E mutation described
herein is involved in these interactions is open to speculation.
Besides its structural properties, αA-crystallin is the target of
posttranslational alterations, and one of its most important
modifications is phosphorylation. Past studies have demonstrated that
the major site of αA-crystallin in vivo phosphorylation is Ser122,
which is phosphorylated in a cAMP-dependent manner.
30 This
phosphorylated form of αA-crystallin could be detected in human
lenses only from adolescent, adult, and senile donors, but not in
infants, suggesting a developmental regulation of this particular kind
of modification.
31 Computer-assisted prediction programs,
such as ProSite (ExPASy), did not support the hypothesis that the V124E
mutation influences the use of the very close phosphorylation site at
Ser122.
However, the most exciting finding concerns the function ofα
A-crystallin as a molecular chaperone.
32 α
A-Crystallin prevents thermal aggregation of several enzymes and
even of β- and γ-crystallins. Chaperone activity is essential for
the lens, because degradation and extrusion of defective proteins is
not possible as it is in other tissues. Moreover, the lens is exposed
to a variety of damaging agents, particularly in light of various
wavelengths leading to oxidative effects on quite a number of lens
proteins. α-Crystallin has a substrate specificity different from
other chaperones and recognizes specific nonnative intermediates formed
during denaturation only.
33
Based on the gene structure, Wistow
34 thought the overall
structure of α-crystallins to consist of a globular N-terminal domain
of two symmetry-related motifs and a somewhat longer C-terminal domain
also consisting of two motifs. The two globular domains, which are
built up by two exons, are fused by a short connecting peptide, which
is extended in the rodent αA
ins-crystallin by
23 amino acids
5 and has a significant reduced chaperone
activity compared with the normal αA-crystallin.
35 36 The C terminus of the αA-crystallin consists of two rather
hydrophilic domains
2 that are exposed to the surface and
tend to form tetrameric assemblies.
37 Moreover, there is
considerable evidence for the involvement of the flexible C-terminal
extension in chaperone activity. Its truncation or immobilization
greatly reduces this capacity, suggesting that the hydrophilic tail is
likely to be important in keeping complexes of chaperones and bound
proteins in solution.
4 The chaperone activity ofα
-crystallin was first localized to amino acid residues 158-173 of
the C-terminal region of αA-crystallin,
38 but recently,
reduced chaperone-like activity was demonstrated also in anα
A-crystallin mutation (Asp69Ser)
39 and by the
introduction of a hydrophobic tryptophan at position
172.
40
The mutation described in this article replaces a neutral, hydrophobic
amino acid (Val) with an acidic amino acid (Glu) in the C-terminal part
(position 124) of the αA-crystallin. The same region of theα
A-crystallin is affected by a mutation in human
CRYAA causing a dominant zonular central cataract phenotypically very similar
to
Aey7. It was characterized by a missense mutation gene
leading to replacement of Arg with Cys at position 116
(R116C).
15 Detailed biochemical analysis of this mutant
protein demonstrated a fourfold reduction in chaperone-like activity,
but it tends to bind to membranes 10 times more than the wild-type
form.
41 Similar explanations may be suggested for
cataractogenesis in the
Cryaa Aey7 mutants.
In mouse and human, a few other mutations in the
Cryaa/
CRYAA genes have already been described.
The loss of the entire
Cryaa gene in mouse knockout mutants
leads to an opacification of the lens only in homozygous
conditions.
13 In the recessive
lop18 mouse
mutants, a missense mutation in the first exon of the
Cryaa gene converting codon 54 Arg to His was demonstrated recently as cause
for the phenotype.
14 A recessive cataract has been also
reported in human, which is caused by the nonsense mutation W9X,
resulting in a premature stop codon of
CRYAA.
16
Of interest and in contrast to the other abundant lens proteins, theβ
- and γ-crystallins, mutations in the Cryaa and CRYAA genes lead to both recessive and dominant phenotypes.
Mutations affecting the N-terminal part or even the loss of the entire
protein result in recessive phenotypes, whereas mutations in the
C-terminal part leads to dominant phenotypes. Most likely, the dominant
mutation inhibits interactions with other proteins affecting the
chaperone activity, affinity with the membranes, or interactions with
the lens intermediate filaments.
The authors thank Ingenium Pharmaceutical, AG, Martinsried,
Germany, for kindly providing the Aey7 mutant; Francoise
André, Gerlinde Bergter, Erika Bürkle, Nicole Hirsch,
Sabine Manz, Andreas Mayer, Dagmar Reinl, and Monika Stadler for expert
technical assistance; and Utz Linzner, GSF-Institute of Experimental
Genetics, Neuherberg, Germany, for providing the oligonucleotides.