Gamma-crystallin Genes Research Articles

Whole genome assembly of the armored loricariid catfish Ancistrus triradiatus highlights herbivory signatures

The catfish Ancistrus triradiatus belongs to the species-rich family Loricariidae. Loricariids display remarkable traits such as herbivory, a benthic lifestyle, the absence of scales but the presence of dermal bony plates. They are exported as ornamental fish worldwide, with escaped fishes becoming a threat locally. Although genetic and phylogenetic studies are continuously increasing and developmental genetic investigations are underway, no genome assembly has been formally proposed for Loricariidae yet. We report a high-quality genome assembly of Ancistrus triradiatus using long and short reads, and a newly assembled transcriptome. The genome assembly is composed of 9530 scaffolds, including 85.6% of ray-finned fish BUSCOs, and 26,885 predicted protein-coding genes. The genomic GC content is higher than in other catfishes, reflecting the higher metabolism associated with herbivory. The examination of the SCPP gene family indicates that the genes presumably triggering scale loss when absent, are present in the scaleless A. triradiatus, questioning their explanatory role. The analysis of the opsin gene repertoire revealed that gene losses associated to the nocturnal lifestyle of catfishes were not entirely found in A. triradiatus, as the UV-sensitive opsin 5 is present. Finally, most gene family expansions were related to immunity except the gamma crystallin gene family which controls pupil shape and sub-aquatic vision. Thus, the genome of A. triradiatus reveals that fish herbivory may be related to the photic zone habitat, conditions metabolism, photoreception and visual functions. This genome is the first for the catfish suborder Loricarioidei and will serve as backbone for future genetic, developmental and conservation studies.

Explosive Expansion of βγ-Crystallin Genes in the Ancestral Vertebrate

In jawed vertebrates, βγ-crystallins are restricted to the eye lens and thus excellent markers of lens evolution. These βγ-crystallins are four Greek key motifs/two domain proteins, whereas the urochordate βγ-crystallin has a single domain. To trace the origin of the vertebrate βγ-crystallin genes, we searched for homologues in the genomes of a jawless vertebrate (lamprey) and of a cephalochordate (lancelet). The lamprey genome contains orthologs of the gnathostome βB1-, βA2- and γN-crystallin genes and a single domain γN-crystallin-like gene. It contains at least two γ-crystallin genes, but lacks the gnathostome γS-crystallin gene. The genome also encodes a non-lenticular protein containing βγ-crystallin motifs, AIM1, also found in gnathostomes but not detectable in the uro- or cephalochordate genome. The four cephalochordate βγ-crystallin genes found encode two-domain proteins. Unlike the vertebrate βγ-crystallins but like the urochordate βγ-crystallin, three of the predicted proteins contain calcium-binding sites. In the cephalochordate βγ-crystallin genes, the introns are located within motif-encoding region, while in the urochordate and in the vertebrate βγ-crystallin genes the introns are between motif- and/or domain encoding regions. Coincident with the evolution of the vertebrate lens an ancestral urochordate type βγ-crystallin gene rapidly expanded and diverged in the ancestral vertebrate before the cyclostomes/gnathostomes split. The β- and γN-crystallin genes were maintained in subsequent evolution, and, given the selection pressure imposed by accurate vision, must be essential for lens function. The γ-crystallin genes show lineage specific expansion and contraction, presumably in adaptation to the demands on vision resulting from (changes in) lifestyle.Electronic supplementary materialThe online version of this article (doi:10.1007/s00239-010-9379-2) contains supplementary material, which is available to authorized users.

Formation of Amyloid Fibrils In Vitro from Partially Unfolded Intermediates of Human γC-Crystallin

Mature-onset cataract results from the formation of light-scattering aggregates of lens crystallins. Although oxidative or mutational damage may be a prerequisite, little is known of the initiation or nucleation of these aggregated states. In mice carrying mutations in gamma-crystallin genes, a truncated form of gamma-crystallin formed intranuclear filamentous inclusions within lens fiber cells. Previous studies have shown that bovine crystallins and human gammaD-crystallin form amyloid fibrils under denaturing conditions in vitro. The amyloid fibril formation of human gammaC-crystallin (HgammaC-Crys) induced by low pH, together with characterization of a partially unfolded intermediate in the process were investigated. HgammaC-Crys was expressed and purified from Escherichia coli. Partially unfolded intermediates were detected by tryptophan fluorescence spectroscopy and UV resonance Raman spectroscopy. The aggregation into amyloid fibrils was monitored by solution turbidity and fluorescence assay. The morphology of aggregates was characterized using transmission electron microscopy (TEM). Secondary structure of the peptides in their fibrillar state was characterized using Fourier transform infrared spectroscopy (FTIR). The structure of HgammaC-Crys was perturbed at low pH. Partially unfolded intermediates were detected when solution pH was lowered to pH 3. At pH 3, HgammaC-Crys aggregated into amyloid fibrils. The kinetics and extent of the reaction was dependent on protein concentration, pH, and temperature. TEM images of aggregates revealed aggregation stages from short to long fibrils and from long fibrils to light-scattering fibril networks. FTIR spectroscopy confirmed the cross-beta character of the secondary structure of these fibrils. HgammaC-Crys formed amyloid fibrils on incubation at low pH via a partially unfolded intermediate. This process could contribute to the early stages of the formation of light-scattering species in the eye lens.

Conversion and Compensatory Evolution of the γ-Crystallin Genes and Identification of a Cataractogenic Mutation That Reverses the Sequence of the Human CRYGD Gene to an Ancestral State

We identified a mutation in the CRYGD gene (P23S) of the gamma-crystallin gene cluster that is associated with a polymorphic congenital cataract that occurs with frequency of approximately 0.3% in a human population. To gain insight into the molecular mechanism of the pathogenesis of gamma-crystallin isoforms, we undertook an evolutionary analysis of the available mammalian and newly obtained primate sequences of the gamma-crystallin genes. The cataract-associated serine at site 23 corresponds to the ancestral state, since it was found in CRYGD of a lower primate and all the surveyed nonprimate mammals. Crystallin proteins include two structurally similar domains, and substitutions in mammalian CRYGD protein at site 23 of the first domain were always associated with substitutions in the structurally reciprocal sites 109 and 136 of the second domain. These data suggest that the cataractogenic effect of serine at site 23 in the N-terminal domain of CRYGD may be compensated indirectly by amino acid changes in a distal domain. We also found that gene conversion was a factor in the evolution of the gamma-crystallin gene cluster throughout different mammalian clades. The high rate of gene conversion observed between the functional CRYGD gene and two primate gamma-crystallin pseudogenes (CRYGEP1 and CRYGFP1) coupled with a surprising finding of apparent negative selection in primate pseudogenes suggest a deleterious impact of recently derived pseudogenes involved in gene conversion in the gamma-crystallin gene cluster.

Tissue-specific Regulation of the Mouse αA-crystallin Gene in Lens via Recruitment of Pax6 and c-Maf to its Promoter

Pax6 is a lineage-restricted DNA-binding transcription factor regulating the formation of mammalian organs including brain, eye and pancreas. Pax6 plays key roles during the initial formation of lens lineage, proliferation of lens progenitor and precursor cells and their terminal differentiation. In addition to Pax6, lens fiber cell differentiation is regulated by c-Maf, Prox1 and Sox1. Crystallins are essential lens structural proteins required for light refraction and transparency. Mouse alphaA-crystallin represents about 17% of all crystallins at the protein level and ranks as one of the most abundant tissue-specific proteins. Lens-specific expression of this gene is regulated at the level of transcription. A promoter fragment of -88 to +46 is capable of driving lens-specific expression in transgenic mouse. Here we provide data suggesting that this lens-specific promoter fragment is comprised of multiple Pax6 and Maf-binding sites. Site-directed mutagenesis of regions within these sites resulted in partially or completely reduced promoter activities in lens cells. Co-transfections using Pax6 and c-Maf alone revealed moderate and strong activations of this promoter, respectively. In contrast to synergistic activation of alphaB-crystallin by Pax6 and c-Maf, Pax6 has a neutral effect on c-Maf-mediated alphaA-crystallin promoter activation. Chromatin immunoprecipitations established in vivo interactions of Pax6 and c-Maf with the alphaA-crystallin promoter in lens cells. Collectively, the present data support a molecular model in which tissue-specific expression of alphaA-crystallin is regulated by recruitment of Pax6 and c-Maf, two proteins regulating multiple processes of lens differentiation, to its promoter. In addition, the data suggest a molecular model of temporal and spatial regulation of alphaB, alphaA and gamma-crystallin genes in mouse embryonic lens by using variants of the Pax6/Maf regulatory module.

HSF4 is required for normal cell growth and differentiation during mouse lens development.

The heat shock transcription factor (HSF) family consists of three members in mammals and regulates expression of heat shock genes via a heat shock element. HSF1 and HSF2 are required for some developmental processes, but it is unclear how they regulate these processes. To elucidate the mechanisms of developmental regulation by HSFs, we generated mice in which the HSF4 gene is mutated. HSF4-null mice had cataract with abnormal lens fiber cells containing inclusion-like structures, probably due to decreased expression of gamma-crystallin, which maintains protein stability. Furthermore, we found increased proliferation and premature differentiation of the mutant lens epithelial cells, which is associated with increased expression of growth factors, FGF-1, FGF-4, and FGF-7. Unexpectedly, HSF1 competed with HSF4 for the expression of FGFs not only in the lens but also in other tissues. These findings reveal the lens-specific role of HSF4, which activates gamma-crystallin genes, and also indicate that HSF1 and HSF4 are involved in regulating expression of growth factor genes, which are essential for cell growth and differentiation.

Study on ultrastructure changes and the genetic locus for a special phenotype cataract

To report the ultrastructure changes and map the genetic locus for a special phenotype autosomal dominant congenital cataract (ADCC) in a large Chinese family. A large four-generation Chinese family affected by this special phenotype ADCC was analyzed. Clinical examinations, light and transmission electron microscopy analysis of the removed lens tissue were performed. Blood samples were taken for DNA extraction and two-point linkage analyses between the polymorphisms of microsatellite markers near the gamma-crystallin gene (CRYG) and the disease-associated locus was also determined. The lens fiber cells displayed abnormal inter- and intracellular alterations, including irregular refractivity, focal degeneration and irregular and enlarged intracellular spaces. Linkage analysis showed that there was linkage between the ADCC disease-associated locus and D2S2208, D2S2382 and D2S164. There were characteristic alterations to the lens fiber cells of this special phenotype ADCC and the CRYGD gene might be the disease-associated.

GammaE-crystallin recruitment to the plasma membrane by specific interaction between lens MIP/aquaporin-0 and gammaE-crystallin.

Major intrinsic protein (MIP), also called aquaporin-0, is essential for lens transparency and is specifically expressed in the lens fiber cell membranes. The goal of the current study was to identify and characterize proteins that interact with MIP and to elucidate the role of these interactions in MIP functions. The C-terminal 74-amino-acid fragment of MIP was used as bait to screen a rat lens cDNA yeast two-hybrid library. The full-length MIP was expressed as enhanced green fluorescent protein (EGFP)-tagged or myc-tagged proteins, and gammaE-crystallin was expressed as FLAG-tagged or red fluorescent protein (HcRed)-tagged proteins, respectively, in the RK13 rabbit kidney epithelial cell line. Protein-protein interactions were analyzed by coimmunoprecipitation assays and visualized by confocal fluorescence microscopy. gammaE-Crystallin, a water-soluble protein that is specifically expressed in lens fibers, was identified as a binding protein to the MIP C-terminal peptide. Coimmunoprecipitation assays demonstrated that gammaE-crystallin interacts specifically with full-length MIP in mammalian cells. MIP did not interact with gammaD-crystallin, another member of the highly conserved gamma-crystallin gene family. Confocal fluorescence microscopy demonstrated that MIP interacted with gammaE-crystallin in individual mammalian cells and that this interaction resulted in the recruitment of gammaE-crystallin from the cytoplasm to the plasma membrane. These experiments provide the first demonstration of MIP interaction with other lens proteins at the molecular level and raise the possibility of a structural role of MIP in the organization of gamma-crystallins in lens fibers.

Cryopreservation of Xenopus transgenic lines.

Xenopus laevis has been widely used for molecular, cellular, and developmental studies. With the development of the sperm-mediated transgenic method, it is now possible to study gene function during vertebrate development by using this popular model. On the other hand, like other animal species, it is labor intensive, and the maintenance of transgenic lines is expensive. In this article, we investigated the possibility of using sperm-cryopreservation as a means to preserve transgenic frog lines. We demonstrated that cryopreserved sperms are viable but not fertile under our in vitro fertilization (IVF) conditions. However, by microinjecting cryopreserved sperm nuclei, we successfully regenerated a transgenic line carrying a double promoter transgene construct, where the marker gene encoding the green fluorescent protein (GFP) is driven by the gamma-crystallin gene promoter and a gene of interest, encoding a fusion protein of GFP with the matrix metalloproteinase stromelysin-3 (ST3-GFP), is driven by a heat shock-inducible promoter. We demonstrated the functional transmission of the ST3-GFP transgene by analyzing the phenotype of the F1 animals after heat-shock to induce its expression. Our method thus provides an inexpensive means to preserve transgenic frog lines and a convenient way for distribution of transgenic lines. Furthermore, the ease with which to microinject nuclei compared to the technically demanding transgenesis procedure with variable outcome should facilitate more laboratories to use transgenic Xenopus laevis for functional studies in vivo. Mol. Reprod. Dev. 67: 65-69, 2004.

Antagonistic action of Six3 and Prox1 at the gamma-crystallin promoter

Gamma-crystallin genes are specifically expressed in the eye lens. Their promoters constitute excellent models to analyse tissue-specific gene expression. We investigated murine CRYGE/f promoters of different length in lens epithelial cell lines. The most active fragment extends from position -219 to +37. Computer analysis predicts homeodomain and paired-domain binding sites for all rodent CRYGD/e/f core promoters. As examples, we analysed the effects of Prox1 and Six3, which are considered important transcription factors involved in lens development. Because of endogenous Prox1 expression in N/N1003A cells, a weak stimulation of CRYGE/f promoter activity was found for PROX1. In contrast, PROX1 stimulated the CRYGF promoter 10-fold in CD5A cells without endogenous PROX1. In both cell lines Six3 repressed the CRYGF promoter to 10% of its basal activity. Our cell transfection experiments indicated that CRYG expression increases as Six3 expression decreases. Prox1 and Six3 act antagonistically on regulation of the CRYGD/e/f promoters. Functional assays using randomly mutated gammaF-crystallin promoter fragments define a Six3-responsive element between -101 and -123 and a Prox1-responsive element between -151 and -174. Since Prox1 and Six3 are present at the beginning of lens development, expression of CRYGD/e/f is predicted to remain low at this time. It increases as Six3 expression decreases during ongoing lens development.

Prox 1 in eye degeneration and sensory organ compensation during development and evolution of the cavefish Astyanax

We have investigated expression of the homeobox gene Prox 1 during eye degeneration and sensory organ compensation in cavefish embryos. The teleost Astyanax mexicanus consists of sighted surface-dwelling forms (surface fish) and several populations of blind cave-dwelling forms (cavefish), which have evolved independently. Eye formation is initiated during cavefish development, but the lens vesicle undergoes apoptosis, and the eye subsequently arrests and degenerates. The requirement of Prox 1 for lens fiber differentiation and gamma-crystallin expression in the mouse suggests that changes in the expression of this gene could be involved in cavefish eye degeneration. Surface fish and cavefish embryos stained with a Prox 1 antibody showed Prox 1 expression in the lens, neuroretina, myotomes, heart, hindbrain, and gut, as reported in other vertebrates. We found that Prox 1 expression is not altered during cavefish lens development. Prox 1 protein was detected in the lens vesicle as soon as it formed and persisted until the time of lens degeneration in each cavefish population. The cavefish lens vesicle was also shown to express a gamma-crystallin gene, suggesting that Prox 1 is functional in cavefish lens development. In addition to the tissues described above, Prox 1 is expressed in developing taste buds and neuromasts in cavefish, which are enhanced to compensate for blindness. It is concluded that the Prox 1 gene is not involved in lens degeneration, but that expansion of the Prox 1 expression domain occurs during taste bud and neuromast development in cavefish.

A 5-base insertion in the gammaC-crystallin gene is associated with autosomal dominant variable zonular pulverulent cataract.

A seven-generation family with 30 members affected by highly variable autosomal dominant zonular pulverulent cataracts has been previously described. We have localized the cataracts to a 19-cM interval on chromosome 2q33-q35 including the gamma-crystallin gene cluster. Maximum lod scores are 4.56 (theta=0.02) with D2S157, 3.66 (theta=0.12) with D2S72, and 3.57 (theta=0.052) with CRYG. Sequencing and allele-specific oligonucleotide analysis of the pseudo gammaE-crystallin promoter region from individuals in the pedigree suggest that activation of the gammaE-crystallin pseudo gene is unlikely to cause the cataracts in the family. In addition, base changes in the TATA box but not the Sp1-binding site have been found in unaffected controls and can be excluded as a sole cause of cataracts. In order to investigate the underlying genetic mechanism of cataracts in this family further, exons of the highly expressed gammaC- and gammaD-crystallin genes have been sequenced. The gammaD-crystallin gene shows no abnormalities, but a 5-bp duplication within exon 2 of the gammaC-crystallin gene has been found in one allele of each affected family member and is absent from both unaffected family members and unaffected controls. This mutation disrupts the reading frame of the gammaC-crystallin coding sequence and is predicted to result in the synthesis of an unstable gammaC-crystallin with 38 amino acids of the first "Greek key" motif followed by 52 random amino acids. This finding suggests that the appropriate association of mutant betagamma-crystallins into oligomers is not necessary to cause cataracts and may give us new insights into the genetic mechanism of cataract formation.

Structure of the crystallins.

The lens is formed from two protein superfamilies, the alpha- and beta gamma-crystallins. Representative three-dimensional structures show they both have a basic 2-beta-sheet domain fold, with the beta gamma-domain being made from two intercalating Greek keys. X-ray structures of monomeric gamma-crystallins and simple oligomeric beta-crystallins show how multiple gene duplications can give rise to highly symmetrical assemblies based on paired domains. These protein folds have been engineered by directed mutagenesis to investigate the roles of the critical region in domain pairing and assembly. Inherited human cataracts have been described that are associated with representatives of each of the crystallin protein families. Mutations to certain beta- and gamma-crystallin genes cause expression of truncated polypeptides that would not be expected to fold properly; instead they would randomly aggregate causing light scattering. As crystallin proteins are not renewed, age-related cataract is a gradual accumulation of small changes to pre-existing normal proteins. The precise sites of post-translational modifications are now being mapped to the various crystallins.

Mouse models of congenital cataract.

Mouse mutants affecting lens development are excellent models for corresponding human disorders. The mutant aphakia has been characterised by bilaterally aphakic eyes (Varnum and Stevens, J Hered 1968;59:147-50); the corresponding gene was mapped to chromosome 19 (Varnum and Stevens, Mouse News Lett 1975;53:35). Recent investigations in our laboratory refined the linkage of 0.6 cM proximal to the marker D19Mit10. Several candidate genes have been excluded (Chuk1, Fgf8, Lbp1, Npm3, Pax2, Pitx3). The Cat3 mutations are characterised by vacuolated lenses caused by alterations in the initial secondary lens fibre cell differentiation. Secondary malformations develop at the cornea and iris, but the retina remains unaffected. The mutation has been mapped to chromosome 10 close to the markers D10Mit41 and D10Mit95. Several candidate genes have been excluded (Dcn, Elk3, Ldc, Mell8, Tr2-11). The series of Cat2 mutations have been mapped close to the gamma-crystallin genes (Cryg; Löster et al., Genomics 1994;23:240-2). The Cat2nop mutation is characterised by a mutation in the third exon of Crygb leading to a truncated gamma B-crystallin and the termination of lens fibre cell differentiation. The Cat2 mutants are interesting models for human cataracts caused by mutations in the human CRYG genes at chromosome 2q32-35.

Cataract mutations and lens development.

The lens plays an essential role for proper eye development. Mouse mutants affecting lens development are excellent models for corresponding human disorders. Moreover, using mutations in particular genes the process of eye and lens development can be dissected into distinct steps. Therefore, three mouse mutants will be described in detail and discussed affecting three essential stages: formation of the lens vesicle, initiation of secondary lens fiber cell formation, and terminal differentiation of the secondary fiber cells. The mutant aphakia (ak) has been characterized by bilaterally apakic eyes [Varnum and Stevens (1968) J. Hered. 59, 147-150], and the corresponding gene was mapped to chromosome 19 [Varnum and Stevens (1975) Mouse News Letters 53, 35]. Recent investigations in our laboratory refined the linkage 0.6 +/- 0.3 N cm proximal to the microsatellite marker D19Mit10. The linked gene Pax2, responsible for proper development of the posterior part of the eye and the optic nerve, was excluded as candidate gene by sequence analysis. Histological analysis of the homozygous ak mutants revealed a persisting lens stalk and subsequently the formation of lens rudiments. The lens defects led to irregular iris development and retinal folding. Congenital aphakia is known as a rare human anomaly. Besides a corneal dystrophy (CDTB), no corresponding disease is localized at the homologous region of human chromosome 10q23. The Cat3 mutations are characterized by vacuolated lenses caused by alterations in the beginning of secondary lens fiber cell differentiation at embryonic day 12.5. Secondary malformations develop at the cornea and the iris, but the retina remains unaffected. Two mutant alleles of the Cat3 locus have been mapped to mouse chromosome 10 very close to the microsatellite markers D10Mit41 and D10Mit95 (less than 0.3 cM). Since Cat3 is mapped to a position, which is homologous to human chromosome 12q21-24, the disorder cornea plana congenita can be considered as a candidate disease. The series of Cat2 mutations have been mapped close to the locus encoding the gamma-crystallin gene cluster Cryg [Löster et al. (1994) Genomics 23, 240-242]. The Cat2nop mutation is characterized by a deletion of 11 bp and an insertion of 4 bp in the 3rd exon of Crygh leading to a truncated gamma B-crystallin. The defect in the Crygh gene is causative for the stop of lens fiber cell differentiation from embryonic day 15.5 onward. Besides the lens, no further ocular tissue is affected. The Cat2 mouse mutants are interesting models for human cataracts caused by mutations in the gamma-crystallin genes at human chromosome 2q32-35. The ak, Cat3 and Cat2 mutants are discussed in the context of other mutants affecting early eye and lens development. Additionally, human congenital cataracts are discussed, which have been characterized similar to the mouse models. The overview of the three types of mutants demonstrates that genes, which affect the early eye development, e.g. at the lens vesicle stage, have consequences for the development of the whole eye. In contrast, if the mutation influences later steps of lens differentiation, the consequences are restricted to the lens only. These data indicate a decreasing effect of the lens for the regulation of eye development during embryogenesis.

Regulation of Expression within a Gene Family: THE CASE OF THE RAT γB- AND γD-CRYSTALLIN PROMOTERS

The six closely related and clustered rat gamma-crystallin genes, the gammaA- to gammaF-crystallin genes, are simultaneously activated in the embryonic lens but differentially shut down during postnatal development with the gammaB-crystallin gene, the last one to be active. We show here that developmental silencing of the gammaD-crystallin promoter correlates with delayed demethylation during lens fiber cell differentiation. Methylation silencing of the gammaD-crystallin promoter is a general effect and does not require the methylation of a specific CpG, nor does methylation interfere with factor binding to the proximal activator. In later development, the gammaD-crystallin promoter is also shut down earlier by a repressor that footprints to the -91/-78 region. A factor with identical properties is present in brain. Hence, a ubiquitous factor has been recruited as a developmental regulator by the lens. All gamma-crystallin promoters tested contain upstream silencers, but at least the gammaB-crystallin silencer is distinct from the gammaD-crystallin silencer. The gamma-crystallin promoters were found to share a proximal activator (the gamma-box; around -50), which behaves as a MARE. The gammaB-box is recognized with much lower avidity than the gammaD-box. By swapping elements between the gammaB- and the gammaD-crystallin promoter, we show that activation by the gammaB-box requires a directly adjacent -46/-38 AP-1 consensus site. These experiments also uncovered another positive element in the gammaD-crystallin promoter, around -10. In the context of the gammaD-crystallin promoter, this element is redundant; in the context of the gammaB-crystallin promoter, it can replace the -46/-38 element.

Sox1 directly regulates the gamma-crystallin genes and is essential for lens development in mice.

gamma-Crystallins are major structural components of the lens fiber cells in amphibians and mammals. Many dominant inherited cataracts in humans and mice have been shown to map within the gamma-crystallin gene cluster. Several transcription factors, including PAX6 and SOX proteins, have been suggested as candidates for crystallin gene regulation. Here we show that the targeted deletion of Sox1 in mice causes microphthalmia and cataract. Mutant lens fiber cells fail to elongate, probably as a result of an almost complete absence of gamma-crystallins. It appears that the direct interaction of the SOX1 protein with a promoter element conserved in all gamma-crystallin genes is responsible for their expression.

INVITED EDITORIAL The Genetics of Cataract: Our Vision Becomes Clearer

I would like to thank Drs. Joe Horwitz, Muriel Kaiser-Kupfer, Mark Kantorow, Eric Wawrousek, and Sam Zigler for a close reading of the manuscript.

Linkage of polymorphic congenital cataract to the gamma-crystallin gene locus on human chromosome 2q33-35

Cataract is one of the major causes of blindness in humans. We describe here an autosomal dominant polymorphic congenital cataract (PCC) which is characterised by wide variations in phenotype of non-nuclear lens opacities, even among affected members of the same family. PCC families included a large, unique pedigree (254 members, 103 affected individuals), and genetic linkage was conducted using a variety of polymorphic markers. Evidence for linkage was found for chromosome 2q33-35 with PCC mapping near D2S72 and TNP1. A tri-nucleotide microsatellite marker for gamma-crystallin B gene (CRYG1) was found to co-segregate with PCC and yielded a maximum lod score of 10.62 at (theta = 0). A multipoint analysis demonstrated that the most probable location of the PCC gene was within an 8 cM genetic interval containing the gamma-crystallin gene cluster. These data provide strong evidence of the existence of an autosomal dominant mutation for PCC in or near the gamma-crystallin gene cluster. This defect is characterised by complete penetrance but variable expression of the cataract phenotype. Our study also suggests that non-nuclear human cataracts might be caused by some abnormality in gamma-crystallin genes.

Cataract mutations as a tool for developmental geneticists.

Lens development as a multistep process can be analyzed by the investigation of distinct cataract mutants. Since the mutant genes are molecularly characterized, the function of the wild-type allele can be deduced. Besides some mutations affecting the lens induction, which are not yet characterized at the molecular level, mainly mutations affecting the crystallin genes are discussed. In particular, for the murine gamma-crystallin genes 8 different mutations are described in the mouse, which lead to different, distinguishable phenotypes. The distinct and complex phenotypes cannot be explained solely by the changed physiochemical properties of the altered crystallin packaging, but point to a regulatory function of the crystallins during lenticular development and differentiation.