Ionizing radiation and genetic risks. II. Nature of radiation-induced mutations in experimental mammalian in vivo systems

This paper reviews data on the nature of spontaneous and radiation-induced mutations in the mouse. The data are from studies using a variety of endpoints scorable at the morphological or the biochemical level and include pre-selected as well as unselected loci at which mutations can lead to recessive or dominant phenotypes. The loci used in the morphological recessive specific-locus tests permit the recovery of a wide spectrum of induced changes. Important variables that affect the nature of radiation-induced mutations (assessed primarily using tests for viability of homozygotes) include: germ cell stage, type of irradiation and the locus. Most of the results pertain to irradiated stem cell spermatogonia. The data on morphological specific-locus mutations show that overall, more than two-thirds of the X- or gamma-ray-induced mutations are lethal when homozygous. This proportion may be lower for those that occur spontaneously, but the numbers of tested mutants are small. For spontaneous mutations, there is evidence for the occurrence of mosaics and for proviral insertions. Most or all tested induced enzyme activity variants, dominant visibles (recovered in specific-locus experiments) and dominant skeletal mutations are lethal when homozygous and this is true of 50% of dominant cataract mutations, but again, the numbers of tested mutants are small. Electrophoretic mobility variants, which are known to be due to base-pair changes, are seldom induced by irradiation. At the histocompatibility loci, no radiation-induced mutations have been recovered, presumably because deletions are incompatible with survival even in heterozygotes. All these findings are consistent with the view that in mouse germ cells, most radiation-induced mutations are DNA deletions. Some mutations (in the morphological specific-locus tests) which had previously been inferred to be deletions on the basis of genetic analyses have now been shown to be DNA deletions by molecular methods. However, the possibility cannot be excluded that at least a small proportion of induced mutations may be intragenic changes. The data on the rates of induction of recessive lethals and of dominant skeletal and dominant cataract mutations (and proportions of the latter two which are homozygous lethal) can be used to estimate the proportions of recessive lethals which are expressed as skeletal abnormalities or cataracts. These calculations show that about 10% of recessive lethals manifest themselves as skeletal and less than 0.2% as cataract mutations.(ABSTRACT TRUNCATED AT 400 WORDS)     

The Let-60 Locus Controls the Switch between Vulval and Nonvulval Cell Fates in Caenorhabditis Elegans

During induction of the Caenorhabditis elegans hermaphrodite vulva by the anchor cell of the gonad, six multipotent vulval precursor cells (VPCs) have two distinct fates: three VPCs generate the vulva and the other three VPCs generate nonspecialized hypodermis. Genes that control the fates of the VPCs in response to the anchor cell signal are defined by mutations that cause all six VPCs to generate vulval tissue (Multivulva or Muv) or that cause all six VPCs to generate hypodermis (Vulvaless or Vul). Seven dominant Vul mutations were isolated as dominant suppressors of a lin-15 Muv mutation. These mutations are dominant alleles of the gene let-60, previously identified only by recessive lethal mutations. Our genetic studies of these dominant Vul recessive lethal mutations, recessive lethal mutations, intragenic revertants of the dominant Vul mutations, and the closely mapping semi-dominant multivulva lin-34 mutations suggest that: (1) loss-of-function mutations of let-60 are recessive lethal at a larval stage, but they also cause a Vul phenotype if the lethality is rescued maternally by a lin-34 gain-of-function mutation. (2) The dominant Vul alleles of let-60 are dominant negative mutations whose gene products compete with wild-type activity. (3) lin-34 semidominant Muv alleles are either gain-of-function mutations of let-60 or gain-of-function mutations of an intimately related gene that elevates let-60 activity. We propose that let-60 activity controls VPC fates. In a wild-type animal, reception by a VPC of inductive signal activates let-60, and it generates into a vulval cell type; in absence of inductive signal, let-60 activity is low and the VPC generates hypodermal cells. Our genetic interaction studies suggest that let-60 acts downstream of let-23 and lin-15 and upstream of lin-1 and lin-12 in the genetic pathway specifying the switch between vulval and nonvulval cell types.     

Spectrum of mutations in the major human skeletal muscle chloride channel gene (CLCN1) leading to myotonia.

Autosomal dominant myotonia congenita and autosomal recessive generalized myotonia (GM) are genetic disorders characterized by the symptom of myotonia, which is based on an electrical instability of the muscle fiber membrane. Recently, these two phenotypes have been associated with mutations in the major muscle chloride channel gene CLCN1 on human chromosome 7q35. We have systematically screened the open reading frame of the CLCN1 gene for mutations by SSC analysis (SSCA) in a panel of 24 families and 17 single unrelated patients with human myotonia. By direct sequencing of aberrant SSCA conformers were revealed 15 different mutations in a total of 18 unrelated families and 13 single patients. Of these, 10 were novel (7 missense mutations, 2 mutations leading to frameshift, and 1 mutation predicted to affect normal splicing). In our overall sample of 94 GM chromosomes we were able to detect 48 (51%) mutant GM alleles. Three mutations (F413C), R894X, and a 14-bp deletion in exon 13) account for 32% of the GM chromosomes in the German population. Our finding that A437T is probably a polymorphism is in contrast to a recent report that the recessive phenotype GM is associated with this amino acid change. We also demonstrate that the R894X mutation may act as a recessive or a dominant mutation in the CLCN1 gene, probably depending on the genetic background. Functional expression of the R894X mutant in Xenopus oocytes revealed a large reduction, but not complete abolition, of chloride currents. Further, it had a weak dominant negative effect on wild-type currents in coexpression studies. Reduction of currents predicted for heterozygous carriers are close to the borderline value, which is sufficient to elicit myotonia.     

Mutations Affecting Functions of the Drosophila Gene Glued

Glued mutations in Drosophila comprise an essential complementation group with complex developmental effects. The original Glued mutation (Gl) has dominant nonlethal effects in heterozygous flies, principally on the morphogenesis of the visual system. Gl also has a recessive lethal effect early in development. Mutations that reverse the dominant visual effects of Gl (GlR mutations) were induced by gamma-radiation or by insertions of the transposable P element. The GlR(G) mutations induced by gamma-radiation do not reverse the lethal effect of Gl; these appear to be null mutations, some of which (and possibly all) delete segments of the Glued region. The GlR(P) mutations induced by insertion of the P element also reverse concomitantly a recessive lethal effect of Gl, suggesting that both the recessive and dominant effects are controlled by the same gene. The reversal of a lethal effect of Gl by the P element is remarkable, since it indicates that an essential gene function can be restored by insertion of unrelated DNA. Another class of lethal Glued mutations was induced in the normal Gl+ strain by ethyl methanesulfonate (EMS). The EMS mutations belong to the same essential complementation group as Gl, but do not have the strong dominant effects of Gl on the visual system. The GlR(P) mutations provide a molecular marker for the Glued gene, which was used to map the gene to the 70C2 band of chromosome 3L by in situ hybridization of a P element probe to polytene chromosomes from the GlR(P) strains and also to isolate clones of Glued genomic DNA for molecular studies of the normal gene and the various Glued mutations.     

A Novel Dominant Transformer Allele of the Sex-Determining Gene Her-1 of Caenorhabditis Elegans

We have characterized a novel dominant allele of the sex-determining gene her-1 of Caenorhabditis elegans. This allele, called n695, results in the incomplete transformation of XX animals into phenotypic males. Previously characterized recessive her-1 alleles transform XO animals into phenotypic hermaphrodites. We have identified five new recessive her-1 mutations as intragenic suppressors of n695. Three of these suppressors are weak, temperature-sensitive alleles. We show that the recessive her-1 mutations are loss-of-function alleles, and that the her-1(n695) mutation results in a gain-of-function at the her-1 locus. The existence of dominant and recessive alleles that cause opposite phenotypic transformations demonstrates that the her-1 gene acts to control sexual identity in C. elegans.     

Novel Mutation to Dominant Fucose Resistance in the l-Arabinose Operon of Escherichia coli

We isolated an unusual mutant in which the arabinose operon carried an araC lesion that led to a dominant fucose resistance while retaining recessive constitutivity. All previously characterized C(c) mutations have been recessive for both fucose resistance and constitutivity.     

Closing the gap on autosomal dominant connexin-26 and connexin-43 mutants linked to human disease

Cells within the vast majority of human tissues communicate directly through clustered arrays of intercellular channels called gap junctions. Gene ablation studies in mouse models have revealed that these intercellular channels are necessary for a variety of organ functions and that some of these genes are essential for survival. Molecular genetics has uncovered that germ line mutations in nearly half of the genes that encode the 21-member connexin family of gap junction proteins are linked to one or more human diseases. Frequently, these mutations are autosomal recessive, whereas in other cases, autosomal dominant mutations manifest as disease. Given the broad and overlapping distribution of connexins in a wide arrangement of tissues, it is hard to predict where connexin-linked diseases will clinically manifest. For instance, the most prevalent connexin in the human body is connexin-43 (Cx43), yet autosomal dominant mutations in the GJA1 gene, which encodes Cx43, exhibit modest developmental disorders resulting in a disease termed oculodentodigital dysplasia. Autosomal recessive mutations in the gene encoding Cx26 result in moderate to severe sensorineural hearing loss, whereas autosomal dominant mutations produce hearing loss and a wide range of skin diseases, including palmoplantar keratoderma. Here, we will focus on autosomal dominant mutations of the genes encoding Cx26 and Cx43 in relation to models that link genotypes to phenotypic outcomes with particular reference to how these approaches provide insight into human disease.     

Cellular pathophysiology of cystic kidney disease: insight into future therapies.

Polycystic kidney disease (PKD) is a developmental kidney disorder which can be inherited as either an autosomal dominant trait, with an incidence of 1:50 to 1:1000, or as an autosomal recessive trait with an incidence of 1:6,000 to 1:40,000. Three different genes have now been cloned that are associated with mutations that cause PKD. Two of these are linked to the most common forms of the dominant disease while the third is associated with the orpk mouse model of recessive polycystic kidney disease. Advances in understanding the molecular genetics of PKD have been paralleled by new insights into the cellular pathophysiology of cyst formation and progressive enlargement. Current data suggest that a number of PKD proteins may interact in a complex, which when disrupted by mutations in PKD genes may lead to altered epithelial proliferative activity, secretion, and cell matrix biology. The identification of a unique cystic epithelial phenotype presents new opportunities for targeted therapies. These include targeted gene therapy, gene complementation, and specific immunological or pharmacological interruption of growth factor pathways.     

The frequency of dominant cataract and recessive specific-locus mutations in mice derived from 80 or 160 mg ethylnitrosourea per kg body weight treated spermatogonia

A systematic comparison of the frequency of dominant cataract and recessive specific-locus mutations in mice has been extended to include results for 80 and 160 mg ethylnitrosourea per kg body weight spermatogonial treatment. The frequency of confirmed dominant cataract mutations in the historical control, 80 and 160 mg/kg ethylnitrosourea treatment groups was 1/22594, 8/5090 and 14/6435, respectively. The frequency of recessive specific-locus mutations in the same dose groups was, respectively, 19/227805, 20/13274 and 35/8658. These present results confirm previous results, which indicate that ethylnitrosourea is effective in inducing both recessive specific-locus and dominant cataract mutations although the per locus mutation rate to recessive alleles was observed to be approximately 6 times greater than the per locus mutation rate to dominant alleles. The exclusion of certain classes of lens opacity variant phenotypes, previously demonstrated not to be due to a dominant mutation, from the group of suspected dominant cataract mutations subjected to a genetic confirmation test has greatly improved the efficiency of the test. A total of 23 dominant cataract mutations were confirmed from a group of 67 phenotypic variants. Of the 23 confirmed dominant cataract mutations, 8 were shown to have reduced transmission to the following generation of offspring expressing the mutant phenotype. These results are also consistent with previous results for ethylnitrosourea or radiation treatment in which it was shown that approximately one-third of the recovered mutations have reduced penetrance. One group of dominant cataract mutations, with phenotypic effects on the polar, sub-capsular or corneal regions, is overly represented in the group of recovered mutations with a reduced transmission of offspring expressing the mutant phenotype. Two hypotheses are suggested for this observation, both dependent on the fact that the regions affected indicate that the mutations are expressed later in the development of the eye. Either all carrier individuals have not expressed the phenotype at the time of examination and classification, or later acting mutations are more subject to environmental interactions resulting in more variable expression. Finally, it is argued that a dominant cataract mutation test represents a most practicable protocol to screen for induced dominant mutations in germ cells of the mouse. The imposition of the criterion that suspected variants be subjected to a genetic confirmation test has at least two advantages beside the fact that results represent unambiguous mutational events.(ABSTRACT TRUNCATED AT 400 WORDS)     

'Haldane's Sieve' in a metapopulation: sifting through plant reproductive polymorphisms

An important result of population genetics is that advantageous mutations will be fixed by selection in a population with a greater probability if they are dominant rather than recessive. This selective filter on new variants entering a population, termed 'Haldane's Sieve', has hitherto been invoked to account for the greater role of dominant than completely recessive mutations in adaptive evolution. Here, we suggest that a process similar to Haldane's Sieve will act on migrants into subpopulations of a metapopulation, and that the repeated action of Haldane's Sieve on alleles maintained by frequency-dependent selection, such as those responsible for many plant reproductive polymorphisms, is expected to bias their frequency distribution in favour of dominant alleles. The genetic and phenotypic signatures left by these processes might provide additional indirect support for the contentious idea that metapopulation dynamics have had an important role in shaping the ecology and evolution of some plant species.     

Chloride channel 7 (CLCN7) gene mutations in intermediate autosomal recessive osteopetrosis

Osteopetrosis is a heterogeneous group of inherited disorders that includes a malignant autosomal recessive form, an intermediate autosomal recessive form and autosomal dominant forms of the disease. Most malignant osteopetroses have been ascribed to mutations in the OC116 gene encoding the human a3 subunit of vacuolar H(+)-ATPase. Few cases of autosomal recessive malignant osteopetrosis have been ascribed to mutations in the chloride channel 7 gene (CLCN7), which accounts for all autosomal dominant type II cases reported to date. Up until now, however, nothing has been known regarding the molecular basis of the intermediate form of osteopetrosis (IARO). Our study of two Portuguese IARO families shows that homozygosity for ClCN7 mutations also accounts for this form of osteopetrosis. The two patients presented with spontaneous fractures in the first years of life and generalised increase of bone density. Direct sequencing of the ClCN7 gene in both patients revealed homozygosity for two mutations (G203D and P470Q). We conclude therefore that ClCN7 mutations not only account for some dominant and malignant forms but also for intermediate forms of osteopetrosis.     

Diploids of Clock Mutants of CHLAMYDOMONAS REINHARDI

Diploids constructed from three clock mutants of Chlamydomonas have been analyzed. In the haploid state, these mutations all lengthen the clock period, and they are at separate genetic loci. One is recessive, one dominant, and one is probably incompletely dominant to their respective wild-type alleles.     

The effects of weak selection on neutral diversity at linked sites

The effects of selection on variability at linked sites have an important influence on levels and patterns of within-population variation across the genome. Most theoretical models of these effects have assumed that selection is sufficiently strong that allele frequency changes at the loci concerned are largely deterministic. These models have led to the conclusion that directional selection for selectively favorable mutations, or against recurrent deleterious mutations, reduces nucleotide site diversity at linked neutral sites. Recent work has shown, however, that fixations of weakly selected mutations, accompanied by significant stochastic changes in allele frequencies, can sometimes cause higher diversity at linked sites when compared with the effects of fixations of neutral mutations. This study extends this work by deriving approximate expressions for the mean conditional times to fixation and loss of mutations subject to selection, and analyzing the conditions under which selection increases rather than reduces these times. Simulations are used to examine the relations between diversity at a neutral site and the fixation and loss times of mutations at a linked site that is subject to selection. It is shown that the long-term level of neutral diversity can be increased over the purely neutral value by recurrent fixations and losses of linked, weakly selected dominant or partially dominant favorable mutations, or linked recessive or partially recessive deleterious mutations. The results are used to examine the conditions under which associative overdominance, as opposed to background selection, is likely to operate.     

Citrullination of fibronectin modulates synovial fibroblast behavior

The hyper-IgE syndromes (HIES; originally named Job's syndrome) are a collection of primary immunodeficiency syndromes resulting in elevated serum IgE levels and typified by recurrent staphylococcal skin abscesses, eczema and pulmonary infections. The disorder has autosomal dominant and recessive forms. Autosomal dominant HIES has been shown to be mainly due to STAT3 mutations and additionally results in connective tissue, skeletal, vascular and dental abnormalities. Autosomal recessive HIES has been shown to be mainly due to mutations in DOCK8; these patients are more prone to viral skin infections instead. This review article discusses the common clinical features of the syndrome, the genetic mutations responsible and the pathogenesis of the disease, as well as treatments currently used.     

Genetic Analysis of Mutants of SACCHAROMYCES CEREVISIAE Resistant to the Herbicide Sulfometuron Methyl

Sulfometuron methyl (SM), a potent new sulfonylurea herbicide, inhibits growth of the yeast Saccharomyces cerevisiae on minimal media. Sixty-six spontaneous mutants resistant to SM were isolated. All of the resistance mutations segregate 2:2 in tetrads; 51 of the mutations are dominant, five are semidominant and ten are recessive. The mutations occur in three linkage groups, designated SMR1, smr2 and smr3. Several lines of evidence demonstrate that the SMR1 mutations (47 dominant and four semidominant) are alleles of ILV2 which encodes acetolactate synthase (ALS), the target of SM. First, SMR1 mutations result in the production of ALS enzyme activity with increased resistance to SM. Second, molecular cloning of the ILV2 gene permitted the isolation of mutations in the cloned gene which result in the production of SM-resistant ALS. Finally, SMR1 mutations map at the ILV2 locus. The smr2 mutations (four recessive, two dominant and one semidominant) map at the pdr 1 (pleiotropic drug resistance) locus and show cross-resistance to other inhibitors, typical of mutations at this locus. The smr3 mutations (six recessive and two dominant) define a new gene which maps approximately midway between ADE2 and HIS3 on the right arm of chromosome XV.     

Myotonia congenita: novel mutations in CLCN1 gene

Myotonia congenita belongs to the group of non-dystrophic myotonia caused by mutations of CLCN1gene, which encodes human skeletal muscle chloride channel 1. It can be inherited either in autosomal dominant (Thomsen disease) or recessive (Becker disease) forms. Here we have sequenced all 23 exons and exon-intron boundaries of the CLCN1 gene, in a panel of 5 unrelated Chinese patients with myotonia congenita (2 with dominant and 3 with recessive form). In addition, detailed clinical analysis was performed in these patients to summarize their clinical characteristics in relation to their genotypes. Mutational analyses revealed 7 different point mutations. Of these, we have found 3 novel mutations including 2 missense (R47W, V229M), one splicing (IVS19+2T>C), and 4 known mutations (Y261C,G523D, M560T, G859D). Our data expand the spectrum of CLCN1 mutations and provide insights for genotype–phenotype correlations of myotonia congenita in the Chinese population.     

Genetic analysis of hypertropic mutants of Phycomyces

A new class of Phycomyces behavioral mutants with enhanced tropic responses has been analyzed genetically to determine the number of genes involved and the nature of their expression. These hypertropic mutants carry pleiotropic nuclear mutations. Besides their effects on sensory behavior, they also affect morphology and meiotic processes. Behavioral analyses of heterokaryons containing hypertropic and wild-type nuclei in varying proportions show that the hypertropic mutations in strains L82, L84, L86, and L88 are strongly dominant. Conversely, the hypertropic mutations carried by the strains L83, L85, and L87 are strongly recessive. We performed recombination analyses between hypertropic mutants and mutants with diminished phototropism, affected in the seven genes madA to madG. We found no evidence of linkage between the hypertropic mutations and any of these mad mutations. From crosses, we isolated double mutants carrying hypertropic mutations together with madC (night blind) and madG (stiff) mutations. The behavioral phenotypes of the double mutants are intermediate between those of the parentals. Complementation analyses show that the three recessive hypertropic mutations affect the same gene, which we call madH. The expression of the recessive hypertropic allele becomes dominant in heterokaryons carrying madC and madH nuclei; the madC gene has been implicated separately with the photoreceptor at the input to the sensory pathway, while the madH gene is associated with the growth control output. This result suggests the physical interaction of both gene products, madH and madC, in a molecular complex for the photosensory transduction chain.     

Myotonia congenita: novel mutations in CLCN1 gene

Myotonia congenita belongs to the group of non-dystrophic myotonia caused by mutations of CLCN1gene, which encodes human skeletal muscle chloride channel 1. It can be inherited either in autosomal dominant (Thomsen disease) or recessive (Becker disease) forms. Here we have sequenced all 23 exons and exon-intron boundaries of the CLCN1 gene, in a panel of 5 unrelated Chinese patients with myotonia congenita (2 with dominant and 3 with recessive form). In addition, detailed clinical analysis was performed in these patients to summarize their clinical characteristics in relation to their genotypes. Mutational analyses revealed 7 different point mutations. Of these, we have found 3 novel mutations including 2 missense (R47W, V229M), one splicing (IVS19+2T>C), and 4 known mutations (Y261C,G523D, M560T, G859D). Our data expand the spectrum of CLCN1 mutations and provide insights for genotype–phenotype correlations of myotonia congenita in the Chinese population.     

Dominant cataract and recessive specific locus mutations in offspring of X-irradiated male mice

Male mice were X-irradiated with 3.0 + 3.0 Gy or 5.1 + 5.1 Gy (fractionation interval 24 h). The offspring were screened for dominant cataract and recessive specific locus mutations. In the 3.0 + 3.0-Gy spermatogonial treatment group, 3 dominant cataract mutations were confirmed in 15 551 offspring examined and 29 specific locus mutations were recovered in 18 139 offspring. In the post-spermatogonial treatment group, 1 dominant cataract mutation was obtained in 1120 offspring and 1 recessive specific locus mutation was recovered in 1127 offspring. The induced mutation rate per locus, per gamete, per Gy calculated for recessive specific locus mutations is 2.0 X 10(-5) in post-spermatogonial stages and 3.7 X 10(-5) in spermatogonia. For dominant cataract mutations, assuming 30 loci, the induced mutation rate is 5.0 X 10(-6) in the post-spermatogonial stages and 1.1 X 10(-6) in spermatogonia. In the 5.1 + 5.1-Gy spermatogonial treatment group, 3 dominant cataract mutations were obtained in 11 205 offspring, whereas in 13 201 offspring 27 recessive specific locus mutations were detected in the spermatogonial group. In the post-spermatogonial treatment group no dominant cataract mutation was observed in 425 offspring and 2 recessive specific locus mutations were detected in 445 offspring. The induced mutation rate per locus, gamete and Gy in spermatogonia for recessive specific locus mutations is 2.8 X 10(-5) and for dominant cataract mutations 0.9 X 10(-6). In post-spermatogonial stages, the mutation rate for recessive specific locus alleles is 6.2 X 10(-5). In the concurrent untreated control group, in 11 036 offspring no dominant cataract mutation and in 23 518 offspring no recessive specific locus mutation was observed. Litter size and the number of carriers at weaning have been determined in the confirmation crosses of the obtained dominant cataract mutants as indicators of viability and penetrance effects. Two mutants had a statistically significantly reduced litter size and one mutant had a statistically significantly reduced penetrance.