New Class Of Mutations Research Articles

Engineering tyrosine residues into hemoglobin enhances heme reduction, decreases oxidative stress and increases vascular retention of a hemoglobin based blood substitute

Hemoglobin (Hb)-based oxygen carriers (HBOC) are modified extracellular proteins, designed to replace or augment the oxygen-carrying capacity of erythrocytes. However, clinical results have generally been disappointing due to adverse side effects, in part linked to the intrinsic oxidative toxicity of Hb. Previously a redox-active tyrosine residue was engineered into the Hb β subunit (βF41Y) to facilitate electron transfer between endogenous antioxidants such as ascorbate and the oxidative ferryl heme species, converting the highly oxidizing ferryl species into the less reactive ferric (met) form. We inserted different single tyrosine mutations into the α and β subunits of Hb to determine if this effect of βF41Y was unique. Every mutation that was inserted within electron transfer range of the protein surface and the heme increased the rate of ferryl reduction. However, surprisingly, three of the mutations (βT84Y, αL91Y and βF85Y) also increased the rate of ascorbate reduction of ferric(met) Hb to ferrous(oxy) Hb. The rate enhancement was most evident at ascorbate concentrations equivalent to that found in plasma (< 100 μM), suggesting that it might be of benefit in decreasing oxidative stress in vivo. The most promising mutant (βT84Y) was stable with no increase in autoxidation or heme loss. A decrease in membrane damage following Hb addition to HEK cells correlated with the ability of βT84Y to maintain the protein in its oxygenated form. When PEGylated and injected into mice, βT84Y was shown to have an increased vascular half time compared to wild type PEGylated Hb. βT84Y represents a new class of mutations with the ability to enhance reduction of both ferryl and ferric Hb, and thus has potential to decrease adverse side effects as one component of a final HBOC product.

A Screen for rfaH Suppressors Reveals a Key Role for a Connector Region of Termination Factor Rho.

ABSTRACTRfaH activates horizontally acquired operons that encode lipopolysaccharide core components, pili, toxins, and capsules. Unlike its paralog NusG, which potentiates Rho-mediated silencing, RfaH strongly inhibits Rho. RfaH is recruited to its target operons via a network of contacts with an elongating RNA polymerase (RNAP) and a specific DNA element called ops to modify RNAP into a pause- and NusG-resistant state. rfaH null mutations confer hypersensitivity to antibiotics and detergents, altered susceptibility to bacteriophages, and defects in virulence. Here, we carried out a selection for suppressors that restore the ability of a ΔrfaH mutant Escherichia coli strain to grow in the presence of sodium dodecyl sulfate. We isolated rho, rpoC, and hns suppressor mutants with changes in regions previously shown to be important for their function. In addition, we identified mutants with changes in an unstructured region that connects the primary RNA-binding and helicase domains of Rho. The connector mutants display strong defects in vivo, consistent with their ability to compensate for the loss of RfaH, and act synergistically with bicyclomycin (BCM), which has been recently shown to inhibit Rho transformation into a translocation-competent state. We hypothesize that the flexible connector permits the reorientation of Rho domains and serves as a target for factors that control the motor function of Rho allosterically. Our results, together with the existing data, support a model in which the connector segment plays a hitherto overlooked role in the regulation of Rho-dependent termination.

Genetic Mutations in Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a clonal malignancy characterized by ineffective hematopoiesis. Most AML patients present different cytogenetic and molecular defects associated with certain biologic and clinical features of the disease. Approximately 50-60 % of de novo AML and 80-95 % of secondary AML patients demonstrate chromosomal aberrations. Structural chromosomal aberrations are the most common cytogenetic abnormalities in about of 40 % of de novo AML patients. A relatively large group of intermediate risk patients with cytogenetically normal (CN) AML demonstrates a variety of outcomes. Current AML prognostic classifications include only some mutations with known prognostic value, namely NPM1, FLT3 and C/EBPa. Patients with NPM1 mutation, but without FLT3-ITD or C/EBPa mutations have a favorable prognosis, whereas patients with FLT3-ITD mutation have a poor prognosis. A new class of mutations affecting genes responsible for epigenetic mechanisms of genome regulations, namely for DNA methylation and histone modification, was found recently. Among them, mutations in genes DNMT3A, IDH1/2, TET2 and some others are the most well-studied mutations to date. A number of studies demonstrated an unfavorable prognostic effect of the DNMT3A mutation in AML. The prognostic significance of the IDH1/2 gene is still unclear. The prognosis is affected by a number of biological factors, including those associated with cytogenetic aberrations and other mutations, especially FLT3 and NPM1. The number of studies of genetic mutations in AML keeps growing. The data on genetic aberrations in AML obtained to date confirm their role in the onset and development of the disease.

Genes and Ontogenes in Drosophila: The Role of RNA Forms

The independent hereditary factors, such as Mendelian genes, are not sufficient for the existence and operation of genetic systems. The hereditary factors of different type were searched for. A new class of mutations, referred to as conditional mutations, was discovered in Drosophila melanogaster. Such a mutation dies in a restrictive genotype but survives and reproduces in a permissive genotype. Besides their conditional nature, mutations in a permissive genotype display a set of specific features that drastically distinguish them from conventional mutations, namely, they (1) are dominant; (2) are as a rule, lethal; (3) have drastically decreased fertility; (4) interact with chromosomal rearrangements; (5) switch the genome from a stable to an unstable state; (6) increase the basal metabolism; (7) induce modifications and morphoses; and (8) their manifestation is inherited in a parental manner. Four properties of these mutations-conditional manifestation together with (1), (4), and (8) suggest that the mutant genes (1) are segments of DNA; (2) their products are RNA duplexes (3) active in germ cells and (4) repeated in the genome. Emergence of morphoses in mutants suggests that the genes are involved in the control of ontogeny. Correspondingly, these genes were named ontogenes. Thus, the genetic system comprises the genes working according to a DNARNA- protein script and the ontogenes following a DNA-RNA script. The first entity is engaged in production of the building material for the organism, proteins, while the second entity controls this process during preparation of the individual developmental program. These different functions of genes depend on the type of transcript formed from DNA as well as the time and place of its origin on DNA.

Functionally Relevant RNA Helicase Mutations in Familial and Sporadic Myeloid Malignancies

In this issue of Cancer Cell, Polprasert and colleagues identified recurrent mutations in the DEAD/H-box RNA helicase gene DDX41 in familial and acquired cases of myelodsyplasia and acute myeloid leukemia. These mutations induce defects in RNA splicing and represent a new class of mutations in myeloid malignancies.

Processed pseudogenes acquired somatically during cancer development

Cancer evolves by mutation, with somatic reactivation of retrotransposons being one such mutational process. Germline retrotransposition can cause processed pseudogenes, but whether this occurs somatically has not been evaluated. Here we screen sequencing data from 660 cancer samples for somatically acquired pseudogenes. We find 42 events in 17 samples, especially non-small cell lung cancer (5/27) and colorectal cancer (2/11). Genomic features mirror those of germline LINE element retrotranspositions, with frequent target-site duplications (67%), consensus TTTTAA sites at insertion points, inverted rearrangements (21%), 5′ truncation (74%) and polyA tails (88%). Transcriptional consequences include expression of pseudogenes from UTRs or introns of target genes. In addition, a somatic pseudogene that integrated into the promoter and first exon of the tumour suppressor gene, MGA, abrogated expression from that allele. Thus, formation of processed pseudogenes represents a new class of mutation occurring during cancer development, with potentially diverse functional consequences depending on genomic context.

X-linked sideroblastic anemia due to ALAS2 intron 1 enhancer element GATA-binding site mutations.

X-linked sideroblastic anemia (XLSA) is the most common form of congenital sideroblastic anemia. In affected males, it is uniformly associated with partial loss-of-function missense mutations in the erythroid-specific heme biosynthesis protein 5-aminolevulinate synthase 2 (ALAS2). Here, we report five families with XLSA owing to mutations in a GATA transcription factor binding site located in a transcriptional enhancer element in intron 1 of the ALAS2 gene. As such, this study defines a new class of mutations that should be evaluated in patients undergoing genetic testing for a suspected diagnosis of XLSA.

Pure intronic rearrangements leading to aberrant pseudoexon inclusion in dystrophinopathy: a new class of mutations?

We report on two unprecedented cases of pseudoexon (PE) activation in the DMD gene resulting from pure intronic double-deletion events that possibly involve microhomology-mediated mechanisms. Array comparative genomic hybridization analysis and direct genomic sequencing allowed us to elucidate the causes of the pathological PE inclusion detected in the RNA of the patients. In the first case (Duchenne phenotype), we showed that the inserted 387-bp PE was originated from an inverted ∼57 kb genomic region of intron 44 flanked by two deleted ∼52 kb and ∼1 kb segments. In the second case (Becker phenotype), we identified in intron 56 two small deletions of 592 bp (del 1) and 29 bp (del 2) directly flanking a 166-bp PE located in very close proximity (134 bp) to exon 57. The key role of del 1 in PE activation was established by using splicing reporter minigenes. However, the analysis of mutant constructs failed to identify cis elements that regulate the inclusion of the PE and suggested that other splicing regulatory factors may be involved such as RNA structure. Our study introduces a new class of mutations in the DMD gene and emphasizes the potential role of underdetected intronic rearrangements in human diseases.

IDH1 Mutations Are Detected in 9.3% of All AML and Are Strongly Associated with Intermediate Risk Karyotype and Unfavourable Prognosis: a Study of 999 Patients

Abstract LBA-3▪▪This icon denotes an abstract that is clinically relevant. Introduction:IDH1 is the gene coding for the soluble isocitrate dehydrogenease 1 (NADP+), which catalyzes the oxidative decaroxylation of isocitrate to 2-oxoglutarate. The gene has been shown to be frequently mutated in high-grade gliomas at residue p.R132, which is located in the substrate binding site of IDH1. So far, several other tumors have been analyzed without detection of the respective mutation (Bleeker et al., Human Mutation 2009). However, recently a next generation sequencing project found IDH1 mutated in 8.5% of AML with normal karyotype (Mardis et al., NEJM, 2009). Aim:To further evaluate the importance of IDH1R132 (IDH1mut) in AML we have analyzed a cohort of 999 comprehensively characterized AML cases. Methods:The respective base exchange was analysed by a LightCycler based melting curve assay with subsequent sequencing of the positive samples. Results:The cohort was comprised of 536 male and 463 female patients (median age: 65.9 years; range: 17.1- 93.3 years). 833 had de novo AML (83.4%), 122 AML following MDS (s-AML,12.1%) and 44 AML after previous treatment of different malignancies (t-AML, 4.4%). Karyotype was available in all cases: 681 had a normal karyotype (NK) AML, and 319 had chromosomal aberrations (t(15;17): n=29; inv(16): n=12, t(8;21): n=23, t(11q23): n=10, t(6;9): n=4, inv(3): n=3; -7: n=27, +8: n=29, +13: n=11, -Y: n=4; complex aberrant: n=60, others: n=106). Overall, in 93 pts IDH1 p.R132 mutations were detected (9.3%). Five different amino acid exchanges were observed: R132C (n=49), R132L (n=22), R132 H and G (n=7, each), and R132S (n=5). With respect to history of the patient IDH1mut were found in 80/833 of de novo AML (9.6%), 11/122 (9.0%) of s-AML, and 2/44 (4.5%) of t-AML, respectively (n.s.). More females (57/463, 12.3%) than males (36/536; 6.7%) had IDH1mut (p=0.003). Age was slightly higher in the mutated cases (63.9 vs. 61.9 years, n.s.). No differences were found for WBC count. IDH1mut were distributed differently between karyotypes: in NK 69/681 (10.1%) and in aberrant karyotypes 24/318 (7.5%). However, IDH1 was never mutated in inv(16), t(8;21), t(6;9), t(11q23), inv(3), or in complex aberrant karyotypes (n=112). In 2 of 27 cases (7.4%) with t(15;17) an IDH1 mutation was detected. Thus, the IDH1 mututations clustered in the intermediate risk karyotype group in comparison to the good or poor risk groups (91/771; 11.8% vs 2/134 (1.5%), p<0.001). The cohort was also characterized for several other molecular mutations. FLT3-ITD was present in 22% (212/954), FLT3-TKD in 6.7% (33/496), NPM1 in 35.4% (329/929), NRAS in 14.6% (48/328), MLL-PTD in 6.9% (64/932), CEBPA mutations in 7.4% (48/645) and RUNX1 mutations in 33.0% (99/299) of analysed cases, respectively. IHD1 mutations were found to be more frequent in NPM1 mutated than in NPM1wt cases (41/329; 12.4% vs 48/598; 8.0%, p= 0.019) and in those with MLL-PTD (11/64; 17.2% vs 77/867; 8.9%, p= 0.031). With lower frequencies IDH1mut were also detected together with RUNX1 mutations (n=8/99), CEBPA mutations (n=2/48), NRAS mutations (n=7/48), and FLT3-TKD (n=1/33). IDH1 was similarly distributed between FLT3-ITD mutated and unmutated cases (18/212; 8.5% vs. 72/744; 9.7%). In 22 (23.7%) of all IDH1mut AML no additional mutation was detected, whereas in 48 (51.6%) one additional, in 22 (23.7%) two additional and in one case three additional mutations were found. An unfavourable effect of IDH1mut on event free survival (EFS) was observed in the total group (median: 272 vs. 456 days; p=0.007) as well as in those with intermediate risk karyotype (median: 272 vs. 449 days; p=0.008). A shorter EFS of the IDH1mut was particularly seen in the NPM1wt cohort (median: 244 vs. 375 days; p=0.038) with a strong trend for an independent effect in a multivariate analysis (p=0.089). Conclusions:IDH1 mutations are frequent in AML and are prognostically unfavourable especially in the NPM1wt cohort. IDH1 mutations seem to be a new class of mutation probably complementing with the classical type 1 and type 2 mutations. Disclosures:Schnittger:MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership. Ulke:MLL Munich Leukemia Lab: Employment. Kaya:MLL Munich Leukemia Lab: Employment. Weiss:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership.

First genomic rearrangement of the RYR1 gene associated with an atypical presentation of lethal neonatal hypotonia

Neonatal hypotonia is frequently observed with a highly variable clinical presentation. Congenital myopathies that are classically characterized by the presence of structural changes of the muscle fibres such as cores, rods and aggregates have been reported to be occasionally associated with this presentation. However, the identification of the causing defect can be a challenging task in severe neonatal forms of the disease since specific structural changes might not always be present in affected newborn’s muscles. The RYR1 gene encodes the skeletal muscle isoform of a calcium channel, the ryanodine receptor, and has been involved in both dominant and recessive congenital myopathies associated with structural changes and presenting with various degree of severity. Here we report the case of a child presenting at birth with a lethal form of neonatal hypotonia associated with an atypical congenital myopathy. Molecular investigations showed that the disease was caused by two novel RYR1 mutations. One of the mutations was a large-sized genomic deletion. This is the first genomic rearrangement identified into the RYR1 gene to our knowledge. This new class of mutation of the RYR1 gene will clearly have consequences for the molecular investigation of RYR1-related diseases.

BCR-ABL truncation due to premature translation termination as a mechanism of resistance to kinase inhibitors

7028 Background: The major mechanism underlying imatinib resistance in patients with chronic myeloid leukemia (CML) is clonal expansion of leukemic cells with point mutations in the BCR-ABL tyrosine kinase. We describe three novel ABL premature termination mutations leading to BCR-ABL truncation in leukemia patients with multidrug (imatinib/nilotinib/dasatinib) resistance. Methods: Peripheral blood or bone marrow samples from drug-resistant CML patients were collected. Total nucleic acids were purified and subjected to two rounds of PCR analysis, with the first PCR designed to eliminate amplification of the wild-type, non-translocated ABL gene. Bi-directional sequencing was then performed. HL60 cells (a Ph-negative myeloid leukemia cell line) and peripheral blood of healthy subjects were used as negative controls; a human CML cell line (K562) was used as a positive control. Results: We identified an exon 7 deletion in three CML patients, a 4-nt insertion (908insCAGG) near the exon 5/6 junction in one CML case, and an exon 6 point mutation (997C&gt;T) in one patient with acute lymphoblastic leukemia (ALL). These mutations all create premature stop codons and cause termination at residues 381, 315, and 333, respectively, leading to truncated proteins with only the first quarter of the kinase domain (P-loop) or lacking the C-terminus of ABL including the A-loop. Conclusions: These novel mutations, and the previously documented 35-nt insertion in exon 8, may constitute a new class of mutations that 1) cause truncation of the BCR-ABL kinase; (2) abolish the regulatory element in the ABL kinase domain and the downstream C-terminal region; and (3) confer significant drug resistance. [Table: see text]

A Protein Complex Restrains a Homicidal Enzyme during Sperm Differentiation

A Protein Complex Restrains a Homicidal Enzyme during Sperm Differentiation

Mutation spectrum in UVB-exposed skin epidermis of a mildly-affectedXpg-deficient mouse

A C-terminal 183 amino acid-truncated mutation of the mouse Xpg gene (XpgDeltaex15) gives rise to a partial deficiency in nucleotide excision repair in homozygously affected cells. We studied the effect of this mutation on UVB-induced mutagenesis in mouse skin, using transgenic mice harboring lambda-phage-based bacterial lacZ genes as a mutational reporter. UVB increased the lacZ mutant frequency in the epidermis moderately in the homozygous mutant mice, but significantly higher than in the wild-type or the heterozygous mice, whereas background mutant frequencies were not appreciably different among the three mouse genotypes. Ninety-eight lacZ mutant sequences isolated from the UVB-exposed epidermis of the XpgDeltaex15-homozygous mice were analyzed and compared with mutant sequences from the wild-type mice. The spectra of the mutations in the two mouse genotypes were not significantly different, and they were highly UV-specific. There were frequent C --> T transitions at dipyrimidine sites and several CC --> TT tandem mutations, although the UV-specific mutations occurred more frequently at CpG sites in the mutant mice. The distribution of the mutations observed in the lacZ transgene and the preferred sequence context of the UV-specific C --> T mutations (5'-TC-3' > 5'-CC-3' > 5'-CT-3') in the Xpg-mutant mice were similar to those found in the wild-type mice. Despite these similarities, we detected a previously unrecognized type of the UV-induced mutation only in the Xpg mutant (6/98 in the mutation spectrum of the mutant vs. 0/76 in the wild-type; P = 0.035), which is characterized by multiple base substitutions or frameshifts within a three-nucleotide sequence containing a dipyrimidine. We propose that this putative new class of mutation, which we refer to as a "triplet mutation", is characteristic of UV-induced mutation in an excision-repair-deficient background.

21 A new class of mutations in CFTR specifically target the pancreas and cause idiopathic pancreatitis through interacting with SPINK1 mutations

21 A new class of mutations in CFTR specifically target the pancreas and cause idiopathic pancreatitis through interacting with SPINK1 mutations

A new class of mutations reveals a novel function for the original phosphatidylinositol 3-kinase binding site.

Previous studies have demonstrated that the specificity of Src homology 2 (SH2) and phosphotyrosine-binding domain interactions are mediated by phosphorylated tyrosines and their neighboring amino acids. Two of the first phosphotyrosine-based binding sites were found on middle T antigen of polyoma virus. Tyr-250 acts as a binding site for ShcA, whereas Tyr-315 forms a binding site for the SH2 domain of the p85 subunit of phosphatidylinositol 3-kinase. However, genetic analysis of a given phosphotyrosine's role in signaling can be complicated when it serves as a binding site for multiple proteins. The situation is particularly difficult when the phosphotyrosine serves as a secondary binding site for a protein with primary binding determinates elsewhere. Mutation of a tyrosine residue to phenylalanine blocks association of all bound proteins. Here we show that the mutation of the amino acids following the phosphorylated tyrosine to alanine can reveal phosphotyrosine function as a secondary binding site, while abrogating the phosphotyrosine motif's role as a primary binding site for SH2 domains. We tested this methodology by using middle T antigen. Our results suggest that Tyr-250 is a secondary binding site for phosphatidylinositol 3-kinase, whereas Tyr-315 is a secondary binding site for a yet-to-be-identified protein, which is critical for transformation.

Lesions in the nuo operon, encoding NADH dehydrogenase complex I, prevent PurF-independent thiamine synthesis and reduce flux through the oxidative pentose phosphate pathway in Salmonella enterica serovar typhimurium.

In Salmonella enterica serovar Typhimurium, PurF-independent thiamine synthesis (or alternative pyrimidine biosynthesis) allows strains, under some growth conditions, to synthesize thiamine in the absence of the first step in the purine biosynthetic pathway. Mutations have been isolated in a number of loci that prevent this synthesis and thus result in an Apb(-) phenotype. Here we identify a new class of mutations that prevent PurF-independent thiamine synthesis and show that they are defective in the nuo genes, which encode the major, energy-generating NADH dehydrogenase of the cell. Data presented here indicated that a nuo mutant has reduced flux through the oxidative pentose phosphate pathway that may contribute to, but is not sufficient to cause, the observed thiamine requirement. We suggest that reduction of the oxidative pentose phosphate pathway capacity in a nuo mutant is an attempt to restore the ratio between reduced and oxidized pyridine nucleotide pools.

Transglutaminase 1 Mutations in Lamellar Ichthyosis: LOSS OF ACTIVITY DUE TO FAILURE OF ACTIVATION BY PROTEOLYTIC PROCESSING

Lamellar ichthyosis is a congenital recessive skin disorder characterized by generalized scaling and hyperkeratosis. It is caused by mutations in the TGM1 gene that encodes the transglutaminase 1 (TGase 1) enzyme, which is critical for the assembly of the cornified cell envelope in terminally differentiating keratinocytes. TGase 1 is a complex enzyme existing as both cytosolic and membrane-bound forms. Moreover, TGase 1 is proteolytically processed, and the major functionally active form consists of a membrane-bound 67/33/10-kDa complex with a myristoylated and palmitoylated amino-terminal 10-kDa membrane anchorage fragment. To understand better how point mutations, deletions, and truncations found in lamellar ichthyosis disease affect the structure and function of TGase 1, we have expressed in baculovirus and keratinocytes a number of reported TGase 1 mutants. The structural implications of these mutations were examined using a homology-derived three-dimensional model of TGase 1 generated from the known x-ray structure of the related coagulation factor XIIIa enzyme. The present studies demonstrate that loss of TGase 1 activity is not restricted to mutations that directly affect the enzymatic activity. We report a new class of mutations that impair the subsequent post-synthetic processing of the protein into its highly active functional forms.

A distamycin A-inducible fragile site, FRA8E, located in the region of the hereditary multiple exostoses gene, is not involved in HPV16 DNA integration and amplification

The rare fragile site is a specific point on a chromosome that is expressed as an isochromatid gap or break under certain conditions of cell culture and is inherited in a Mendelian codominant fashion. Five folate-sensitive fragile sites were cloned, and the molecular basis of fragile site mutation was shown to be a new class of mutation, called dynamic mutation, resulting from an allelic expansion of (CCG)n repeats. The mechanism responsible for other types of rare fragile sites, i.e., distamycin A-inducible and BrdU-requiring, is unknown, although cytogenetic studies suggested that these fragile sites play a mechanistic role in breakage and recombination and may also be integration and modification sites of foreign viral DNA genomes. A distamycin A-inducible fragile site, FRA8E, is mapped to 8q24.1 in which various loci implicated in genomic instability are located. Here we identified a YAC clone spanning both FRA8E and the hereditary multiple exostosis (EXT1) gene, using fluorescence in situ hybridization (FISH) analysis of a yeast artificial chromosome (YAC) contig. By using P1 clones as probes, the FRA8E locus was further localized to a 400-kb region including the EXT1 gene. Furthermore, the integration and amplification site of human papillomovirus 16 DNA in the ASCC (argyrophil small cell carcinoma) cells were shown not to coincide with FRA8E, but to be involved in an extensively broad genomic region of 8q24.1, including the c-myc gene.

The CAG/polyglutamine tract diseases: gene products and molecular pathogenesis.

In the past few years, a new type of genetic mutation, expansion of trinucleotide repeats, has been shown to cause neurologic disease. This new class of mutations was first identified in 1991 as the underlying genetic defect in spinal and bulbar muscular atrophy and the fragile X syndrome, and in recent years, trinucleotide repeat expansions have been found to be the causative mechanism in 10 other neurologic diseases. These mutations are produced by heritable unstable DNA and are termed "dynamic mutations" because of changes in the number of repeat units inherited from generation to generation. In the normal population, these repeat units, although polymorphic, are stably inherited. To date four types of trinucleotide repeat expansions have been identified: (1) long cytosine-guanine-guanine (CGG) repeats in the two fragile X syndromes (FRAXA and FRAXE), (2) long cytosine-thymine-guanine (CTG) repeat expansions in myotonic dystrophy, (3) long guanine-adenine-adenine repeat expansions in Friedreich's ataxia and (4) short cytosine-adenine-guanine repeat expansions (CAG) which are implicated in eight neurodegenerative disorders and are the focus of this review. Diseases that are caused by trinucleotide repeat expansions exhibit a phenomenon called anticipation that can not be explained by conventional Mendelian genetics. Anticipation is defined as increase in the severity of disease with an earlier age of onset of symptoms in successive generations. Anticipation is often influenced by the sex of the transmitting parent, and for most CAG repeat disorders, the disease is more severe when paternally transmitted. The severity and the age of onset of the disease have been correlated with the size of the repeats on mutant alleles, with the age of onset being inversely correlated with the size of the expansion. In all eight disorders caused by CAG repeat expansion, the repeat is located within the coding region of the gene involved and in all cases it is translated into a stretch of polyglutamines in the respective proteins. All the proteins are unrelated outside of the polyglutamine stretch and most are novel with exception of the androgen receptor and the voltage gated alpha 1A calcium channel, which are mutated in spinal and bulbar muscular atrophy and spinocerebellar ataxia type 6. It is intriguing that the proteins are ubiquitously expressed in both peripheral and nervous tissue but in each disorder only a select population of nerve cells are targeted for degeneration as a consequence of the expanded CAG repeat. Current thinking among scientists working on the molecular mechanisms of neurodegeneration in these diseases is that the presence of an expanded polyglutamine confers a gain of function onto the involved protein. To understand the mechanisms underlying the pathogenesis of these diseases, investigators have turned to generating transgenic mice which recapitulate some of the features of the human disease and hence are excellent model systems to study the progression of the disease in vivo.

Clinical aspects of CAG repeat diseases.

Seven neurodegenerative disorders are known to be caused by unstable expansions of the trinucleotide CAG within human genes, and more will be discovered in the coming years. These disorders share some clinical similarities, as well as some differences, which are summarized here. These diseases have unusual clinical genetic properties related to the dynamic nature of CAG repeat expansions, including instability of the repeat expansion in meiosis, particularly male meiosis; a strong correlation between onset age and size of the repeat expansion; anticipation (earlier disease onset in succeeding generations); new mutations arising from unstable, mutable alleles with a high-normal CAG repeat number; and reduced penetrance for alleles in the low-affected range. Much more remains to be learned about the molecular biology and clinical pathophysiology of this new class of genetic diseases.