AGG Interspersion Patterns Research Articles

Screening for Fragile X Syndrome Among Filipino Children with Autism Spectrum Disorder.

Individuals with autism spectrum disorder present with difficulties in social communication, restricted interests or behaviors and other co-morbidities. About 2 to 10% of cases of autism have a genetic cause, and Fragile X Syndrome (FXS) is reported in 0 to 6.5% of individuals with autism. However, the FXS and premutation prevalence among Filipino children has never been reported. The aim of the study was to establish the presence of FXS or premutation carriers among Filipino children with autism and to describe the phenotypic characteristic of cases identified. Blood was collected from 235 children aged 2-6 years old and diagnosed with autism. Samples were analyzed using PCR methods to amplify CGG repeats in the FMRI gene. The diagnosis of autism was confirmed through the Autism Diagnostic Observation Schedule-2. Additional characteristics were documented from a physical examination, Griffiths Scales of Child Development assessment and a parent-answered questionnaire using the Vineland Adaptive Behavior Scale. Fragile X testing through PCR methods in 235 children with diagnosed autism showed 220 (93.6%) were negative, no full mutations, 1 (0.436%) premutation carrier and 14 (5.95%) cases contained intermediate alleles. The FXS testing was limited to confirmed cases of autism, which is considered a high-risk group and does not provide prevalence for the general Filipino population. Subjects were self-referred or referred by clinicians, which may not represent the Filipino autism population with a bias towards those with means for clinical consultations and ability to travel to the place of testing. Samples were not measured for mosaicism, DNA methylation or AGG interspersion patterns. These may have effects on the CGG repeat expansion and overall presentation of FXS. Findings from a single premutation carrier cannot characterize features distinctly present in Filipinos with the mutation. Nevertheless, these results support the data that the prevalence of FXS in Asian populations may be lower than non-Asian populations. This can contribute to a better understanding of FXS and genetic causes of autism in the Philippines and other Asian populations.

Analysis of AGG interspersion of the FMR1 gene in fragile X syndrome

To analyze (CGG)n repeats sequence and AGG interspersion correlated with unstable expansion of FMR1 gene in a general Chinese population. AmplideX FMR1 PCR Kit was used to amplify 380 X chromosomes from randomly selected 176 males and 102 females, 11 permutation carriers and 10 full mutation patients have served as controls. Results of capillary electrophoresis were analyzed with GeneMapper software Version 4.0. SPSS 11.0 software was used for statistical analysis. The ratio of heterozygous females was 64.70%. The number of alleles in general males and females was 15 and 14, the classes of AGG pattern was 26 and 27, respectively. The range of alleles was between 17 to 45 CGG repeats in males and 21 to 44 CGG repeats in females, and 1 male was identified as gray zone carrier. The most frequent allele was 29 CGG repeats, which was followed by 30 and 36 repeats, while 28 CGG repeats were absent. The most common AGG pattern was 9A9A9, 99.21% of the population was detected with different forms and numbers of AGG interruption, and 6A interruption pattern was found in 10.02% samples especially in individuals with more CGG repeats. However, 57.58% of control samples had no AGG interruption, and none of the controls had 6A interruption pattern. No significant difference was observed in allele frequent distribution of (CGG)n repeats and AGG interspersion patterns between the males and females (P > 0.05), and AGGs was significantly different between general population and controls (P < 0.05). AGGs and AGG pattern may have important roles in maintaining (CGG)n stability in general population of China, 9A9A6A9 may be a special pattern for preventing (CGG)n unstable expansion in Asian populations.

Fragile X Syndrome in Mentally Retarded Patients from Latvia

Fragile X Syndrome in Mentally Retarded Patients from Latvia The aim of this study was to estimate the prevalence of FXS in Latvia and characterise the FMR1 CGG-repeat structure in Latvian patients exhibiting mental retardation. A group of 352 unrelated patients with mental retardation (MR) referred from clinical geneticists was screened by PCR for the normal allele. In a sample of 245 chromosomes the CGG repeat number was determined by Applied Biosystems protocol on ABI Prism 310. Prevalence of 29, 30, and 31 CGG repeats was found for the normal allele. Five affected patients were detected (detection rate 2.56%). AGG interspersion pattern analysis showed stability of transmission to the next generation for 12 intermediate alleles. The found detection rate of FXS in our survey among MR patients was similar to the detection rate reported in literature. Taking into account the number of confirmed FXS cases we suggest that FXS is still clinically unrecognized in paediatrician practice.

Fragile X Gene Stability in Basque Valleys: Prevalence of Premutation and Intermediate Alleles

Fragile X syndrome is the most common form of inherited mental retardation. The molecular basis is usually the unstable expansion of a CGG repeat in the FMR1 gene. We previously analyzed a sample of two Basque valleys. In the present work we extend the study to another five isolated valleys. The results show that differences in factors implicated in CGG repeat instability—CGG repeat size, XS548/FRAXAC1 haplotypes, and AGG interspersion pattern—are present in the Basque populations analyzed.

Fragile X Gene Stability in Basque Valleys: Prevalence of Premutation and Intermediate Alleles

Fragile X syndrome is the most common form of inherited mental retardation. The molecular basis is usually the unstable expansion of a CGG repeat in the FMR1 gene. We previously analyzed a sample of two Basque valleys. In the present work we extend the study to another five isolated valleys. The results show that differences in factors implicated in CGG repeat instability--CGG repeat size, XS548/FRAXAC1 haplotypes, and AGG interspersion pattern-are present in the Basque populations analyzed.

Haplotype and AGG Interspersion Analysis of FMR1 Alleles in a Croatian Population: No Founder Effect Detected in Patients with Fragile X Syndrome

Several studies have suggested that fragileX syndrome (FRAXA), the most common inherited form of mental retardation, originated from a limited number of founder chromosomes. The aim of this study is to assess the genetic origin of fragile X syndrome in a Croatian population. We performed a haplotype analysis of the polymorphic loci DXS548 and FRAXAC1 in 18 unrelated fragile X and 56 control chromosomes. The AGG interspersion pattern of the FMR1 CGG repeat region was analyzed by sequencing. This is the first report on haplotype and AGG interspersion analysis of the fragile X syndrome gene in a Croatian population—the only eastern European population of Slavic origin analyzed so far. Our findings are intriguing, because they show a distinct distribution of the DXS548 and FRAXAC1 alleles in our fragile X population compared to other European fragile X populations. The DXS548/FRAXAC1 haplotype 194/154 (7-3), which is common among normal populations, was found to be the most frequent haplotype in our fragile X population as well. The AGG interspersion analysis indicated that AGG loss rather than haplotype may determine FMR1 allele instability. Our results suggest that no common ancestral X chromosome is associated with fragile X syndrome in the Croatian population studied. Further analysis of the origin of fragile X syndrome among other Slavic populations will be necessary to better define its eastern European distribution.

Haplotype and AGG Interspersion Analysis of FMR1 Alleles in a Croatian Population: No Founder Effect Detected in Patients with Fragile X Syndrome

Several studies have suggested that fragile X syndrome (FRAXA), the most common inherited form of mental retardation, originated from a limited number of founder chromosomes. The aim of this study is to assess the genetic origin of fragile X syndrome in a Croatian population. We performed a haplotype analysis of the polymorphic loci DXS548 and FRAXAC1 in 18 unrelated fragile X and 56 control chromosomes. The AGG interspersion pattern of the FMR1 CGG repeat region was analyzed by sequencing. This is the first report on haplotype and AGG interspersion analysis of the fragile X syndrome gene in a Croatian population-the only eastern European population of Slavic origin analyzed so far. Our findings are intriguing, because they show a distinct distribution of the DXS548 and FRAXAC1 alleles in our fragile X population compared to other European fragile X populations. The DXS548/FRAXAC1 haplotype 194/154 (7-3), which is common among normal populations, was found to be the most frequent haplotype in our fragile X population as well. The AGG interspersion analysis indicated that AGG loss rather than haplotype may determine FMR1 allele instability. Our results suggest that no common ancestral X chromosome is associated with fragile X syndrome in the Croatian population studied. Further analysis of the origin of fragile X syndrome among other Slavic populations will be necessary to better define its eastern European distribution.

AGG interspersion analysis of the FMR1 CGG repeats in mental retardation of unspecific cause

Objectives To study the AGG interspersion pattern in mentally retarded patients of unspecified cause. Methods FMR1 CGG substructure in 104 normal and 232 mentally retarded (MR) males was determined by CGG repeat and AGG interspersion analyses. Genomic DNA of the study subjects was obtained for PCR and Southern hybridization analyses. Results All study subjects had less than 53 CGG repeats and none had fragile X syndrome of mental retardation. There was a significant difference ( P < 0.006) in the AGG interspersion pattern. MR males had (1) more variable internal substructures, (2) proportionally less 2 and 3 AGG but more 0 and 1 AGG, less (CGG) 9AGG(CGG) 9AGG(CGG) 9 but more (CGG) 9AGG(CGG) 19 alleles and (3) a longer pure 3′ CGG repeat. Conclusions Our results suggest that the MR alleles have a lesser number of interspersed AGG and a longer pure 3′ CGG repeat than the normal population. They are thus more prone to instability and expansion to long repeat lengths as in the fragile X syndrome of mental retardation.

AGG interspersion patterns in the CGG repeat of the FMR1 gene and linked DXS548/FRAXAC1 haplotypes in Brazilian populations

DOI 10.1002/ajmg.a.30376 In grouping the authors by geographic locality when publishing this letter, an error was introduced. The correct order of the contributors should have read: Cláudia B. Angeli,1 Leonardo P. Capelli,1 Maria Teresa B.M. Auricchio,1 Emygdya R. Leal-Mesquita,2 Ândrea K.C. Ribeiro-dos-Santos,3 Iris Ferrari,4 Silviene F. Oliveira,4 Maria de Nazaré Klautau-Guimarães,4 Angela M. Vianna-Morgante,1 and Regina C. Mingroni-Netto1 1Centro de Estudos do Genoma Humano, Departamento de Biologia, Instituto de Biociências, Universidade de São Paulo, São Paulo, Brazil 2Departamento de Biologia, Universidade Federal do Maranhão, Maranhão, Brazil 3Centro de Ciências Biológicas, Universidade Federal do Pará, Pará, Brazil 4Departamento de Genética e Morfologia, Instituto de Ciências Biológicas, Universidade de Brasília, Brasília, Brazil The Publisher regrets the error.

Structure and stability upon maternal transmission of common and intermediate FMR1 (Fragile X Mental Retardation 1) alleles in a sample of the Brazilian population

In order to investigate the stability of the FMR1 (Fragile X Mental Retardation 1) alleles from the normal population, when maternally inherited, we analyzed 75 mother-to-son transmissions. Sixty-eight alleles fell within the common range with 20-40 CGG repeats, and seven alleles were intermediate, with 41-48 repeats. No change was observed either in the length or in the structure of these repeats upon transmission. Fifty-three alleles were ascertained in different families, and their size distribution was similar to those described for European and European-derived populations, with three peaks of frequency: 66% of the alleles with (CGG)29, (CGG)30 or (CGG)31, 7.5% with (CGG)20, and 5.7% with (CGG)23. Regarding the AGG interspersion pattern, 69.8% had two AGG repeats, 20.8% had one, 5.7% had three and 3.8% had none. The most common patterns were 10+9+9 (30.2%), 9+9+9 (18.9%), 10+9 (7.5%), and 10+9+10 (7.5%). About 70% of the alleles with up to 40 repeats were linked to the DXS548/FRAXAC1 haplotype 7-3, the most commonly reported in normal populations. Four out of five intermediate alleles were in linkage with the two haplotypes most frequently associated to the FMR1 full mutation, 2-1 and 6-4. These four alleles showed long uninterrupted CGG repeats at the 3' end. The 9+9+22, 9+9+23 and 9+9+28 alleles were linked to the haplotype 2-1, and the 9+37 allele, to the haplotype 6-4. The pattern of AGG interspersion of these alleles and the associated haplotypes were in accordance with the two main pathways toward mutation previously proposed.

AGG interspersion patterns in the CGG repeat of the FMR1 gene and linked DXS548/FRAXAC1 haplotypes in Brazilian populations

AGG interspersion patterns in the CGG repeat of the <i>FMR1</i> gene and linked DXS548/FRAXAC1 haplotypes in Brazilian populations

A new insight into fragile X syndrome among Basque population.

The expansion of a trinucleotide repeat [CGG]n located in the FMR1 X-linked gene is the main cause of fragile X syndrome, the most common form of inherited mental retardation. We have analyzed the factors known, to date, to influence the instability of the repeat in 158 normal X chromosomes from the Spanish Basque population. These factors included length of the repeat, AGG interspersion pattern, length of uninterrupted CGG and DXS548-FRAXAC1 markers associated haplotype. Previous investigations on Basques showed an absence of this disorder among mentally retarded individuals that was likely due to a low prevalence of large CGG alleles and the presence of AGG interruptions on them. The present report suggests that, although the frequency of large alleles is low and they do maintain AGG interruptions, different mutational pathways that might lead to fragile X syndrome could be occurring among Basques. These pathways mainly include alleles with internal sequences 9 + 9 + n and 9 + 12 + 9 that show fragile X associated haplotypes. Besides, the lack of the most proximal AGG interruption, proposed recently as a novel factor involved in CGG repeat instability, was highly identified among alleles with long pure CGG tracts, which showed an internal sequence n + 9. The data suggest that, despite the lower incidence of large alleles, the prevalence of potentially unstable alleles among Basques is similar to that of other Caucasian populations and that these alleles could become fragile X chromosomes.

Analysis of FMR1 (CGG)(n) alleles and DXS548-FRAXAC1 haplotypes in three European circumpolar populations: traces of genetic relationship with Asia.

Fragile X syndrome, the most common form of inherited mental retardation, is caused by expansion of a (CGG)(n) repeat located in the FMR1 gene. The molecular factors involved in the mutation process from stable (CGG)(n) alleles towards unstable alleles are largely unknown, although family transmission studies and population studies have suggested that loss of AGG interruptions in the (CGG)(n) repeat is essential. We have analysed the AGG interspersion pattern of the FMR1 (CGG)(n) repeat and the haplotype distribution of closely located microsatellite markers DXS548 and FRAXAC1, in three circumarctic populations: Norwegians, Nenets and Saami. The data confirm the conservation, reported in all human populations studied so far, of an AGG interruption for each 9-10 CGG and support the stabilising effect of AGG interruptions. The data also indicate the existence of chromosomes of Asian origin in the Saami and Nenets population, thereby confirming a genetic relationship between Northern Europe and Asia. DXS548-FRAXAC1 haplotype frequencies were compared between 24 Norwegian fragile X males and 119 normal males. Significant linkage disequilibrium were found between the fragile X mutation and haplotype 6-4 and between normal (CGG)(n) alleles and haplotype 7-3.

Haplotype and AGG-interspersion analysis of FMR1 (CGG)(n) alleles in the Danish population: implications for multiple mutational pathways towards fragile X alleles.

The AGG interspersion pattern and flanking microsatellite markers and their association with instability of the FMR1 (CGG)(n) repeat, involved in the fragile X syndrome, were analyzed in DNA from filter-paper blood spots randomly collected from the Danish newborn population. Comparison of DXS548-FRAXAC1 haplotype frequencies in the normal population and among fragile X patients suggested strong linkage disequilibrium between normal alleles and haplotype 7-3 and between fragile X alleles and haplotype 2-1 and 6-4. Comparison of the AGG interspersion pattern in 143 alleles, ranging in size from 34-62 CGG, and their associated haplotypes indicates the existence of at least three mutational pathways from normal alleles toward fragile X alleles in the Danish population. Two subgroups of normal alleles, with internal sequences of (CGG)(10)AGG(CGG)(19) and (CGG)(9)AGG(CGG)(12) AGG(CGG)(9), possibly predisposed for expansion, were identified in the data set. When alleles larger than 34 CGG were investigated, comparing the length of 3' uninterrupted CGG triplets (uCGG), we found that alleles associated with haplotype 2-1 and 6-4 contain significantly longer stretches of uCGG than alleles associated with haplotype 7-3. Thus, the data support that (CGG)(n) instability is correlated to the length of uCGG.

Analysis of FMR1 (CGG)n alleles and FRAXA microsatellite haplotypes in the population of Greenland: implications for the population of the New World from Asia.

The fragile X syndrome is caused by the expansion of a polymorphic (CGG)n tract in the promoter region of the FMR1 gene. Apparently the incidence of fragile X syndrome is rare in the population of Greenland. In order to examine population-related factors involved in stability of the (CGG)n sequence, DNA samples obtained randomly from the Greenlandic population were analysed for size and AGG interspersion pattern of the FMR1 (CGG)n region and associated DXS548-FRAXAC1 haplotypes. In addition a large Greenland family with unstable transmission in the premutation range was analysed. The (CGG)n allele sizes in the Greenland population showed a narrow distribution similar to that reported for Asian populations. DNA sequencing of alleles with 36 CGG repeats revealed an AGG(CGG)6 insertion previously reported exclusively in Asian populations and a high frequency of alleles with a (CGG)10AGG(CGG)9AGG(CGG)9 or (CGG)9AGG(CGG)9AGG(CGG)6AGG(CGG)9 sequence pattern was found. Thus the data confirm the Asian origin of the Greenlandic (Eskimo) population and indicates that some (CGG)n alleles have remained stable for 15-30,000 years, since the population of the New World arrived from Asia via the Bering Strait.

Re-examination of factors associated with expansion of CGG repeats using a single nucleotide polymorphism in FMR1.

In at least 98% of fragile X syndrome cases, the disease results from expansion of the CGG repeat in the 5' end of FMR1. The use of microsatellite markers in the FMR1 region has revealed a disparity of risk between haplotypes for CGG repeat expansion. Although instability appears to depend on both the haplotype and the AGG interspersion pattern of the repeat, these factors alone do not completely describe the molecular basis for the linkage disequilibrium between normal and fragile X chromosomes, in part due to instability of the marker loci themselves. In an effort to better understand the mechanism of dynamic mutagenesis, we have searched for and discovered a single nucleotide polymorphism in intron 1 of FMR1 and characterized this marker, called ATL1, in 564 normal and 152 fragile X chromosomes. The G allele of this marker is found in 40% of normal chromosomes, in contrast to 83% of fragile X chromosomes. Not only is the G allele exclusively linked to haplotypes over-represented in fragile X syndrome, but G allele chromosomes also appear to transition to instability at a higher rate on haplotypes negatively associated with risk of expansion. The two alleles of ATL1 also reveal a highly significant linkage disequilibrium between unstable chromosomes and the 5' end of the CGG repeat itself, specifically the position of the first AGG interruption. The data expand the number of haplotypes associated with FMR1 and specifically allow discrimination, by ATL1 alleles, of single haplotypes with differing predispositions to expansion. Such haplotypes should prove useful in further defining the mechanism of dynamic mutagenesis.

Fragile X “gray zone” alleles: AGG patterns, expansion risks, and associated haplotypes

The risk for fragile X {open_quotes}gray-zone{close_quotes} alleles to expand appears to depend on the absence of stabilizing AGGs, which interrupt the CGG repeat region. To characterize such alleles better, we analyzed a series of 101 chromosomes with triplet repeat lengths ranging from 35 to 59 for variations in their AGG interspersion patterns. Among these, 11.9% had 3 AGGs, 59.3%, had 2, 24.8% had 1, and 4.0% had 0. An inverse relationship between FMR1 repeat length and the number of interrupting AGGs was observed. Within the range of 35-44 repeats, 98.7% of alleles were found to have a pure CGG repeat length (P{sub CGG}) of less than 33. However, among alleles with 45-59 repeats, 50% were found to have 0 or 1 AGG and a P{sub CGG} of more than 33. Thus, gray-zone alleles with 45-59 repeats frequently have a long stretch of pure CGGs and thus are more likely to be unstably inherited than alleles with 35-44 repeats. We found length associations of P{sub GCC} with 2 flanking microsatellites, DXS548 and FRAXAC 1: a P{sub CGG} 20 was more strongly associated with the haplotype 25-21. This resultmore » could reflect a founder effect or a generalized instability of CGGs and microsatellites. 28 refs., 2 figs., 4 tabs.« less

Fragile X "gray zone" alleles: AGG patterns, expansion risks, and associated haplotypes.

The risk for fragile X "gray-zone" alleles to expand appears to depend on the absence of stabilizing AGGs, which interrupt the CGG repeat region. To characterize such alleles better, we analyzed a series of 101 chromosomes with triplet repeat lengths ranging from 35 to 59 for variations in their AGG interspersion patterns. Among these, 11.9% had 3 AGGs, 59.3% had 2, 24.8% had 1, and 4.0% had 0. An inverse relationship between FMR1 repeat length and the number of interrupting AGGs was observed. Within the range of 35-44 repeats, 98.7% of alleles were found to have a pure CGG repeat length (PCGG) of less than 33. However, among alleles with 45-59 repeats, 50% were found to have 0 or 1 AGG and a PCGG of more than 33. Thus, gray-zone alleles with 45-59 repeats frequently have a long stretch of pure CGGs and thus are more likely to be unstably inherited than alleles with 35-44 repeats. We found length associations of PCGG with 2 flanking microsatellites, DXS548 and FRAXAC1: a PCGG < or = 20 was strongly associated with haplotype 20-19, whereas a PCGG > 20 was more strongly associated with the haplotype 25-21. This result could reflect a founder effect or a generalized instability of CGGs and microsatellites.

Fragile X founder effects and new mutations in Finland.

The apparent associations between fragile X mutations and nearby microsatellites may reflect both founder effects and microsatellite instability. To gain further insight into their relative contributions, we typed a sample of 56 unrelated control and 37 fragile X chromosomes from an eastern Finnish population for FMR1 CGG repeat lengths, AGG interspersion patterns, DXS548, FRAXAC1, FRAXE and a new polymorphic locus, Alu-L. In the controls, the most common FMR1 allele was 30 repeats with a range of 20 to 47 and a calculated heterozygosity of 88%. A strong founder effect was observed for locus DXS548 with 95% of fragile X chromosomes having the 21 CA repeat (196 bp) allele compared to 17% of controls, while none of the fragile X but 69% of controls had the 20 repeat allele. Although the FRAXAC1 locus is much closer than DXS548 to FMR1 (7 kb vs. 150 kb), there was no significant difference between fragile X and control FRAXAC1 allele distributions. The FRAXE repeat, located 600 kb distal to FMR1, was found to show strong linkage disequilibrium as well. A newly defined polymorphism, Alu-L, located at approximately 40 kb distal to the FMR1 repeat, showed very low polymorphism in the Finnish samples. Analysis of the combined loci DXS548-FRAXAC1-FRAXE showed three founder haplotypes. Haplotype 21-19-16 was found on 27 (75%) of fragile X chromosomes but on none of controls. Three (8.4%) fragile X chromosomes had haplotypes 21-19-15, 21-19-20, and 21-19-25 differing from the common fragile X haplotype only in FRAXE. These could have arisen by recombination or from mutations of FRAXE. A second haplotype 21-18-17 was found in four (11.1%) fragile X chromosomes but only one (1.9%) control. This may represent a more recent founder mutation. A third haplotype 25-21-15, seen in two fragile X chromosomes (5.6%) and one (1.9%) control, was even less common and thus may represent an even more recent mutation or admixture of immigrant types. Analysis of the AGG interspersions within the FMR1 CGG repeat showed that 7/8 premutation chromosomes lacked an AGG whereas all controls had at least one AGG. This supports the hypothesis that the mutation of AGG to CGG leads to repeat instability and mutational expansion.