scholarly journals A Genotype-Phenotype Study of High-Resolution FMR1 Nucleic Acid and Protein Analyses in Fragile X Patients with Neurobehavioral Assessments

2020 ◽  
Vol 10 (10) ◽  
pp. 694 ◽  
Author(s):  
Dejan B. Budimirovic ◽  
Annette Schlageter ◽  
Stela Filipovic-Sadic ◽  
Dragana D. Protic ◽  
Eran Bram ◽  
...  
Keyword(s):  
Cell Lines ◽  
Genomic Dna ◽  
Fragile X ◽  
Genetic Disorder ◽  
Critical Role ◽  
Autism Spectrum ◽  
Total N ◽  
Cgg Repeat ◽  
Repeat Expansions ◽  
High Level

Fragile X syndrome (FXS) is caused by silencing of the FMR1 gene, which encodes a protein with a critical role in synaptic plasticity. The molecular abnormality underlying FMR1 silencing, CGG repeat expansion, is well characterized; however, delineation of the pathway from DNA to RNA to protein using biosamples from well characterized patients with FXS is limited. Since FXS is a common and prototypical genetic disorder associated with intellectual disability (ID) and autism spectrum disorder (ASD), a comprehensive assessment of the FMR1 DNA-RNA-protein pathway and its correlations with the neurobehavioral phenotype is a priority. We applied nine sensitive and quantitative assays evaluating FMR1 DNA, RNA, and FMRP parameters to a reference set of cell lines representing the range of FMR1 expansions. We then used the most informative of these assays on blood and buccal specimens from cohorts of patients with different FMR1 expansions, with emphasis on those with FXS (N = 42 total, N = 31 with FMRP measurements). The group with FMRP data was also evaluated comprehensively in terms of its neurobehavioral profile, which allowed molecular–neurobehavioral correlations. FMR1 CGG repeat expansions, methylation levels, and FMRP levels, in both cell lines and blood samples, were consistent with findings of previous FMR1 genomic and protein studies. They also demonstrated a high level of agreement between blood and buccal specimens. These assays further corroborated previous reports of the relatively high prevalence of methylation mosaicism (slightly over 50% of the samples). Molecular-neurobehavioral correlations confirmed the inverse relationship between overall severity of the FXS phenotype and decrease in FMRP levels (N = 26 males, mean 4.2 ± 3.3 pg FMRP/ng genomic DNA). Other intriguing findings included a significant relationship between the diagnosis of FXS with ASD and two-fold lower levels of FMRP (mean 2.8 ± 1.3 pg FMRP/ng genomic DNA, p = 0.04), in particular observed in younger age- and IQ-adjusted males (mean age 6.9 ± 0.9 years with mean 3.2 ± 1.2 pg FMRP/ng genomic DNA, 57% with severe ASD), compared to FXS without ASD. Those with severe ID had even lower FMRP levels independent of ASD status in the male-only subset. The results underscore the link between FMR1 expansion, gene methylation, and FMRP deficit. The association between FMRP deficiency and overall severity of the neurobehavioral phenotype invites follow up studies in larger patient cohorts. They would be valuable to confirm and potentially extend our initial findings of the relationship between ASD and other neurobehavioral features and the magnitude of FMRP deficit. Molecular profiling of individuals with FXS may have important implications in research and clinical practice.

scholarly journals A Genotype-Phenotype Study of High-Resolution FMR1 Nucleic Acid and Protein Analyses in Fragile X Patients With Neurobehavioral Assessments

2020 ◽  
Author(s):  
Dejan B Budimirovic ◽  
Annette Schlageter ◽  
Stela Sadic-Filipovic ◽  
Dragana Protic ◽  
Eran Bram ◽  
...  
Keyword(s):  
Cell Lines ◽  
Fragile X ◽  
Genetic Disorder ◽  
Critical Role ◽  
Autism Spectrum ◽  
Total N ◽  
Cgg Repeat ◽  
High Prevalence ◽  
Repeat Expansions ◽  
High Level

Abstract Background. Fragile X syndrome (FXS) is caused by silencing of the FMR1 gene, which encodes a protein with a critical role in synaptic plasticity. The molecular abnormality underlying FMR1 silencing, CGG repeat expansion, is well characterized; however, delineation of the pathway from DNA to RNA to protein using biosamples from well characterized patients with FXS is limited. Since FXS is a common and prototypical genetic disorder associated with intellectual disability (ID) and autism spectrum disorder (ASD), a comprehensive assessment of the FMR1 DNA-RNA-protein pathway and its correlations with the neurobehavioral phenotype is a priority.Methods. We applied nine sensitive and quantitative assays evaluating FMR1 DNA, RNA, and FMRP parameters to a reference set of cell lines representing the range of FMR1 expansions. We then used the most informative of these assays on blood and buccal specimens from cohorts of patients with different FMR1 expansions, with emphasis on those with FXS (N = 42 total, N = 31 with FMRP measurements). The group with FMRP data was also evaluated comprehensively in terms of its neurobehavioral profile, which allowed molecular-neurobehavioral correlations. Results. FMR1 CGG repeat expansions, methylation levels, and FMRP levels, in both cell lines and blood samples, were consistent with previous FMR1 genomic and protein studies. They also demonstrated a high level of agreement between blood and buccal specimens. These assays further corroborated previous reports of the relatively high prevalence of methylation mosaicism. Molecular-neurobehavioral correlations confirmed the inverse relationship between overall severity of the FXS phenotype and decrease in FMRP levels. Other intriguing findings included a potential relationship between diagnosis of FXS with ASD and very low levels of FMRP, compared to FXS without ASD. Conclusions. The results underscore the link between FMR1 expansion, gene methylation, and FMRP deficit. The association between FMRP deficiency and overall severity of the neurobehavioral phenotype invites follow up studies in larger patient cohorts. They would be valuable to confirm and potentially extend our initial findings of a relationship between ASD and other neurobehavioral features and the magnitude of FMRP deficit. Molecular profiling of individuals with FXS may have important implications in research and clinical practice.

scholarly journals Cascade Testing for Fragile X Syndrome in a Rural Setting in Cameroon (Sub-Saharan Africa)

Genes ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 136
Author(s):  
Karen Kengne Kamga ◽  
Séraphin Nguefack ◽  
Khuthala Minka ◽  
Edmond Wonkam Tingang ◽  
Alina Esterhuizen ◽  
...  
Keyword(s):  
Fragile X Syndrome ◽  
Fragile X ◽  
Autism Spectrum ◽  
Sub Saharan Africa ◽  
Rural Setting ◽  
Premature Menopause ◽  
Molecular Testing ◽  
Full Mutation ◽  
Cgg Repeat ◽  
Cgg Repeats

Fragile X Syndrome (FXS), an X-linked dominant monogenic condition, is the main genetic cause of intellectual disability (ID) and autism spectrum disorder (ASD). FXS is associated with an expansion of CGG repeat sequence in the Fragile X Mental Retardation gene 1 (FMR1) on chromosome X. Following a neuropediatric assessment of two male siblings who presented with signs of FXS that was confirmed with molecular testing, we provided cascade counselling and testing to the extended family. A total of 46 individuals were tested for FXS; among them, 58.70% (n = 27) were females. The mean age was 9.4 (±5) years for children and 45.9 (±15.9) years for adults. Pedigree analysis suggested that the founder of these families was likely a normal transmitting male. Four out of 19 males with clinical ID were confirmed to have a full mutation for FXS, while 14/27 females had a pathologic CGG expansion (>56 CGG repeats) on one of their X chromosomes. Two women with premature menopause were confirmed of being carriers of premutation (91 and 101 CGG repeats). We also identified maternal alleles (91 and 126 CGG repeats) which expanded to a full mutation in their offspring (>200 CGG repeats). This study is a rare report on FXS from Africa and illustrates the case scenario of implementing genetic medicine for a neurogenetic condition in a rural setting.

scholarly journals Enhanced detection of nucleotide repeat mRNA with hybridization chain reaction

2021 ◽  
Author(s):  
Mary Rebecca Glineburg ◽  
Yuan Zhang ◽  
Elizabeth M Tank ◽  
Sami Barmada ◽  
Peter Todd
Keyword(s):  
Fragile X ◽  
Data Interpretation ◽  
Repeat Expansion ◽  
Hybridization Chain Reaction ◽  
Chain Reaction ◽  
Cgg Repeat ◽  
Hexanucleotide Repeat ◽  
Rna Foci ◽  
Repeat Expansions ◽  
The Impact

RNAs derived from expanded nucleotide repeats form detectable foci in patient cells and these foci are thought to contribute to disease pathogenesis. The most widely used method for detecting RNA foci is fluorescence in situ hybridization (FISH). However, FISH is prone to low sensitivity and photo-bleaching that can complicate data interpretation. Here we applied hybridization chain reaction (HCR) as an alternative approach to repeat RNA foci detection of GC-rich repeats in two neurodegenerative disorders: GGGGCC (G4C2) hexanucleotide repeat expansions in C9orf72 that cause amyotrophic lateral sclerosis and frontotemporal dementia (C9 ALS/FTD) and CGG repeat expansions in FMR1 that cause Fragile X-associated tremor/ataxia syndrome. We found that HCR of both G4C2 and CGG repeats has comparable specificity to traditional FISH, but is >40x more sensitive and shows repeat-length dependence in its intensity. HCR is better than FISH at detecting both nuclear and cytoplasmic foci in human C9 ALS/FTD fibroblasts, patient iPSC derived neurons, and patient brain samples. We used HCR to determine the impact of integrated stress response (ISR) activation on RNA foci number and distribution. G4C2 repeat RNA did not readily co-localize with the stress granule marker G3BP1, but ISR induction increased both the number of detectible nuclear RNA foci and the nuclear/cytoplasmic foci ratio in patient fibroblasts and patient derived neurons. Taken together, these data suggest that HCR can be a useful tool for detecting repeat expansion mRNA in C9 ALS/FTD and other repeat expansion disorders.

scholarly journals Screening for FMR1 CGG Repeat Expansion in Thai Patients with Autism Spectrum Disorder

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Areerat Hnoonual ◽  
Charunee Jankittunpaiboon ◽  
Pornprot Limprasert
Keyword(s):  
Autism Spectrum Disorder ◽  
Fragile X ◽  
Single Gene ◽  
Autism Spectrum ◽  
Spectrum Disorder ◽  
Fmr1 Gene ◽  
Normal Male ◽  
Cgg Repeat ◽  
Cgg Repeats ◽  
Normal Controls

Autism spectrum disorder (ASD) is a complex disorder with a heterogeneous etiology. Fragile X syndrome (FXS) is recognized as the most common single gene mutation associated with ASD. FXS patients show some autistic behaviors and may be difficult to distinguish at a young age from autistic children. However, there have been no published reports on the prevalence of FXS in ASD patients in Thailand. In this study, we present a pilot study to analyze the CGG repeat sizes of the FMR1 gene in Thai autistic patients. We screened 202 unrelated Thai patients (168 males and 34 females) with nonsyndromic ASD and 212 normal controls using standard FXS molecular diagnosis techniques. The distributions of FMR1 CGG repeat sizes in the ASD and normal control groups were similar, with the two most common alleles having 29 and 30 CGG repeats, followed by an allele with 36 CGG repeats. No FMR1 full mutations or premutations were found in either ASD individuals or the normal controls. Interestingly, three ASD male patients with high normal CGG and intermediate CGG repeats (44, 46, and 53 CGG repeats) were identified, indicating that the prevalence of FMR1 intermediate alleles in Thai ASD patients was approximately 1% while these alleles were absent in the normal male controls. Our study indicates that CGG repeat expansions of the FMR1 gene may not be a common genetic cause of nonsyndromic ASD in Thai patients. However, further studies for mutations other than the CGG expansion in the FMR1 gene are required to get a better information on FXS prevalence in Thai ASD patients.

203 PLASMA MEMBRANE Ca2+-PUMPING ATPase 1 IS ABUNDANTLY EXPRESSED AND DISTINCTLY REGULATED BY ESTROGEN IN HUMAN ENDOMETRIUM DURING THE MENSTRUAL CYCLE

2011 ◽  
Vol 23 (1) ◽  
pp. 201
Author(s):  
H. Yang ◽  
E.-B. Jeung
Keyword(s):  
Plasma Membrane ◽  
Menstrual Cycle ◽  
Critical Role ◽  
Human Endometrium ◽  
Cycle Phase ◽  
Menstrual Phase ◽  
Menstrual Cycle Phase ◽  
Secretory Phase ◽  
Total N ◽  
High Level

Plasma membrane Ca2+-pumping ATPases (PMCA) play a critical role in maintaining cellular Ca2+ homeostasis. The PMCA mRNA are encoded on 4 genes, designated PMCA1 to PMCA4. In a previous study, we found that both PMCA1 and PMCA4 are expressed at similar levels in astrocytes and in neurons. Although PMCA1b is expressed in the uterus of rats during the oestrous cycle, the expression of PMCA1 and its potential roles has not been elucidated during the menstrual cycle in the human endometrium. Thus, in the current study, the expression pattern of PMCA1 was examined to predict its roles in the human endometrium during the menstrual cycle. Human uterine tissues (total n = 40) were separated into 3 groups according to menstrual cycle phase: menstrual phase, proliferative phase (early, mid, late), and secretory phase (early, mid, late). Using real-time PCR and Western blot analysis, uterine expression of PMCA1 mRNA and protein increased to 1.5-fold in the early-, mid- and late-proliferative phases in the endometrium of the human uterus, compared with other menstrual phases. In addition, uterine PMCA1 was abundantly localised in the cytoplasm of the luminal and glandular epithelial cells in the menstrual phases, indicating that this protein may participate in the uterine Ca balance of the human endometrium during the menstrual cycle. Taken together, these results suggest that a high level of uterine PMCA1 expression may be involved in reproductive functions during the menstrual cycle of humans.

scholarly journals Cascade Screening for Fragile X Syndrome/CGG Repeat Expansions in Children Attending Special Education in Sri Lanka

PLoS ONE ◽  
2015 ◽  
Vol 10 (12) ◽  
pp. e0145537 ◽  
Author(s):  
C. H. W. M. R. Bhagya Chandrasekara ◽  
W. S. Sulochana Wijesundera ◽  
Hemamali N. Perera ◽  
Samuel S. Chong ◽  
Indhu-Shree Rajan-Babu
Keyword(s):  
Special Education ◽  
Sri Lanka ◽  
Fragile X Syndrome ◽  
Fragile X ◽  
Cascade Screening ◽  
Cgg Repeat ◽  
Repeat Expansions

scholarly journals CGG repeat RNA G-quadruplexes interact with FMRpolyG to cause neuronal dysfunction in fragile X-related tremor/ataxia syndrome

2021 ◽  
Vol 7 (3) ◽  
pp. eabd9440
Author(s):  
Sefan Asamitsu ◽  
Yasushi Yabuki ◽  
Susumu Ikenoshita ◽  
Kosuke Kawakubo ◽  
Moe Kawasaki ◽  
...  
Keyword(s):  
Therapeutic Strategy ◽  
Aminolevulinic Acid ◽  
Fragile X ◽  
Protoporphyrin Ix ◽  
Neuronal Dysfunction ◽  
Cgg Repeat ◽  
Repeat Expansions ◽  
Ataxia Syndrome ◽  
And Behavior ◽  
Ran Translation

Fragile X-related tremor/ataxia syndrome (FXTAS) is a neurodegenerative disease caused by CGG triplet repeat expansions in FMR1, which elicit repeat-associated non-AUG (RAN) translation and produce the toxic protein FMRpolyG. We show that FMRpolyG interacts with pathogenic CGG repeat-derived RNA G-quadruplexes (CGG-G4RNA), propagates cell to cell, and induces neuronal dysfunction. The FMRpolyG polyglycine domain has a prion-like property, preferentially binding to CGG-G4RNA. Treatment with 5-aminolevulinic acid, which is metabolized to protoporphyrin IX, inhibited RAN translation of FMRpolyG and CGG-G4RNA–induced FMRpolyG aggregation, ameliorating aberrant synaptic plasticity and behavior in FXTAS model mice. Thus, we present a novel therapeutic strategy to target G4RNA prionoids.

scholarly journals Inhibitors of Histone Deacetylases Are Weak Activators of the FMR1 Gene in Fragile X Syndrome Cell Lines

2017 ◽  
Vol 2017 ◽  
pp. 1-5 ◽  
Author(s):  
Alexander A. Dolskiy ◽  
Vladimir O. Pustylnyak ◽  
Andrey A. Yarushkin ◽  
Natalya A. Lemskaya ◽  
Dmitry V. Yudkin
Keyword(s):  
Fragile X Syndrome ◽  
Cell Lines ◽  
Histone Deacetylases ◽  
Neuronal Development ◽  
Fragile X ◽  
Repeat Expansion ◽  
Symptomatic Therapy ◽  
Cgg Repeat ◽  
Clinic Practice ◽  
Patient Cell

Fragile X syndrome is the most common cause of inherited intellectual disability in humans. It is a result of CGG repeat expansion in the 5′ untranslated region (5′ UTR) of the FMR1 gene. This gene encodes the FMRP protein that is involved in neuronal development. Repeat expansion leads to heterochromatinization of the promoter, gene silencing, and the subsequent absence of FMRP. To date, there is no specific therapy for the syndrome. All treatments in clinic practice provide symptomatic therapy. The development of drug therapy for Fragile X syndrome treatment is connected with the search for inhibitors of enzymes that are responsible for heterochromatinization. Here, we report a weak transcriptional activity of the FMR1 gene and the absence of FMRP protein after Fragile X syndrome cell lines treatment with two FDA approved inhibitors of histone deacetylases, romidepsin and vorinostat. We demonstrate that romidepsin, an inhibitor of class I histone deacetylases, does not activate FMR1 expression in patient cell cultures, whereas vorinostat, an inhibitor of classes I and II histone deacetylases, activates a low level of FMR1 expression in some patient cell lines.

scholarly journals FMR1 and Autism, an Intriguing Connection Revisited

Genes ◽  
2021 ◽  
Vol 12 (8) ◽  
pp. 1218
Author(s):  
William Fyke ◽  
Milen Velinov
Keyword(s):  
Fragile X ◽  
Major Gene ◽  
Single Gene ◽  
Autism Spectrum ◽  
Protein Product ◽  
Loss Of Function ◽  
Cgg Repeat ◽  
Statistical Manual ◽  
The Us ◽  
The Central Nervous System

Autism Spectrum Disorder (ASD) represents a distinct phenotype of behavioral dysfunction that includes deficiencies in communication and stereotypic behaviors. ASD affects about 2% of the US population. It is a highly heritable spectrum of conditions with substantial genetic heterogeneity. To date, mutations in over 100 genes have been reported in association with ASD phenotypes. Fragile X syndrome (FXS) is the most common single-gene disorder associated with ASD. The gene associated with FXS, FMR1 is located on chromosome X. Accordingly, the condition has more severe manifestations in males. FXS results from the loss of function of FMR1 due to the expansion of an unstable CGG repeat located in the 5′′ untranslated region of the gene. About 50% of the FXS males and 20% of the FXS females meet the Diagnostic Statistical Manual 5 (DSM-5) criteria for ASD. Among the individuals with ASD, about 3% test positive for FXS. FMRP, the protein product of FMR1, is a major gene regulator in the central nervous system. Multiple pathways regulated by FMRP are found to be dysfunctional in ASD patients who do not have FXS. Thus, FXS presents the opportunity to study cellular phenomena that may have wider applications in the management of ASD and to develop new strategies for ASD therapy.

scholarly journals Abundancy of polymorphic CGG repeats in the human genome suggest a broad involvement in neurological disease

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Dale J. Annear ◽  
Geert Vandeweyer ◽  
Ellen Elinck ◽  
Alba Sanchis-Juan ◽  
Courtney E. French ◽  
...  
Keyword(s):  
Human Disease ◽  
Fragile X ◽  
Disease Genes ◽  
Cgg Repeat ◽  
Neurodevelopmental Disease ◽  
Repeat Units ◽  
Cgg Repeats ◽  
Repeat Expansions ◽  
Ataxia Syndrome ◽  
Strong Candidate

AbstractExpanded CGG-repeats have been linked to neurodevelopmental and neurodegenerative disorders, including the fragile X syndrome and fragile X-associated tremor/ataxia syndrome (FXTAS). We hypothesized that as of yet uncharacterised CGG-repeat expansions within the genome contribute to human disease. To catalogue the CGG-repeats, 544 human whole genomes were analyzed. In total, 6101 unique CGG-repeats were detected of which more than 93% were highly variable in repeat length. Repeats with a median size of 12 repeat units or more were always polymorphic but shorter repeats were often polymorphic, suggesting a potential intergenerational instability of the CGG region even for repeats units with a median length of four or less. 410 of the CGG repeats were associated with known neurodevelopmental disease genes or with strong candidate genes. Based on their frequency and genomic location, CGG repeats may thus be a currently overlooked cause of human disease.

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