Detection and Quantification of the Fragile X Mental Retardation Protein 1 (FMRP)

LaFauci, Giuseppe; Adayev, Tatyana; Kascsak, Richard J.; Brown, W. Ted

doi:10.3390/genes7120121

Cited by 19 publications

(17 citation statements)

References 67 publications

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“…Thus, we investigated if the S500D mutation causes major structural changes within FMRP by circular dichroism spectroscopy and by fluorescence spectroscopy. We used FMRP and FMRP S500D at 1 μM and 5 μM concentrations, as it has been proposed that at high μM concentrations FMRP tends to aggregate [52]. As seen in Fig 6A there are no major structural differences between FMRP and FMRP S500D, as both their CD spectra overlap almost perfectly at both protein concentrations investigated.…”

Section: Resultsmentioning

confidence: 99%

FMRP - G-quadruplex mRNA - miR-125a interactions: Implications for miR-125a mediated translation regulation of PSD-95 mRNA

et al. 2019

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Fragile X syndrome, the most common inherited form of intellectual disability, is caused by the CGG trinucleotide expansion in the 5’-untranslated region of the Fmr1 gene on the X chromosome, which silences the expression of the fragile X mental retardation protein (FMRP). FMRP has been shown to bind to a G-rich region within the PSD-95 mRNA, which encodes for the postsynaptic density protein 95, and together with microRNA-125a to mediate the reversible inhibition of the PSD-95 mRNA translation in neurons. The miR-125a binding site within the PSD-95 mRNA 3’-untranslated region (UTR) is embedded in a G-rich region bound by FMRP, which we have previously demonstrated folds into two parallel G-quadruplex structures. The FMRP regulation of PSD-95 mRNA translation is complex, being mediated by its phosphorylation. While the requirement for FMRP in the regulation of PSD-95 mRNA translation is clearly established, the exact mechanism by which this is achieved is not known. In this study, we have shown that both unphosphorylated FMRP and its phosphomimic FMRP S500D bind to the PSD-95 mRNA G-quadruplexes with high affinity, whereas only FMRP S500D binds to miR-125a. These results point towards a mechanism by which, depending on its phosphorylation status, FMRP acts as a switch that potentially controls the stability of the complex formed by the miR-125a-guided RNA induced silencing complex (RISC) and PSD-95 mRNA.

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Section: Resultsmentioning

confidence: 99%

FMRP - G-quadruplex mRNA - miR-125a interactions: Implications for miR-125a mediated translation regulation of PSD-95 mRNA

et al. 2019

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“…These findings are contributory since concordance in patterns of FMR1 mosaicism across tissues continues to be a controversial issue in FXS [ 46 , 59 , 60 ]. Moreover, very little is known about the correlation between blood FMRP measurements and postmortem brain tissue FMRP levels in the same subject [ 61 , 62 ]. Only a single case study “verified” reduced expression of FMR1 mRNA and FMRP in both peripheral blood and brain leading to the FXS [ 61 ].…”

Section: Discussionmentioning

confidence: 99%

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

et al. 2020

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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.

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“…Further research is needed to clarify the potential role for FMRP in sleep–wake regulation. Moreover, the concentration of FMRP in human blood serum is low (in the pg range), rendering its quantification difficult, and depends on various possible factors, including genetic influences [66]. Cautious interpretation and independent replication of this result are, thus, crucial.…”

Section: Discussionmentioning

confidence: 99%

Dynamic changes in cerebral and peripheral markers of glutamatergic signaling across the human sleep–wake cycle

Weigend

Holst

Treyer

et al. 2019

Sleep

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Sleep and brain glutamatergic signaling are homeostatically regulated. Recovery sleep following prolonged wakefulness restores efficient functioning of the brain, possibly by keeping glutamatergic signaling in a homeostatic range. Evidence in humans and mice suggested that metabotropic glutamate receptors of subtype-5 (mGluR5) contribute to the brain’s coping mechanisms with sleep deprivation. Here, proton magnetic resonance spectroscopy in 31 healthy men was used to quantify the levels of glutamate (Glu), glutamate-to-glutamine ratio (GLX), and γ-amino-butyric-acid (GABA) in basal ganglia (BG) and dorsolateral prefrontal cortex on 3 consecutive days, after ~8 (baseline), ~32 (sleep deprivation), and ~8 hours (recovery sleep) of wakefulness. Simultaneously, mGluR5 availability was quantified with the novel radioligand for positron emission tomography, [18F]PSS232, and the blood levels of the mGluR5-regulated proteins, fragile X mental retardation protein (FMRP) and brain-derived neurotrophic factor (BDNF) were determined. The data revealed that GLX (p = 0.03) in BG (for Glu: p < 0.06) and the serum concentration of FMRP (p < 0.04) were increased after sleep loss. Other brain metabolites (GABA, N-acetyl-aspartate, choline, glutathione) and serum BDNF levels were not altered by sleep deprivation (pall > 0.6). By contrast, the night without sleep enhanced whole-brain, BG, and parietal cortex mGluR5 availability, which was normalized by recovery sleep (pall < 0.05). The findings provide convergent multimodal evidence that glutamatergic signaling is affected by sleep deprivation and recovery sleep. They support a role for mGluR5 and FMRP in sleep–wake regulation and warrant further studies to investigate their causality and relevance for regulating human sleep in health and disease.Clinical Trial Registration: www.clinicaltrials.gov (study identifier: NCT03813082)

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Detection and Quantification of the Fragile X Mental Retardation Protein 1 (FMRP)

Cited by 19 publications

References 67 publications

FMRP - G-quadruplex mRNA - miR-125a interactions: Implications for miR-125a mediated translation regulation of PSD-95 mRNA

FMRP - G-quadruplex mRNA - miR-125a interactions: Implications for miR-125a mediated translation regulation of PSD-95 mRNA

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

Dynamic changes in cerebral and peripheral markers of glutamatergic signaling across the human sleep–wake cycle

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