RESUMO
BACKGROUND: Spinocerebellar ataxia type 12 (SCA12) is a neurodegenerative disease caused by a CAG/CTG repeat expansion at the PPP2R2B locus. OBJECTIVE: We investigated how the CAG repeat expansion within the PPP2R2B 7B7D transcript influences the expression of Bß1 and a potential protein containing a long polyserine tract. METHODS: Transcript and protein expression were measured using quantitative PCR (qPCR) Role of Bß1 overexpression in the pathogenesis of SCA12 and Western blot, respectively, in an SK-N-MC cell model that overexpresses the full-length PPP2R2B 7B7D transcript. The apoptotic effect of a protein containing a long polyserine tract on SK-N-MC cells was evaluated using caspase 3/7 activity. RESULTS: The CAG repeat expansion increases the expression of the PPP2R2B 7B7D transcript, as well as Bß1 protein, in an SK-N-MC cell model in which the full-length PPP2R2B 7B7D transcript is overexpressed. The CAG repeat expansion within the 7B7D transcript is translated into a long polyserine tract that triggers apoptosis in SK-N-MC cells. CONCLUSIONS: The SCA12 mutation leads to overexpression of PPP2R2B Bß1 and to expression of a protein containing a long polyserine tract; both these effects potentially contribute to SCA12 pathogenesis. © 2024 International Parkinson and Movement Disorder Society.
RESUMO
BACKGROUND: Genes, principal units of genetic information, vary in complexity and evolutionary history. Less-complex genes (e.g., long non-coding RNA (lncRNA) expressing genes) readily emerge de novo from non-genic sequences and have high evolutionary turnover. Genesis of a gene may be facilitated by adoption of functional genic sequences from retrotransposon insertions. However, protein-coding sequences in extant genomes rarely lack any connection to an ancestral protein-coding sequence. RESULTS: We describe remarkable evolution of the murine gene D6Ertd527e and its orthologs in the rodent Muroidea superfamily. The D6Ertd527e emerged in a common ancestor of mice and hamsters most likely as a lncRNA-expressing gene. A major contributing factor was a long terminal repeat (LTR) retrotransposon insertion carrying an oocyte-specific promoter and a 5' terminal exon of the gene. The gene survived as an oocyte-specific lncRNA in several extant rodents while in some others the gene or its expression were lost. In the ancestral lineage of Mus musculus, the gene acquired protein-coding capacity where the bulk of the coding sequence formed through CAG (AGC) trinucleotide repeat expansion and duplications. These events generated a cytoplasmic serine-rich maternal protein. Knock-out of D6Ertd527e in mice has a small but detectable effect on fertility and the maternal transcriptome. CONCLUSIONS: While this evolving gene is not showing a clear function in laboratory mice, its documented evolutionary history in Muroidea during the last ~ 40 million years provides a textbook example of how a several common mutation events can support de novo gene formation, evolution of protein-coding capacity, as well as gene's demise.
Assuntos
Muridae , RNA Longo não Codificante , Animais , RNA Longo não Codificante/genéticaRESUMO
Spinocerebellar ataxia type 8 (SCA8) is caused by a bidirectionally transcribed CTG·CAG expansion that results in the in vivo accumulation of CUG RNA foci, an ATG-initiated polyGln and a polyAla protein expressed by repeat-associated non-ATG (RAN) translation. Although RAN proteins have been reported in a growing number of diseases, the mechanisms and role of RAN translation in disease are poorly understood. We report a novel toxic SCA8 polySer protein which accumulates in white matter (WM) regions as aggregates that increase with age and disease severity. WM regions with polySer aggregates show demyelination and axonal degeneration in SCA8 human and mouse brains. Additionally, knockdown of the eukaryotic translation initiation factor eIF3F in cells reduces steady-state levels of SCA8 polySer and other RAN proteins. Taken together, these data show polySer and WM abnormalities contribute to SCA8 and identify eIF3F as a novel modulator of RAN protein accumulation.
Assuntos
Envelhecimento/metabolismo , Fator de Iniciação 3 em Eucariotos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Degenerações Espinocerebelares/metabolismo , Substância Branca/metabolismo , Envelhecimento/genética , Envelhecimento/patologia , Animais , Fator de Iniciação 3 em Eucariotos/genética , Células HeLa , Humanos , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Degenerações Espinocerebelares/genética , Degenerações Espinocerebelares/patologia , Substância Branca/patologiaRESUMO
Short polyserine (polyS) repeats are frequently found in proteins and longer ones are produced in neurological disorders such as Huntington disease (HD) owing to translational frameshifting or non-ATG-dependent translation, together with polyglutamine (polyQ) and polyalanine (polyA) repeats, forming intracellular aggregates. However, the physiological and pathological structures of polyS repeats are not clearly understood. Early studies highlighted their structural versatility, similar to other homopolymers whose conformation is influenced by the surrounding protein context. As polyS stretches are frequently near polyQ and polyA repeats, which can be part of coiled coil (CC) structures, and the frameshift-derived polyS repeats in HD directly flank CC heptads important for aggregation, we investigate here the structural and aggregation properties of polyS in the context of CC structures. We have taken advantage of peptide models, previously used to study polyQ and polyA in CCs, in which we inserted polyS repeats of variable length and studied them in comparison with polyQ and polyA peptides. We found that polyS repeats promote CC-mediated polymerization and fibrillization as revealed by circular dichroism, chemical crosslinking, and atomic force microscopy. Furthermore, they promote CC-based, length-dependent intracellular aggregation, which is negligible with 7 and widespread with 49 serines. These findings show that polyS repeats can participate in the formation of CCs, as previously found for polyQ and polyA, conferring to peptides distinctive structural properties with aggregation kinetics that are intermediate between those of polyA and polyQ CCs, and contribute to an overall structural definition of the pathophysiogical roles of homopolymeric repeats in CC structures.
Assuntos
Peptídeos/química , Agregados Proteicos , Conformação Proteica , Dicroísmo Circular , Humanos , Cinética , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/metabolismo , Peptídeos/genética , Agregação Patológica de ProteínasRESUMO
Kasugamycin residues (KASU), a pest control antibiotic, was reported as an ecosystem threat owing to its over-application in plant protection to meet the growing global need for agronomic products. Therefore, we report herein the first electrochemical sensor for fast and sensitive analysis of KASU in vegetables based on the synergetic hybridization between conducting polyserine film (poly (SER)), and carbon nanomaterials including functionalized multiwalled carbon nanotubes (fMWCNTs) and reduced graphene oxide (rGO). The sensor was characterized morphologically using Scanning electron (SEM) and atomic force Microscopy (AFM), while cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used for electrochemical characterization. Under the optimized conditions using differential pulse voltammetry (DPV), the sensor exhibited an outstanding sensitivity and selectivity, with a good linear response of 3-106 µg/mL and an assessed limit of detection and quantification of 0.40 and 1.33 µg/mL, respectively. Furthermore, the electrochemical sensor was effectively applied to quantify KASU in cucumber, zucchini, and carrots with a recovery range 95.5-100.1%, and RSD lower than 4.1% (n = 3), showing its applicability and efficiency for selective analysis of KASU in foodstuffs.
Assuntos
Grafite , Nanotubos de Carbono , Limite de Detecção , Verduras , Técnicas Eletroquímicas/métodos , Nanotubos de Carbono/química , Ecossistema , Aminoglicosídeos , Antibacterianos , Eletrodos , Grafite/químicaRESUMO
Carbon nanotubes (CNTs) display exceptional properties that predispose them to wide use in technological or biomedical applications. To remove the toxicity of CNTs and to protect them against undesired protein adsorption, coverage of the CNT sidewall with poly(ethylene oxide) (PEO) is often considered. However, controversial results on the antifouling effectiveness of PEO layers have been reported so far. In this work, the interactions of pristine CNT and CNT covered with the PEO chains at different grafting densities with polyglycine, polyserine, and polyvaline are studied using molecular dynamics simulations in vacuum, water, and saline environments. The peptides are adsorbed on CNT in all investigated systems; however, the adsorption strength is reduced in aqueous environments. Save for one case, addition of NaCl at a physiological concentration to water does not appreciably influence the adsorption and structure of the peptides or the grafted PEO layer. It turns out that the flexibility of the peptide backbone allows the peptide to adopt more asymmetric conformations which may be inserted deeper into the grafted PEO layer. Water molecules disrupt the internal hydrogen bonds in the peptides, as well as the hydrogen bonds formed between the peptides and the PEO chains.