TY - JOUR T1 - Cortical neurons derived from human pluripotent stem cells lacking FMRP display altered spontaneous firing patterns. JF - Mol Autism Y1 - 2020 A1 - Das Sharma, Shreya A1 - Pal, Rakhi A1 - Reddy, Bharath Kumar A1 - Selvaraj, Bhuvaneish T A1 - Raj, Nisha A1 - Samaga, Krishna Kumar A1 - Srinivasan, Durga J A1 - Ornelas, Loren A1 - Sareen, Dhruv A1 - Livesey, Matthew R A1 - Bassell, Gary J A1 - Svendsen, Clive N A1 - Kind, Peter C A1 - Chandran, Siddharthan A1 - Chattarji, Sumantra A1 - Wyllie, David J A AB -
BACKGROUND: Fragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder. Notwithstanding the extensive studies using rodent and other pre-clinical models of FXS, which have provided detailed mechanistic insights into the pathophysiology of this disorder, it is only relatively recently that human stem cell-derived neurons have been employed as a model system to further our understanding of the pathophysiological events that may underlie FXS. Our study assesses the physiological properties of human pluripotent stem cell-derived cortical neurons lacking fragile X mental retardation protein (FMRP).
METHODS: Electrophysiological whole-cell voltage- and current-clamp recordings were performed on two control and three FXS patient lines of human cortical neurons derived from induced pluripotent stem cells. In addition, we also describe the properties of an isogenic pair of lines in one of which FMR1 gene expression has been silenced.
RESULTS: Neurons lacking FMRP displayed bursts of spontaneous action potential firing that were more frequent but shorter in duration compared to those recorded from neurons expressing FMRP. Inhibition of large conductance Ca-activated K currents and the persistent Na current in control neurons phenocopies action potential bursting observed in neurons lacking FMRP, while in neurons lacking FMRP pharmacological potentiation of voltage-dependent Na channels phenocopies action potential bursting observed in control neurons. Notwithstanding the changes in spontaneous action potential firing, we did not observe any differences in the intrinsic properties of neurons in any of the lines examined. Moreover, we did not detect any differences in the properties of miniature excitatory postsynaptic currents in any of the lines.
CONCLUSIONS: Pharmacological manipulations can alter the action potential burst profiles in both control and FMRP-null human cortical neurons, making them appear like their genetic counterpart. Our studies indicate that FMRP targets that have been found in rodent models of FXS are also potential targets in a human-based model system, and we suggest potential mechanisms by which activity is altered.
VL - 11 IS - 1 ER - TY - JOUR T1 - Improved detection of RNA foci in amyotrophic lateral sclerosis post-mortem tissue using BaseScope™ shows a lack of association with cognitive dysfunction. JF - Brain Commun Y1 - 2020 A1 - Mehta, Arpan R A1 - Selvaraj, Bhuvaneish T A1 - Barton, Samantha K A1 - McDade, Karina A1 - Abrahams, Sharon A1 - Chandran, Siddharthan A1 - Smith, Colin A1 - Gregory, Jenna M AB -The hexanucleotide repeat expansion is the commonest known genetic mutation in amyotrophic lateral sclerosis. A neuropathological hallmark is the intracellular accumulation of RNA foci. The role that RNA foci play in the pathogenesis of amyotrophic lateral sclerosis is widely debated. Historically, RNA foci have been identified using hybridization. Here, we have implemented BaseScope™, a high-resolution modified hybridization technique. We demonstrate that previous studies have underestimated the abundance of RNA foci in neurons and glia. This improved detection allowed us to investigate the abundance, regional distribution and cell type specificity of antisense RNA foci in post-mortem brain and spinal cord tissue of six deeply clinically phenotyped patients and six age- and sex-matched controls. We find a correlation between RNA foci and the accumulation of transactive response DNA-binding protein of 43 kDa in spinal motor neurons ( = 0.93; = 0.008), but not in glia or cortical motor neurons. We also demonstrate that there is no correlation between the presence of RNA foci and the accumulation of transactive response DNA binding protein of 43 kDa in extra-motor brain regions. Furthermore, there is no association between the presence of RNA foci and cognitive indices. These results highlight the utility of BaseScope™ in the clinicopathological assessment of the role of antisense RNA foci in .
VL - 2 IS - 1 ER - TY - JOUR T1 - FMRP Interacts with C/D Box snoRNA in the Nucleus and Regulates Ribosomal RNA Methylation. JF - iScience Y1 - 2018 A1 - D'Souza, Michelle Ninochka A1 - Gowda, Naveen Kumar Chandappa A1 - Tiwari, Vishal A1 - Babu, Rosana Ottakandathil A1 - Anand, Praveen A1 - Dastidar, Sudhriti Ghosh A1 - Singh, Randhir A1 - James, Owen G A1 - Selvaraj, Bhuvaneish A1 - Pal, Rakhi A1 - Ramesh, Arati A1 - Chattarji, Sumantra A1 - Chandran, Siddharthan A1 - Gulyani, Akash A1 - Palakodeti, Dasaradhi A1 - Muddashetty, Ravi S AB -FMRP is an RNA-binding protein that is known to localize in the cytoplasm and in the nucleus. Here, we have identified an interaction of FMRP with a specific set of C/D box snoRNAs in the nucleus. C/D box snoRNAs guide 2'O methylations of ribosomal RNA (rRNA) on defined sites, and this modification regulates rRNA folding and assembly of ribosomes. 2'O methylation of rRNA is partial on several sites in human embryonic stem cells, which results in ribosomes with differential methylation patterns. FMRP-snoRNA interaction affects rRNA methylation on several of these sites, and in the absence of FMRP, differential methylation pattern of rRNA is significantly altered. We found that FMRP recognizes ribosomes carrying specific methylation patterns on rRNA and the recognition of methylation pattern by FMRP may potentially determine the translation status of its target mRNAs. Thus, FMRP integrates its function in the nucleus and in the cytoplasm.
VL - 9 ER -