biostudies-arrayexpressAnna-Leigh BrownHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-15653TDP-43 knockdown produces aberrant NMD-sensitive cryptic exons. To discover where TDP-43 cryptic exons were localized and which were sensitive to NMD-inhibition, we performed cellular fractionation as well and NMD-inhibition with TDP-43 KDbiostudies-arrayexpressLibrary Construction - RNA from the nuclear and the cytosolic fractions was extracted with the Direct-zol kit (Zymo) with on-column DNase I treatment. For each experimental condition, 2 μg of cytoplasmic RNA and an equal volume of nuclear RNA fraction were reverse-transcribed with RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific) according to the manufacturer's instructionsSequencing - Samples were sequenced at 2 x 150 base pairs on an Illumina HiSeq 2500 machine.Sample Treatment - For the fractionation experiments, SH-SY5Y cells were treated for 10 days with 25 ng/ml doxycycline hyclate (Sigma) to induce the shRNA against TDP-43, and for 24 hours with 0.5 μM SMG1-11j inhibitor32 to block the NMD machinery. After 10 days, cells were trypsinised, pelleted and resuspended in 1X PBS (Thermo). A fraction of the resuspended cells was pelleted and used for the subcellular fractionation with the Ambion PARIS Kit (Life Technologies), according to the manufacturer's instructions. RNA from the nuclear and the cytosolic fractions was extracted with the Direct-zol kit (Zymo) with on-column DNase I treatment. For each experimental condition, 2 μg of cytoplasmic RNA and an equal volume of nuclear RNA fraction were reverse-transcribed with RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific) according to the manufacturer's instructions.Sample Collection - s, SH-SY5Y cells were treated for 10 days with 25 ng/ml doxycycline hyclate (Sigma) to induce the shRNA against TDP-43, and for 24 hours with 0.5 μM SMG1-11j inhibitor32 to block the NMD machinery. After 10 days, cells were trypsinised, pelleted and resuspended in 1X PBS (Thermo).Nucleic Acid Extraction - RNA from the nuclear and the cytosolic fractions was extracted with the Direct-zol kit (Zymo) with on-column DNase I treatment. For each experimental condition, 2 μg of cytoplasmic RNA and an equal volume of nuclear RNA fraction were reverse-transcribed with RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific) according to the manufacturer's instructionsOrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesSequence Alignment - Raw sequences (in fastq format) were trimmed using Fastp33 with the parameter “qualified_quality_phred: 10”, and aligned using STAR (v2.7.0f)34 to the GRCh38 with gene models from GENCODE v4035.Data Transformation - Trimmed fastqc files were aligned to the transcriptome using Salmon (v1.5.1)37, outputting isoform-specific counts used for differential gene expression, performed using DeSEQ238 without covariates, using an index built from GENCODE v4035.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsIllumina HiSeq 2500NARNA-seq of coding RNAHomo sapiensAnna-Leigh BrownfalseRNA-sequencing in SH-SY5Y neuroblastoma cell line with TDP-43 KD and nuclear and cytoplasmic fractionation as well and treatment with the nonsense-mediated decay inhibition using SMG1-11j inhibitorTDP-43 knockdown produces aberrant NMD-sensitive cryptic exons. To discover where TDP-43 cryptic exons were localized and which were sensitive to NMD-inhibition, we performed cellular fractionation as well and NMD-inhibition with TDP-43 KD2026-07-04T00:00:00Z2026-07-04T01:00:55.286Z2025-10-01T12:22:54.784ZE-MTAB-15653ERP180868E-MTAB-15433EFO_0002944EFO_0004170EFO_0004917EFO_0005518EFO_0003816EFO_0003738EFO_0004184EFO_0003969