biostudies-arrayexpressBen SzaroXenopus laevishttps://www.ebi.ac.uk/biostudies/studies/E-GEOD-25207Cells use their wide variety of RNPs to integrate the expression of functionally inter-related proteins by forming RNP complexes with cis-elements that are shared among co-regulated RNAs. In this study, we identified the the associated mRNAs that co-precipitated with hnRNP K in developing juvenile frog brain. three replicates of immunoprecipitation experiments for both hnRNP K and beta-galactosidase(control)biostudies-arrayexpressNucleic Acid Extraction - The complete mRNP IP protocol is provided as a supplementary file on the Series record (GSE25207_IP_of_mRNP_complexes_protocol.txt). RNA from Immunoprecipitated samples was isolated by phenol chloroform extraction and further purified on Qiagen Rneasy columns before performing quality control analysis on Agilent Bioanalyzer chips.Hybridization - Standard Affymetrix protocol for Exon arrays.Labeling - Standard Affymetrix protocol for Exon arrays.MIAME ScoreRaw DataOrganizationAssays and DataProcessed DataMAGE-TAB FilesArray DesignsFeature Extraction - Raw data were corrected for background and transcripts scored as present or absent, using MAS 5.0 and further filtered to remove the bottom 30th percentile (GeneSpring GX 10).Assay Data Transformation - ID_REF = <br>VALUE = MAS5.0 signal intensityImage Adquisition - GeneChips were scanned using Affymetrix GeneChip Scanner.UnknownTranscriptomicsGenomicsProteomicsThe RNA-binding protein, hnRNP K, is essential for axonogenesis. Suppressing its expression in Xenopus embryos yields terminally specified neurons with severely disorganized microtubules, microfilaments and neurofilaments, raising the hypothesis that hnRNP K post-transcriptionally regulates multiple transcripts of proteins that organize the axonal cytoskeleton. To identify downstream candidates for this regulation, RNAs that co-immunoprecipitated from juvenile brain with hnRNP K were identified on microarrays. A substantial number of these transcripts were linked to the cytoskeleton and to intracellular localization, trafficking and transport. Injection into embryos of a non-coding RNA bearing multiple copies of an hnRNP K RNA-binding consensus sequence found within these transcripts largely phenocopied hnRNP K knockdown, further supporting the idea that it regulates axonogenesis through its binding to downstream target RNAs. For further study of regulation by hnRNP K of the cytoskeleton during axon outgrowth, we focused on three validated RNAs representing elements associated with all three polymers - Arp2, tau and an α-internexin-like neurofilament. All three were co-regulated post-transcriptionally by hnRNP K, as hnRNP K knockdown yielded comparable defects in their nuclear export and translation but not transcription. Directly knocking down expression of all three together, but not each one individually, substantially reproduced the axonless phenotype, providing further evidence that regulation of axonogenesis by hnRNP K occurs largely through pleiotropic effects on cytoskeletal-associated targets. These experiments provide evidence that hnRNP K is the nexus of a novel post-transcriptional regulatory module controlling the synthesis of proteins that integrate all three cytoskeletal polymers to form the axon.otherXenopus laevishnRNP K post-transcriptionally co-regulates multiple cytoskeletal genes needed for axonogenesis.Yuanyuan LiuLiu Y, Szaro BGBen SzarofalseRIP-Chip Analysis of hnRNP K in Xenopus juvenile brainCells use their wide variety of RNPs to integrate the expression of functionally inter-related proteins by forming RNP complexes with cis-elements that are shared among co-regulated RNAs. In this study, we identified the the associated mRNAs that co-precipitated with hnRNP K in developing juvenile frog brain. three replicates of immunoprecipitation experiments for both hnRNP K and beta-galactosidase(control)2011-07-05T00:00:00Z2023-08-11T15:55:41.281Z2022-02-02T21:46:27.6ZE-GEOD-25207GSE252072169352310.1242/dev.066993