OMICS Analyses Unraveling Related Gene and Protein Driven Molecular Mechanisms Underlying PACAP 38 Induced Neurite Outgrowth in PC12 Cells
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PROVIDER: PRJNA925830 | ENA |
REPOSITORIES: ENA
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Project description:Recently, our group has shown that the pituitary adenylate cyclase-activating polypeptide (PACAP)-induced neurite projection elongation in rat adrenal-derived pheochromocytoma cell line (PC12) is mediated via PAC1 receptor-mediated dephosphorylation of CRMP2 majorly through PI3K/AKT and MEK/ERK pathways. However, the mechanism of neuronal outgrowth by PACAP, including CRMP2 dephosphorylation, remains unclear. In our previous report, we found that GSK-3β, CDK5, and Rho/ROCK dephosphorylated CRMP2 within 3 hours after the addition of PACAP, but the early dephosphorylation of CRMP2 by PACAP remains unclear. Thus, we considered it important to identify early factors in PACAP-induced neurite projection elongation and performed omics-based transcriptomic (whole genome DNA microarray) and proteomic (TMT-labeling in conjunction with liquid chromatography-tandem mass spectrometry) analyses of gene and protein expression profiles from 5-120 minutes after PACAP addition. Results surprisingly revealed a number of key regulators involved in neurite outgrowth, including known ones. We also identified factors that may be involved in CRMP2 dephosphorylation. Cross-referencing previous research, we tried to map these molecular components onto potential pathways. The results of this comprehensive analysis may provide important new information on the molecular mechanisms of neuronal differentiation induced by PACAP.
2023-01-25 | GSE223333 | GEO
Project description:Recently, our group has shown that the pituitary adenylate cyclase-activating polypeptide (PACAP)-induced neurite projection elongation in rat adrenal-derived pheochromocytoma cell line (PC12) is mediated via PAC1 receptor-mediated dephosphorylation of CRMP2 majorly through PI3K/AKT and MEK/ERK pathways. However, the mechanism of neuronal outgrowth by PACAP, including CRMP2 dephosphorylation, remains unclear. In our previous report, we found that GSK-3β, CDK5, and Rho/ROCK dephosphorylated CRMP2 within 3 hours after the addition of PACAP, but the early dephosphorylation of CRMP2 by PACAP remains unclear. Thus, we considered it important to identify early factors in PACAP-induced neurite projection elongation and performed omics-based transcriptomic (whole genome DNA microarray) and proteomic (TMT-labeling in conjunction with liquid chromatography-tandem mass spectrometry) analyses of gene and protein expression profiles from 5-120 minutes after PACAP addition. Results surprisingly revealed a number of key regulators involved in neurite outgrowth, including known ones. We also identified factors that may be involved in CRMP2 dephosphorylation. Cross-referencing previous research, we tried to map these molecular components onto potential pathways. The results of this comprehensive analysis may provide important new information on the molecular mechanisms of neuronal differentiation induced by PACAP.
2023-03-11 | PXD039992 | Pride
Project description:This SuperSeries is composed of the following subset Series:; GSE9738: Analysis of gene expression during neurite outgrowth and regeneration (430A and 420A 2.0 array); GSE9739: Analysis of gene expression during neurite outgrowth and regeneration (MG-U74A); GSE9740: Analysis of gene expression during neurite outgrowth and regeneration MG-U74B Experiment Overall Design: Refer to individual Series
2008-04-10 | E-GEOD-9744 | biostudies-arrayexpress
Project description:We have undertaken a genome-wide study of transcriptional activity in embryonic superior cervical ganglia (SCG) and dorsal root ganglia (DRG) during a time course of neurite outgrowth in vitro. Gene expression observed in these models likely includes both developmental gene expression patterns and regenerative responses to axotomy, which occurs as the result of tissue dissection. Comparison across both models revealed many genes with similar gene expression patterns during neurite outgrowth. These patterns were minimally affected by exposure to the potent inhibitory cue Semaphorin3A, indicating that this extrinsic cue does not exert major effects at the level of nuclear transcription. We also compared our data to several published studies of DRG and SCG gene expression in animal models of regeneration, and found the expression of a large number of genes in common between neurite outgrowth in vitro and regeneration in vivo. Experiment Overall Design: We wished to determine the transcriptional profiles of neurons undergoing neurite outgrowth in vitro. We were particularly interested in finding genes whose expression is generally associated with the process of neurite outgrowth, rather than with cell type-specific effects. Thus, in order to avoid focusing on transcripts unique to one tissue type versus another, we used a comparative strategy to look for effects that were common to two tissue types and therefore more likely to be involved in the general process of neurite outgrowth. While these explants contain multiple cell types, we felt this was preferable to the more disruptive conditions required to dissociate neurons or obtain a pure neuron population. To this end, we monitored gene expression in cultured explants from SCG and DRG using DNA microarrays. We initiated our studies by culturing embryonic day 13 (E13) mouse SCG in vitro and harvesting tissue for RNA isolation at time points from 2 to 65 hours. Time points were selected to detect both fast, short-term responses (2, 5 and 12 hours), as well as sustained, long-term changes (24, 40, and 65 hours). Samples were hybridized to Affymetrix MG-U74v2 A and B microarrays, with RNA from acutely dissected explants serving as a baseline reference. We followed these experiments with a parallel analysis of a more heterogeneous tissue type, the DRG, which is more frequently used than SCG for in vivo studies of neurite regeneration. Cervical and upper thoracic DRG from E12 embryos were cultured with NGF (the same trophic support as in SCG cultures), harvested at time points from 2 to 40 hours, and hybridized to Affymetrix MOE 430A microarrays.
2008-06-16 | E-GEOD-9738 | biostudies-arrayexpress
Project description:MicroRNAs (miRNAs) are small non-coding RNAs that control protein expression through translational inhibition or mRNA degradation. MiRNAs have been implicated in diverse biological processes such as development, proliferation, apoptosis and differentiation. Upon treatment with nerve growth factor (NGF), rat pheochromocytoma PC12 cells elicit neurite outgrowth and differentiae into neuron-like cells. NGF plays a critical role not only in neuronal differentiation but also in protection against apoptosis. In an attempt to identify NGF-regulated miRNAs in PC12 cells, we performed miRNAM-cM-^@M-^@microarray analysis using total RNAs harvested from cells treated with NGF. In response to NGF treatment, expression of 8 and 12 miRNAs were up- and down-regulated, respectively. Quantitative RT-PCR analysis confirmed increased expression of miR-221, miR-181a* and miR-326, and decreased expression of miR-143, miR-210 and miR-532-3p after NGF treatment, among which miR-221 was drastically up-regulated. Overexpression of miR-221 induced neurite outgrowth of PC12 cells in the absence of NGF treatment, and also enhanced neurite outgrowth caused by low-dose NGF. More importantly, knockdown of miR-221 by antagomir attenuated NGF-mediated neurite outgrowth. Finally, miR-221 decreased expression of Foxo3a and Apaf-1, both of which are involved in apoptosis in PC12 cells. Our results indicate that miR-221 plays a critical role for neuronal differentiation as well as protection against apoptosis in PC12 cells. NGF induced miRNAs expression in rat pheochromocytoma PC12 cells was measured in cells treated with 100 ng/ml NGF for 0, 12, 24 and 48 hr.
2012-05-15 | E-GEOD-32122 | biostudies-arrayexpress
Project description:MicroRNAs (miRNAs) are small non-coding RNAs that control protein expression through translational inhibition or mRNA degradation. MiRNAs have been implicated in diverse biological processes such as development, proliferation, apoptosis and differentiation. Upon treatment with nerve growth factor (NGF), rat pheochromocytoma PC12 cells elicit neurite outgrowth and differentiae into neuron-like cells. NGF plays a critical role not only in neuronal differentiation but also in protection against apoptosis. In an attempt to identify NGF-regulated miRNAs in PC12 cells, we performed miRNA microarray analysis using total RNAs harvested from cells treated with NGF. In response to NGF treatment, expression of 8 and 12 miRNAs were up- and down-regulated, respectively. Quantitative RT-PCR analysis confirmed increased expression of miR-221, miR-181a* and miR-326, and decreased expression of miR-143, miR-210 and miR-532-3p after NGF treatment, among which miR-221 was drastically up-regulated. Overexpression of miR-221 induced neurite outgrowth of PC12 cells in the absence of NGF treatment, and also enhanced neurite outgrowth caused by low-dose NGF. More importantly, knockdown of miR-221 by antagomir attenuated NGF-mediated neurite outgrowth. Finally, miR-221 decreased expression of Foxo3a and Apaf-1, both of which are involved in apoptosis in PC12 cells. Our results indicate that miR-221 plays a critical role for neuronal differentiation as well as protection against apoptosis in PC12 cells.
2012-05-16 | GSE32122 | GEO
Project description:The present research investigates the molecular mechanism of neurite outgrowth (protrusion elongation) under pituitary adenylate cyclase-activating polypeptide (PACAP) 38 treatments using a rat adrenal-derived pheochromocytoma cell line-PC12. This study specifically looks into the regulation of PACAP38-induced collapsing response mediator protein 2 (CRMP2) previously identified in a mouse brain ischemia model and which could be recovered by PACAP38 treatment. Previously, DNA microarray analysis revealed that PACAP 38-mediated neuroprotection involved not only CRMP2 but also pathways related to glycogen synthase kinase-3β (GSK-3β) and other signaling components. Thus, to clarify whether CRMP2 acts directly on PACAP38 or through GSK-3β as part of the mechanism of PACAP38-induced neurite outgrowth, we observed neurite outgrowth in the presence of GSK-3β inhibitors and activators. PC12 cells were treated with PACAP38 being added to the cell culture medium at concentrations of 10-7 M, 10-8 M, and 10-9 M. Post PACAP38 treatment, immunostaining was used to confirm protrusion elongation of the PC12 cells, while RT-PCR, two-dimensional gel electrophoresis in conjunction with Western blotting, and inhibition experiments were performed to confirm the expression of the PACAP gene, its receptors, and downstream signaling components. Our data show that neurite protrusion elongation by PACAP38 (10-7 M) in PC12 cells is mediated through the PAC1-R receptor as demonstrated by its suppression by a specific inhibitor PA-8. Inhibitor experiments suggested that PACAP38-triggered neurite protrusion follows a GSK-3β-regulated pathway, where the AKT and cAMP/ERK pathways are involved and where the inhibition of Rho/Roc could enhance neurite protrusion under PACAP38 stimulation. Although we could not yet confirm the exact role and position of CRMP2 in PACAP38-mediated PC12 cell elongation, it appears that its phosphorylation and dephosphorylation have a correlation with the neurite protrusion elongation through the interplay of CDK5, which needs to be investigated further.
| S-EPMC8371652 | biostudies-literature
Project description:The Dp71 protein is the most abundant dystrophin in the central nervous system (CNS). Several dystrophin Dp71 isoforms have been described and are classified into three groups, each with a different C- terminal end. However, the functions of Dp71 isoforms remain unknown. In the present study, we analyse the effect of Dp71eΔ71 overexpression on the neuronal differentiation of PC12 Tet-On cells. Overexpression of the dystrophin Dp71eΔ71 stimulates neuronal differentiation, increasing the percentage of cells with neurites and the neurite length. According to the 2-DE analysis, Dp71eΔ71 overexpression modified the protein expression profile of PC12 Tet-On cells that had been treated with neuronal growth factor (NGF) for nine days. Interestingly, all differentially expressed proteins were up-regulated compared to the control. The proteomic analysis showed that Dp71eΔ71 increases the expression of proteins with important roles in the differentiation process, such as the HspB1, S100A6, and K8 proteins involved in the cytoskeletal structure and the HCNP protein involved in neurotransmitter synthesis. The expression of the neuronal marker TH was also up- regulated.
2018-04-09 | PXD009114 | Pride
Project description:The study aimed to understand mechanism/s of neuronal outgrowth in the rat adrenal-derived pheochromocytoma cell line (PC12) under pituitary adenylate cyclase-activating polypeptide (PACAP) treatment. Neurite projection elongation was suggested to be mediated via Pac1 receptor-mediated dephosphorylation of CRMP2, where GSK-3β, CDK5, and Rho/ROCK dephosphorylated CRMP2 within 3 h after addition of PACAP, but the dephosphorylation of CRMP2 by PACAP remained unclear. Thus, we attempted to identify the early factors in PACAP-induced neurite projection elongation via omics-based transcriptomic (whole genome DNA microarray) and proteomic (TMT-labeled liquid chromatography-tandem mass spectrometry) analyses of gene and protein expression profiles from 5-120 min after PACAP addition. The results revealed a number of key regulators involved in neurite outgrowth, including known ones, called 'Initial Early Factors', e.g., genes Inhba, Fst, Nr4a1,2,3, FAT4, Axin2, and proteins Mis12, Cdk13, Bcl91, CDC42, including categories of 'serotonergic synapse, neuropeptide and neurogenesis, and axon guidance'. cAMP signaling and PI3K-Akt signaling pathways and a calcium signaling pathway might be involved in CRMP2 dephosphorylation. Cross-referencing previous research, we tried to map these molecular components onto potential pathways, and we may provide important new information on molecular mechanisms of neuronal differentiation induced by PACAP. Gene and protein expression data are publicly available at NCBI GSE223333 and ProteomeXchange, identifier PXD039992.
| S-EPMC9964364 | biostudies-literature
Project description:We have undertaken a genome-wide study of transcriptional activity in embryonic superior cervical ganglia (SCG) and dorsal root ganglia (DRG) during a time course of neurite outgrowth in vitro. Gene expression observed in these models likely includes both developmental gene expression patterns and regenerative responses to axotomy, which occurs as the result of tissue dissection. Comparison across both models revealed many genes with similar gene expression patterns during neurite outgrowth. These patterns were minimally affected by exposure to the potent inhibitory cue Semaphorin3A, indicating that this extrinsic cue does not exert major effects at the level of nuclear transcription. We also compared our data to several published studies of DRG and SCG gene expression in animal models of regeneration, and found the expression of a large number of genes in common between neurite outgrowth in vitro and regeneration in vivo. Keywords: time course
2007-12-04 | GSE9739 | GEO