Unknown,Transcriptomics,Genomics,Proteomics

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The expression profiles of 30 transcription factors in 17 inducible gain-of-function lines after activation of overexpression, including 5 time points

ABSTRACT: We performed a large-scale expression analysis with inducible gain-of-function (GOF) lines which contain a C-terminal fusion protein of the TF of interest and a glucocorticoid receptor (GR) domain, driven by a constitutive 35S promoter. Such fusion proteins reside in the cytosol and can only translocate to the nucleus in the presence of dexamethasone (DEX), enabling the TF to regulate its downstream target genes (Corrado and Karali, 2009). In total 18 GOF lines were used of the following TFs: ERF-1 (AT4G17500), ERF2 (AT5G47220), ERF5 (AT5G47230), ERF6 (AT4G17490), ERF11 (AT1G28370), ERF8 (AT1G53170), ERF9 (AT5G44210), ERF59 (AT1G06160), ERF98 (AT3G23230), MYB51 (AT1G18570), STZ (AT1G27730), WRKY28 (AT4G18170), WRKY33 (AT2G38470), WRKY15 (AT2G23320), ZAT6 (AT5G04340), WRKY48 (AT5G49520), RAP2.6L (AT5G13330) and the control 35S::GFP-GR. To get an indication of which genes are direct or indirect targets of the induced TF, we performed a time-course experiment. All GOF lines were transferred at 15 DAS to DEX-containing medium and the third leaf was harvested at 1 h, 2 h, 4 h, 8 h and 24 h after transfer. The expression of 30 TFs (17 TFs listed above and WRKY6 (AT1G62300), WRKY30 (AT5G24110) and WRKY40 (AT1G80840), GA2OX6 (AT1G02400), GA20OX1 (AT4G25420), ACS6 (AT4G11280), MPK3 (AT3G45640), DEL1 (AT3G48160), UVI4 (AT2G42260), CYCA3;1 (AT5G43080), CYCB1;2 (AT5G06150), XET (AT1G10550), EXPA5 (AT3G29030)) was measured with nCounter Nanostring (Geiss et al, 2008). The time-course experiment gives an indication of whether a gene is putatively a direct, and thus induced during the early time points, or an indirect target of the induced TF.

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: Dirk Inze

PROVIDER: E-MTAB-6209 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Background: Molecular adaptations in the striatum mediated by dopamine (DA) denervation or Levodopa (L-dopa) treatment have been implicated with the motor deficits found in Parkinsonâs disease (PD). Alterations in glutamatergic neurotransmission and anti-oxidant mechanisms are reported to play important roles in mediating these changes. However, the mechanisms mediating the molecular adaptations in the striatum are not well understood. In recent years, microRNAs (miRNAs) have been recognized as potent post-transcriptional regulators of gene expression with fundamental roles in numerous biological processes. miRNAs are known to influence the development and maintenance of striatal neurons. To determine the contribution of miRNAs in molecular adaptations found in PD, we screened the expression of 800 miRNAs in postmortem striatum samples obtained from PD and neurologically normal controls. We detected the expression of approximately 250 miRNAs in postmortem human putamen samples collected from patients with PD and healthy controls. There was an abundance of a subset of 17 miRNAs (10 up- and 7 down-regulated) differing substantially between PD and the control, suggesting that miRNAs are altered in the striatum during the course of PD. Computational analysis show that many of these miRNAs targets pro-inflammatory factors in the striatum. Therefore, we sought to detrmine the expression of experimentally validated pro-inflammatory transcripts in PD striatum. Methods: Using a digital gene expression platform to quantify 134 gene transcripts, we compared human postmortem putamen tissues from patients with PD and neurologically normal controls. Results: We identified deregulated expression of 10 gene transcripts (4 up- and 6 down-regulated mRNAs) in postmortem human putamen samples collected fromPD patients compared to controls. The expression of these genes negative correlates with the expression of many of the differentially epxressed miRNAs. Moreover, many of these genes are predicted targets of the differentially expressed miRNAs. Conclusions: We identified deregulated mRNAs most likely associated with anti-oxidant mechanims in PD striatum. This approach may provide insights into pathogenesis of the disease. Human postmortem putamen tissues from 12 patients with PD symptoms and 12 neurologically normal controls were used in the study. Samples were obtained from the Human Brain and Spinal Fluid Resource Center, Los Angeles, CA through NIH NeuroBioBank, and stored at -80Â°C until RNA isolation. Total RNA was extracted using the mirVana RNA isolation kit, following the manufacturerâs instructions (Ambion). Using 225ng total RNA, mRNA levels were assayed by direct digital detection using Nanostring nCounter assay kits (Nanostring Technologies).

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The expression profiles of 30 transcription factors in 17 inducible gain-of-function lines after activation of overexpression, including 5 time points

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