Unknown,Transcriptomics,Genomics,Proteomics

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Expression data from primary MEF lacking either HDAC1, HDAC2 or both

ABSTRACT: Previously published data suggested some redundant functions between HDAC1 and HDAC2 in mouse. To test this hypothesis, we used microarrays to have a genome wide analysis at the transcription level of primary MEFs lacking HDAC1, HDAC2. MEF HDAC1 F/F were were transduced with two different retroviruses: one virus expresses the Tamoxifen-inducible cre recombinase Cre-ERT2 and the second virus expresses either a small hairpin micro-RNA against HDAC2 or a scrambled version. HDAC1F/F MEFs expressing either a scrambled micro-RNA or a micro-RNA against HDAC2 can be induced by addition of Tamoxifen to delete HDAC1, thereby generating four different genotypes: WT, HDAC1 KO, HDAC2 knockdown (Kd) and HDAC1/2 KO/Kd.

ORGANISM(S): Mus musculus

SUBMITTER: Patrick Matthias

PROVIDER: E-GEOD-20100 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Histone deacetylases (Hdac) remove acetyl groups from proteins, influencing global and specific gene expression. Hdacs control inflammation, as shown by Hdac inhibitor-dependent protection from DSS-induced murine colitis. While tissue-specific Hdac knockouts show redundant and specific functions, little is known of their intestinal epithelial cell (IEC) role. We have shown previously that dual Hdac1/Hdac2 IEC-specific loss disrupts cell proliferation and determination, with decreased secretory cell numbers and altered barrier function. We thus investigated how compound Hdac1/Hdac2 or Hdac2 IEC-specific deficiency alters the inflammatory response. Floxed Hdac1 and Hdac2 and villin-Cre mice were interbred. Compound Hdac1/Hdac2 IEC-deficient mice showed chronic basal inflammation, with increased basal Disease Activity Index (DAI) and deregulated Reg gene colonic expression. DSS-treated dual Hdac1/Hdac2 IEC-deficient mice displayed increased DAI, histological score, intestinal permeability and inflammatory gene expression. In contrast to double knockouts, Hdac2 IEC-specific loss did not affect IEC determination and growth, nor result in chronic inflammation. However, Hdac2 disruption protected against DSS colitis, as shown by decreased DAI, intestinal permeability and caspase-3 cleavage. Hdac2 IEC-specific deficient mice displayed increased expression of IEC gene subsets, such as colonic antimicrobial Reg3b and Reg3g mRNAs, and decreased expression of immune cell function-related genes. Our data show that Hdac1 and Hdac2 are essential IEC homeostasis regulators. IEC-specific Hdac1 and Hdac2 may act as epigenetic sensors and transmitters of environmental cues and regulate IEC-mediated mucosal homeostatic and inflammatory responses. Different levels of IEC Hdac activity may lead to positive or negative outcomes on intestinal homeostasis during inflammation Total RNAs from the colon of three control and three Hdac2 IEC-specific knockout mice were isolated with the Rneasy kit (Qiagen, Mississauga, ON, Canada).

Project description:Histone deacetylases (HDAC) are enzymes that modify key residues in histones to regulate chromatin architecture, and play a vital role in cell survival, cell cycle progression, and tumorigenesis. To understand the function of Hdac3, a critical component of the N-CoR/SMRT repression complex, a conditional allele of Hdac3 was engineered. Given the vital role of HDAC3 in normal cells and in the generation and treatment of various cancers, a conditional deletion of Hdac3 was engineered in mice. The deletion of Hdac3 in the germ line yielded very early embryonic lethality. Therefore, mouse embryonic-derived fibroblasts (MEFs) containing the Hdac3 conditional allele were used to examine the function of Hdac3 in cell viability and in cell cycle control. Given the links between Hdac3 and transcriptional repression, we used oligonucleotide micro arrays to examine RNA isolated from control and Cre recombinase expressing MEFs to define the changes in gene expression upon inactivation of Hdac3. Initially, we compared RNA from Ad-Cre infected Hdac3FL/+ and Hdac3FL/- MEFs and then we extended the analysis to Hdac3FL/- MEFs expressing Cre-ER (tamoxifen responsive Cre-estrogen receptor fusion protein). When the gene expression of ethanol-treated (vehicle) and tamoxifen-treated MEFs was compared at 72 hr post-induction, 83 genes were induced and 111 genes were down-regulated at least 1.5-fold upon inactivation of Hdac3. Approximately, 48 genes were up-regulated in both Ad-Cre and tamoxifen-treated MEFs at the 72 hr time point. The majority of the affected genes were associated with signal transduction and metabolism. Overall, few of the up-regulated genes at the 72hr time point in Ad-Cre and tamoxifen-treated MEFs are associated with apoptosis or with cell cycle control. Thus, these metabolic regulatory genes may be “core” Hdac3-regulated genes, and alterations in gene expression are unlikely to trigger apoptosis. Keywords: Conditional knock out analysis, genetic modification

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Expression data from primary MEF lacking either HDAC1, HDAC2 or both

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