Mouse prostate strain-associated gene expression and analysis in human prostate tissues: FVB/N and C57BL/6

Genes differentially expressed in prostates of FVB/N and C57BL/6 strains. Heat map colors reflect fold ratio values between sample and reference pool. Columns 1-4 represent biological replicates for each strain. Rows represent individual genes. Values shown in red are relatively larger than the overall mean; values shown in green are relatively smaller than the overall mean. Transcript abundance levels in benign human prostate tissues associated with high grade (7-10) or low grade (≤6) adenocarcinomas for each gene determined to be altered in mouse strain comparisons where a corresponding ortholog was identified. Genes depected in (a) and (b) are in identical order. Black box (b) and text (a) represent genes with significant differential expression in the human datasets altered in the expected orientation. Gray box (b) and text (a) represent genes with significant differential expression in the human datasets altered in the opposite orientation. Transcript alterations for selected genes in benign tissue samples associating with high (Gleason 7-10) and low (Gleason ≤6) prostate cancers. Plots represent the 95% confidence intervals of logexpression ratios of tissues samples relative to a cell line reference.<p><b>Copyright information:</b></p><p>Taken from "Genetic background influences murine prostate gene expression: implications for cancer phenotypes"</p><p>http://genomebiology.com/2007/8/6/R117</p><p>Genome Biology 2007;8(6):R117-R117.</p><p>Published online 18 Jun 2007</p><p>PMCID:PMC2394769.</p><p></p

Analysis of strain-dependent differences in prostate gene expression by qRT-PCR

RNAs from preparations used in the microarray analysis or microdissected epithelium were reverse transcribed and amplified using qRT-PCR with primers specific for (), (), () and (). Ribosomal protein S16 expression levels were used to normalize qRT-PCR data. Normalized results are expressed relative to the lowest expressing value. Error bars indicate the standard deviation of four biological independent replicates. qRT-PCR for microdissected epithelium is represented by one sample per strain for each gene. White bars denote measurements from the microarray analysis. Black bars denote measurements generated by qRT-PCR from whole prostate. Diagonal lines denote measurements generated by qRT-PCR from microdissected prostate epithelium.<p><b>Copyright information:</b></p><p>Taken from "Genetic background influences murine prostate gene expression: implications for cancer phenotypes"</p><p>http://genomebiology.com/2007/8/6/R117</p><p>Genome Biology 2007;8(6):R117-R117.</p><p>Published online 18 Jun 2007</p><p>PMCID:PMC2394769.</p><p></p

Mouse prostate strain-associated gene expression and analysis in human prostate tissues: BALB/c and C57BL/6

Genes differentially expressed in prostates of BALB/c (BALB) and C57BL/6 (C57) strains. Heat map colors reflect fold ratio values between sample and reference pool. Columns 1-4 represent biological replicates for each strain. Rows represent individual genes. Values shown in red are relatively larger than the overall mean; values shown in green are relatively smaller than the overall mean. Transcript abundance levels in benign human prostate tissues associated with high grade (7-10) or low grade (≤6) adenocarcinomas for each gene determined to be altered in mouse strain comparisons where a corresponding ortholog was identified. Genes depicted in (a) and (b) are in identical order. Black box (b) and text (a) represent genes with significant differential expression in the human datasets altered in the expected orientation. Gray box (b) and text (a) represent genes with significant differential expression in the human datasets altered in the opposite orientation. Transcript alterations for selected genes in benign tissue samples associating with high (Gleason 7-10) and low (Gleason ≤6) prostate cancers. Plots represent the 95% confidence intervals of logexpression ratios of tissues samples relative to a cell line reference.<p><b>Copyright information:</b></p><p>Taken from "Genetic background influences murine prostate gene expression: implications for cancer phenotypes"</p><p>http://genomebiology.com/2007/8/6/R117</p><p>Genome Biology 2007;8(6):R117-R117.</p><p>Published online 18 Jun 2007</p><p>PMCID:PMC2394769.</p><p></p

Expression of Hairy Target Genes Is Disrupted in <i>hairy</i> Mutant Embryos

<p>Whole mount in situ hybridization on wild-type (A, C, E, G, I, K, and M) or <i>hairy^7H</i> mutant (B, D, F, H, J, L, and N) embryos with probes recognizing <i>prd</i> (A and B), <i>stg</i> (C and D), <i>ImpL2</i> (E and F), <i>mae</i> (G and H), <i>egh</i> (I and J), <i>kayak</i> (K and L), or <i>Idgf2</i> (M and N). Anterior is to the left. Dorsal is up, except in (M) and (N), which are dorsal views.</p

Hairy Target Gene Expression Is Disrupted in the Mutant Background of the Cofactors Associated with a Particular Target

<div><p>Whole mount in situ hybridization on wild-type (A, E, and I), <i>groucho</i> germline clone</p> <p>(B, F, and J), <i>dCtBP</i> germline clone (C, G, and K), and <i>dSir2</i> mutant (D, H, and L) embryos with probes recognizing <i>stg</i> (A–D), <i>kayak</i> (E–H), or <i>prd</i> (I–L). Anterior is to the left. Dorsal is up.</p></div

Binding of Hairy to Class C (C-Box) Sites in Putative Targets In Vitro

<div><p>(A) Schematic diagram (not to scale) of C-boxes within putative Hairy targets. C-boxes (Hairy binding sites) are denoted by white boxes, black arrows indicate transcription start sites (Ra, Rb, and Rc), ATG denotes the initiating methionine, and capital letters indicate bases matching with the Hairy consensus C-box. The distances in kilobases of the C-boxes from transcription start sites are noted in gray.</p> <p>(B) EMSA with either GST or GST–Hairy and the <i>ac</i> h/E-1 oligonucleotide. Lane 1, probe alone; lane 2, binding to probe by GST; lanes 3–5, binding to probe by GST–Hairy. In lanes 4 and 5, binding to probe by GST–Hairy was in the presence of competitor unlabeled oligos. An arrow indicates the Hairy–DNA complex; comp_wt and comp_mut indicate wild-type and mutated cold probes, respectively.</p> <p>(C) EMSA with either GST or GST–Hairy to the C-boxes within the <i>stg</i> and <i>prd</i> genes. Lanes 1–5, GST and GST–Hairy binding to the <i>stg</i> C-box (location: 25072658); lanes 6–10, GST and GST–Hairy binding to the <i>prd</i> C-box. (location: 12074032). Lane order and annotations are as in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.0020178#pbio-0020178-g004" target="_blank">Figure 4</a>B.</p> <p>(D) EMSA with GST–Hairy to the same C-box within the <i>stg</i> 4.9-kb genomic fragment is not competed by the presence of mutant competitor unlabeled oligo. Lane 1, probe alone; lane 2, binding to GST; lane 3, binding to probe by GST–Hairy; lanes 4 and 5, binding to probe by GST–Hairy in the presence of wild-type and mutant competitor unlabeled oligos, respectively.</p> <p>(E) Differential binding to C-boxes within the <i>egh</i> gene. EMSA with either GST or GST–Hairy to C-boxes within the <i>egh</i> promoter and transcribed region. Binding to three putative C-box sites is shown: egh1 (location: 2341609), egh2 (location: 2350367), and egh3 (location: 2352168). Lanes 1, 5, and 9: probe alone; lanes 2, 6, and 10: binding to probes by GST; lanes 3, 7, and 11: binding to probes with GST–Hairy. Lanes 4, 8, and 12: binding with GST–Hairy in the presence of unlabeled wild-type competitor. C-box locations and promoter information generated using Apollo (Berkeley <i>Drosophila</i> Genome Project).</p></div

Hairy Shows Context-Dependent Association with Its Cofactors

<div><p>(A) Sites of Hairy binding and Hairy cofactor recruitment based on DamID. The gray lines depict the relative position on the chromosomes of the approximately 6200 cDNAs on the microarray chip. The blue dots below the line represent Hairy binding sites while the green (Groucho), red (dCtBP), and yellow (dSir2) dots represent the positions of cofactor recruitment.</p> <p>(B–D) Cofactor recruitment visualized on third instar larval salivary gland chromosomes. Polytene chromosome sets stained (green) with antibodies to Groucho (B), dCtBP (C), and dSir2 (D). All chromosomes were counterstained with DAPI (blue) to visualize the DNA.</p> <p>(E) Higher magnification view of chromosome arms 2L and 2R costained with Groucho (red) and Hairy (green), and the merged image.</p> <p>(F) Higher magnification view of chromosome arm 2L costained with dCtBP (red) and Hairy (green), and the merged image, compared to the predicted DamID map. Note that both the DamID projected map and polytene chromosomes have more dCtBP recruitment sites to the left of the dashed line than to the right of the dashed line.</p> <p>(G) Chromosome arm 3R stained with dSir2 (green), highlighting regional specificity of dSir2 recruitment.</p> <p>(H and I) Higher magnification view of the distal ends of chromosome arms 2R (H) and 3L (I) from (D), stained with dSir2 (green), showing regional specificity and lack of dSir2 recruitment, respectively.</p></div

Hairy Overlaps with Cofactors Differentially

<div><p>(A–C) Venn diagram showing the overlap between Hairy targets and those loci also binding to the cofactors Groucho (A), dCtBP (B), and dSir2 (C).</p> <p>(D) Venn diagram showing combined overlaps of Hairy with its three known cofactors.</p></div

Hairy Binds to a Specific Set of Transcriptional Targets

<div><p>(A and B) Comparison of DamID-identified targets for Hairy with the <i>Drosophila</i> Myc and Mad/Mnt family proteins. Venn diagram comparing DamID-identified Hairy downstream targets in Kc cells compared to the transcriptional activator dMyc (A) and the transcriptional repressor dMnt (B).</p> <p>(C) Venn diagram comparing DamID-identified Hairy targets from Kc cells and embryos.</p></div

Hairy Binds to Putative Target Loci on Polytene Chromosomes

<div><p>(A) Hairy binds to polytene region 1A, the location of the Hairy target, <i>ac.</i></p> <p>(B) Hairy is not found at 84A, the cytological location for <i>ftz.</i></p> <p>(C–F) Hairy also binds to polytene region 99A, the location of <i>stg</i> (C); polytene region 3A, the location of <i>egh</i> (D); polytene region 33C, the location of <i>prd</i> (E); and polytene region 82A, the location of <i>hkb</i> (F).</p> <p>(G–I) Hairy is recruited to the insertion site for the pstg βE-4.9 reporter construct (arrow in [H] and [I]). Compare to the equivalent region of wild-type X chromosomes marked by brackets in (A), (D), and (G).</p> <p>(J) In situ hybridization to polytene chromosomes from pstg βE-4.9 larvae showing that this line has two insertions on the X chromosome at 1F and 6C. The probe also recognizes sequences to the endogenous <i>white</i> locus (asterisk).</p></div