1999 Media Guide

1999 Men\u27s Track and Field Media Guide, George Fox College

<i>BORIS</i> and <i>SBSN</i> expression in NSCLC clinical samples.

<p><i>BORIS</i> and <i>SBSN</i> expression in 11 samples from healthy individuals (N, left) and 28 samples from NSCLC patients (T, right). Expression was quantified relative to GAPDH. <i>SBSN</i> and <i>BORIS</i> co-express with p-value  = 0.0004 (by Fisher exact test). Clinical samples were ranked by <i>BORIS</i> expression level.</p

Reorganization of <i>SBSN</i> chromatin structure upon BORIS induction.

<p><i>BORIS</i> expression was induced by 0, 0.0313 or 1 µg/ml doxycycline 24 hours after transfection with control empty vector or with BORIS expressing vector. Cell nuclei were isolated and digested with MNase. Purified DNA after 8 min of digestion with MNase (Figure S6) was used in PCR for primer extension experiments as described in the Methods. Relative localization of the TSS and region of chromatin reorganization is indicated. DNA digestion outside of <i>SBSN</i> promoter was used as loading control. Note: DNA was most accessible for MNase digestion at 0 µg/ml doxycycline, even at overall underloading of this sample. M – Hi-Lo DNA ladder (BioRad).</p

Changes in histone modifications around <i>SBSN</i> TSS upon BORIS induction.

<p>(A) Enrichment of active chromatin modification, H3K4me3, near the <i>SBSN</i> TSS was measured by ChIP experiment with H3K4me3 antibodies for H358 transfected with BORIS and induced by indicated doxycycline concentrations. The region from +202 bp to +374 bp was analyzed. Note that cell lysates from the same experiments was used for all ChIP experiments (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g002" target="_blank">Figure 2C</a>). Enrichment level was measured relative to IgG in qRT-PCR as described in the Methods. (B) Enrichment of active chromatin modification – H3Ac. (C) Enrichment of repressive chromatin modification, H3K9me3. (*, p-value <0.00002 (A); p-value <0.004 (B); p-value <0.03 (C)).</p

Dose-Dependent Activation of Putative Oncogene <em>SBSN</em> by BORIS

<div><p>Testis-specific transcription factor BORIS (Brother of the Regulator of Imprinted Sites), a paralog and proposed functional antagonist of the widely expressed CTCF, is abnormally expressed in multiple tumor types and has been implicated in the epigenetic activation of cancer-testis antigens (CTAs). We have reported previously that suprabasin (<em>SBSN</em>), whose expression is restricted to the epidermis, is epigenetically derepressed in lung cancer. In this work, we establish that <em>SBSN</em> is a novel non-CTA target of BORIS epigenetic regulation. With the use of a doxycycline-inducible BORIS expressing vector, we demonstrate that relative BORIS dosage is critical for <em>SBSN</em> activation. At lower concentrations, BORIS induces demethylation of the <em>SBSN</em> CpG island and disruption and activation of chromatin around the <em>SBSN</em> transcription start site (TSS), resulting in a 35-fold increase in <em>SBSN</em> expression in the H358 human lung cancer cell line. Interestingly, increasing BORIS concentrations leads to a subsequent reduction in <em>SBSN</em> expression via chromatin repression. In a similar manner, increase in BORIS concentrations leads to eventual decrease of cell growth and colony formation. This is the first report demonstrating that different amount of BORIS defines its varied effects on the expression of a target gene via chromatin structure reorganization.</p> </div

Smaller concentrations of BORIS induce H358 cell growth.

<p>Cell proliferation assay for H358 cells transfected with BORIS or control empty vectors and induced by indicated doxycycline concentrations. Values are the mean ± SEM of pentaplicate cultures in 96-well dish. (*, p-value <0.04).</p

Dose-dependent changes in <i>SBSN</i> parameters induced by BORIS.

<p>Relative changes in different parameters were ranked from (−) to (++++) in according to results from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g001" target="_blank">Figures 1</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g002" target="_blank">2</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g003" target="_blank">3</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g004" target="_blank">4</a>, and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040389#pone-0040389-g005" target="_blank">5</a>.</p

BORIS-dependent DNA demethylation of <i>SBSN</i> CpG island.

<p>DNA was isolated from H358 cells transfected with BORIS expressing vector and induced by indicated doxycycline concentrations. DNA was treated by bisulfite, purified and used for QMSP with <i>SBSN</i>-specific primers and probes at <i>SBSN</i> as described in methods. Values are normalized to beta-actin unmethylated control. (*, p-value <0.03).</p

Aberrant promoter methylation in the DNAs from salivary rinses collected with or without an exfoliating brush from 57 HNSCC cancer patients.

<p>Each column represents a patient, and each row the methylation status of the given gene in salivary samples collected with brush or without brush. Black shading indicates promoter hypermethylation and white indicates lack of promoter methylation. WB, salivary rinses collected with an exfoliating brush; WOB, salivary rinses collected without brush.</p

Concordance between salivary rinses collected with or without an exfoliating brush from 57 HNSCC patients.

§<p>WB = salivary rinses collected with an exfoliating brush.</p>‡<p>WOB = salivary rinses collected without a brush.</p>§§<p>CI = Confidence Interval.</p