Identification of transcription repressors that regulate
the expression of HMOX1 gene with SureFIND Transcriptome PCR Array
Heme oxygenase-1 (HMOX1) is an inducible enzyme that catalyzes oxidative
degradation of heme to form biliverdin, carbon monoxide and free iron.
HMOX1 expression is elevated in advanced stage breast-, gastric- and
prostate- cancers and shown to have protumoral function. HMOX1 response
also plays a role in drug resistance during chemotherapy (1). Although
function of HMOX1 gene has been intensively studied, the molecular
mechanism behind gene expression regulation of HMOX1, especially the
repression mechanism that normally active when no stimulants available is
important for understanding the tumor development and its drug resistance.
Transcription Factor Knockdown Transcriptome PCR Array provides
comprehensive experimental methods to screen for transcriptional regulators
for a gene of interest.
Materials and methods
In this study, we employed Cancer Transcription Factor Knockdown
Transcriptome PCR Array to simultaneously analyze the transcription
regulators for HMOX1 gene expression. SYBR green qPCR Assay was performed
to quantify the expression of HMOX1 (gene of interest) and RPL13A (house
keeping gene). Fold change in HMOX1 gene expression as a result of each
transcription regulator specific siRNA treatment relative to negative siRNA
control were calculated and normalized to RPL13A. Fold changes were
converted to log2 and subjected to MAD analysis for positive hits
Figure 1. Screening of HMOX1 gene expression repressors with
SureFIND. MCF7 cell based SureFIND Transcriptome PCR Array (Cancer Transcription
Factor Knockdown) was used with its 90 targets gene indicated. RT˛ qPCR Primer
Assays (SYBRŽ Green-based) for HMOX1 and RPL13A was performed. HMOX1 gene
expression level are expressed as Log2 fold changes based on Ct calculation
using RPL13A as house-keeping gene and non-target siRNA treated sample well (VTC)
as negative control.
Our results show that BACH1, MYBL2 and YY1 negatively regulate HMOX1 gene
expression. Knock down of BACH1, MYBL2 and YY1, showed about 12-fold, 8-fold and
10-fold increase in the level of HMOX1 mRNA compared to the negative control,
which was from cells treated with non-targeting siRNA, respectively (Figure1).
Interestingly, BACH1 is well known repressor for HMOX1 expression (2), whereas
MYBL2 and YY1 are novel regulators.
Transcription Factor Knockdown Transcriptome PCR Array identified three
transcription factors that repress HMOX1 gene expression and two of these (MYBL2
and YY1) were novel finding.
1. Jin Song, et al. (2009) Suppression of Annexin A11 in Ovarian Cancer:
Implications in Chemoresistance. Neoplasia. 11(6): 605.
2. Jiying Sun, et al. (2002) Hemoprotein Bach1 regulates enhancer availability
of heme oxygenase-1 gene. The EMBO Journal. 21(19). 5216
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