Prostate apoptosis response protein 4 sensitizes human colon cancer cells to chemotherapeutic 5-FU through mediation of an NFκB and microRNA network

Bi Dar Wang, Christina L.B. Kline, Danielle M. Pastor, Thomas L. Olson, Bryan Frank, Truong Luu, Arun Sharma, Gavin Robertson, Matthew T. Weirauch, Steven R. Patierno, Joshua M. Stuart, Rosalyn Irby, Norman H. Lee

Research output: Contribution to journalArticle

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Abstract

Background: Diminished expression or activity of prostate apoptosis response protein 4 (Par-4) has been demonstrated in a number of cancers, although reports on Par-4 expression during colon cancer progression are lacking. An understanding of the molecular events in conjunction with the genetic networks affected by Par-4 is warranted.Results: Colon cancer specimens derived from patients have significantly diminished expression of Par-4 mRNA relative to paired normal colon. Hence, the functional consequences of reintroducing Par-4 into HT29 colon cancer cells were assessed. Overexpression augmented the interaction of Par-4 with NFκB in the cytosol but not nucleus, and facilitated apoptosis in the presence of 5-fluorouracil (5-FU). Analogous findings were obtained when AKT1 pro-survival signaling was inhibited. Transcriptome profiling identified ~700 genes differentially regulated by Par-4 overexpression in HT29 cells. Nearly all Par-4-regulated genes were shown by promoter analysis to contain cis-binding sequences for NFκB, and meta-analysis of patient expression data revealed that one-third of these genes exist as a recurrent co-regulated network in colon cancer specimens. Sets of genes involved in programmed cell death, cell cycle regulation and interestingly the microRNA pathway were found overrepresented in the network. Noteworthy, Par-4 overexpression decreased NFκB occupancy at the promoter of one particular network gene DROSHA, encoding a microRNA processing enzyme. The resulting down-regulation of DROSHA was associated with expression changes in a cohort of microRNAs. Many of these microRNAs are predicted to target mRNAs encoding proteins with apoptosis-related functions. Western and functional analyses were employed to validate several predictions. For instance, miR-34a up-regulation corresponded with a down-regulation of BCL2 protein. Treating Par-4-overexpressing HT29 cells with a miR-34a antagomir functionally reversed both BCL2 down-regulation and apoptosis by 5-FU. Conversely, bypassing Par-4 overexpression by direct knockdown of DROSHA expression in native HT29 cells increased miR-34a expression and 5-FU sensitivity.Conclusion: Our findings suggest that the initiation of apoptotic sensitivity in colon cancer cells can be mediated by Par-4 binding to NFκB in the cytoplasm with consequential changes in the expression of microRNA pathway components.

Original languageEnglish (US)
Article number98
JournalMolecular Cancer
Volume9
DOIs
StatePublished - Apr 30 2010

Fingerprint

MicroRNAs
Fluorouracil
Colonic Neoplasms
HT29 Cells
Apoptosis
Down-Regulation
Genes
Proto-Oncogene Proteins c-bcl-2
prostate apoptosis response-4 protein
Messenger RNA
Gene Regulatory Networks
Gene Expression Profiling
Protein Binding
Cytosol
Meta-Analysis
Prostate
Cell Cycle
Colon
Cytoplasm
Cell Death

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Oncology
  • Cancer Research

Cite this

Wang, Bi Dar ; Kline, Christina L.B. ; Pastor, Danielle M. ; Olson, Thomas L. ; Frank, Bryan ; Luu, Truong ; Sharma, Arun ; Robertson, Gavin ; Weirauch, Matthew T. ; Patierno, Steven R. ; Stuart, Joshua M. ; Irby, Rosalyn ; Lee, Norman H. / Prostate apoptosis response protein 4 sensitizes human colon cancer cells to chemotherapeutic 5-FU through mediation of an NFκB and microRNA network. In: Molecular Cancer. 2010 ; Vol. 9.
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abstract = "Background: Diminished expression or activity of prostate apoptosis response protein 4 (Par-4) has been demonstrated in a number of cancers, although reports on Par-4 expression during colon cancer progression are lacking. An understanding of the molecular events in conjunction with the genetic networks affected by Par-4 is warranted.Results: Colon cancer specimens derived from patients have significantly diminished expression of Par-4 mRNA relative to paired normal colon. Hence, the functional consequences of reintroducing Par-4 into HT29 colon cancer cells were assessed. Overexpression augmented the interaction of Par-4 with NFκB in the cytosol but not nucleus, and facilitated apoptosis in the presence of 5-fluorouracil (5-FU). Analogous findings were obtained when AKT1 pro-survival signaling was inhibited. Transcriptome profiling identified ~700 genes differentially regulated by Par-4 overexpression in HT29 cells. Nearly all Par-4-regulated genes were shown by promoter analysis to contain cis-binding sequences for NFκB, and meta-analysis of patient expression data revealed that one-third of these genes exist as a recurrent co-regulated network in colon cancer specimens. Sets of genes involved in programmed cell death, cell cycle regulation and interestingly the microRNA pathway were found overrepresented in the network. Noteworthy, Par-4 overexpression decreased NFκB occupancy at the promoter of one particular network gene DROSHA, encoding a microRNA processing enzyme. The resulting down-regulation of DROSHA was associated with expression changes in a cohort of microRNAs. Many of these microRNAs are predicted to target mRNAs encoding proteins with apoptosis-related functions. Western and functional analyses were employed to validate several predictions. For instance, miR-34a up-regulation corresponded with a down-regulation of BCL2 protein. Treating Par-4-overexpressing HT29 cells with a miR-34a antagomir functionally reversed both BCL2 down-regulation and apoptosis by 5-FU. Conversely, bypassing Par-4 overexpression by direct knockdown of DROSHA expression in native HT29 cells increased miR-34a expression and 5-FU sensitivity.Conclusion: Our findings suggest that the initiation of apoptotic sensitivity in colon cancer cells can be mediated by Par-4 binding to NFκB in the cytoplasm with consequential changes in the expression of microRNA pathway components.",
author = "Wang, {Bi Dar} and Kline, {Christina L.B.} and Pastor, {Danielle M.} and Olson, {Thomas L.} and Bryan Frank and Truong Luu and Arun Sharma and Gavin Robertson and Weirauch, {Matthew T.} and Patierno, {Steven R.} and Stuart, {Joshua M.} and Rosalyn Irby and Lee, {Norman H.}",
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Prostate apoptosis response protein 4 sensitizes human colon cancer cells to chemotherapeutic 5-FU through mediation of an NFκB and microRNA network. / Wang, Bi Dar; Kline, Christina L.B.; Pastor, Danielle M.; Olson, Thomas L.; Frank, Bryan; Luu, Truong; Sharma, Arun; Robertson, Gavin; Weirauch, Matthew T.; Patierno, Steven R.; Stuart, Joshua M.; Irby, Rosalyn; Lee, Norman H.

In: Molecular Cancer, Vol. 9, 98, 30.04.2010.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Prostate apoptosis response protein 4 sensitizes human colon cancer cells to chemotherapeutic 5-FU through mediation of an NFκB and microRNA network

AU - Wang, Bi Dar

AU - Kline, Christina L.B.

AU - Pastor, Danielle M.

AU - Olson, Thomas L.

AU - Frank, Bryan

AU - Luu, Truong

AU - Sharma, Arun

AU - Robertson, Gavin

AU - Weirauch, Matthew T.

AU - Patierno, Steven R.

AU - Stuart, Joshua M.

AU - Irby, Rosalyn

AU - Lee, Norman H.

PY - 2010/4/30

Y1 - 2010/4/30

N2 - Background: Diminished expression or activity of prostate apoptosis response protein 4 (Par-4) has been demonstrated in a number of cancers, although reports on Par-4 expression during colon cancer progression are lacking. An understanding of the molecular events in conjunction with the genetic networks affected by Par-4 is warranted.Results: Colon cancer specimens derived from patients have significantly diminished expression of Par-4 mRNA relative to paired normal colon. Hence, the functional consequences of reintroducing Par-4 into HT29 colon cancer cells were assessed. Overexpression augmented the interaction of Par-4 with NFκB in the cytosol but not nucleus, and facilitated apoptosis in the presence of 5-fluorouracil (5-FU). Analogous findings were obtained when AKT1 pro-survival signaling was inhibited. Transcriptome profiling identified ~700 genes differentially regulated by Par-4 overexpression in HT29 cells. Nearly all Par-4-regulated genes were shown by promoter analysis to contain cis-binding sequences for NFκB, and meta-analysis of patient expression data revealed that one-third of these genes exist as a recurrent co-regulated network in colon cancer specimens. Sets of genes involved in programmed cell death, cell cycle regulation and interestingly the microRNA pathway were found overrepresented in the network. Noteworthy, Par-4 overexpression decreased NFκB occupancy at the promoter of one particular network gene DROSHA, encoding a microRNA processing enzyme. The resulting down-regulation of DROSHA was associated with expression changes in a cohort of microRNAs. Many of these microRNAs are predicted to target mRNAs encoding proteins with apoptosis-related functions. Western and functional analyses were employed to validate several predictions. For instance, miR-34a up-regulation corresponded with a down-regulation of BCL2 protein. Treating Par-4-overexpressing HT29 cells with a miR-34a antagomir functionally reversed both BCL2 down-regulation and apoptosis by 5-FU. Conversely, bypassing Par-4 overexpression by direct knockdown of DROSHA expression in native HT29 cells increased miR-34a expression and 5-FU sensitivity.Conclusion: Our findings suggest that the initiation of apoptotic sensitivity in colon cancer cells can be mediated by Par-4 binding to NFκB in the cytoplasm with consequential changes in the expression of microRNA pathway components.

AB - Background: Diminished expression or activity of prostate apoptosis response protein 4 (Par-4) has been demonstrated in a number of cancers, although reports on Par-4 expression during colon cancer progression are lacking. An understanding of the molecular events in conjunction with the genetic networks affected by Par-4 is warranted.Results: Colon cancer specimens derived from patients have significantly diminished expression of Par-4 mRNA relative to paired normal colon. Hence, the functional consequences of reintroducing Par-4 into HT29 colon cancer cells were assessed. Overexpression augmented the interaction of Par-4 with NFκB in the cytosol but not nucleus, and facilitated apoptosis in the presence of 5-fluorouracil (5-FU). Analogous findings were obtained when AKT1 pro-survival signaling was inhibited. Transcriptome profiling identified ~700 genes differentially regulated by Par-4 overexpression in HT29 cells. Nearly all Par-4-regulated genes were shown by promoter analysis to contain cis-binding sequences for NFκB, and meta-analysis of patient expression data revealed that one-third of these genes exist as a recurrent co-regulated network in colon cancer specimens. Sets of genes involved in programmed cell death, cell cycle regulation and interestingly the microRNA pathway were found overrepresented in the network. Noteworthy, Par-4 overexpression decreased NFκB occupancy at the promoter of one particular network gene DROSHA, encoding a microRNA processing enzyme. The resulting down-regulation of DROSHA was associated with expression changes in a cohort of microRNAs. Many of these microRNAs are predicted to target mRNAs encoding proteins with apoptosis-related functions. Western and functional analyses were employed to validate several predictions. For instance, miR-34a up-regulation corresponded with a down-regulation of BCL2 protein. Treating Par-4-overexpressing HT29 cells with a miR-34a antagomir functionally reversed both BCL2 down-regulation and apoptosis by 5-FU. Conversely, bypassing Par-4 overexpression by direct knockdown of DROSHA expression in native HT29 cells increased miR-34a expression and 5-FU sensitivity.Conclusion: Our findings suggest that the initiation of apoptotic sensitivity in colon cancer cells can be mediated by Par-4 binding to NFκB in the cytoplasm with consequential changes in the expression of microRNA pathway components.

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