Inhibition of transcription factor activity by nuclear compartment- associated Bcl-2

Cynthia A. Massaad, Bryce P. Portier, Giulio Taglialatela

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

Using a reporter gene assay in PC12, HEK293, HeLa, and NIH-3T3 cells, we show that the anti-apoptotic protein Bcl-2 significantly inhibits transcriptional activation of various transcription factors, including NFκB, AP1, CRE, and NFAT. A Bcl-2 mutant lacking its BH4 domain (ΔBH4) also inhibited transcription, whereas a Bcl-2 mutant lacking its transmembrane domain (ΔTM) was ineffective. Furthermore, Bcl-2 cliimeric proteins containing transmembrane domains from the mitochondrial protein monoamine oxidase B (HaoB) or the endoplasmic reticulum protein cytochrome b5 showed no effect on transcription factor activity. Subcellular localization studies showed that under conditions of transient transfection, the active Bcl-2 forms (wild type and ΔBH4) were predominantly found in the nuclear fraction, whereas the non-active forms (ΔTM, MaoB, and cytochrome b5) were in the non-nuclear fraction. Additionally, stably expressed Bcl-2 loses its ability to inhibit transcriptional activation and localizes predominantly to the non-nuclear fraction. Expression of FKBP38 (a chaperone that shuttles Bcl-2 to the mitochondria) removes co-expressed Bcl-2 from the nuclear fraction and reverses its effect on transcription factor activity. Finally, using an indticible gene expression system, we show that nuclear compartment-associated Bcl-2 prevents entry of NFκB subunits to the nucleus without affecting NFκB release from its cytosolic inhibitory subunit IκBα. These results suggest that (a) Bcl-2 suppresses transcriptional activity of multiple transcription factors; (b) Bcl-2 does not interfere with NFκB activation but prevents entrance of its active subunits to the nucleus; (c) membrane anchoring is required for this function of Bcl-2; and (d) association of Bcl-2 with the nuclear compartment is also necessary. We speculate that nuclear compartment-associated Bcl-2 may affect nuclear trafficking of multiple factors necessary for transcriptional activity.

Original languageEnglish (US)
Pages (from-to)54470-54478
Number of pages9
JournalJournal of Biological Chemistry
Volume279
Issue number52
DOIs
StatePublished - Dec 24 2004

Fingerprint

Transcription Factors
Cytochromes b5
Chemical activation
Transcriptional Activation
NIH 3T3 Cells
Apoptosis Regulatory Proteins
Mitochondria
Mitochondrial Proteins
Monoamine Oxidase
Transcription
Reporter Genes
Gene expression
Endoplasmic Reticulum
Transfection
Assays
Proteins
Genes
Association reactions
Membranes
Gene Expression

ASJC Scopus subject areas

  • Biochemistry

Cite this

Inhibition of transcription factor activity by nuclear compartment- associated Bcl-2. / Massaad, Cynthia A.; Portier, Bryce P.; Taglialatela, Giulio.

In: Journal of Biological Chemistry, Vol. 279, No. 52, 24.12.2004, p. 54470-54478.

Research output: Contribution to journalArticle

Massaad, Cynthia A. ; Portier, Bryce P. ; Taglialatela, Giulio. / Inhibition of transcription factor activity by nuclear compartment- associated Bcl-2. In: Journal of Biological Chemistry. 2004 ; Vol. 279, No. 52. pp. 54470-54478.
@article{beb47e2d8b3740568313f0ec1e06f9ab,
title = "Inhibition of transcription factor activity by nuclear compartment- associated Bcl-2",
abstract = "Using a reporter gene assay in PC12, HEK293, HeLa, and NIH-3T3 cells, we show that the anti-apoptotic protein Bcl-2 significantly inhibits transcriptional activation of various transcription factors, including NFκB, AP1, CRE, and NFAT. A Bcl-2 mutant lacking its BH4 domain (ΔBH4) also inhibited transcription, whereas a Bcl-2 mutant lacking its transmembrane domain (ΔTM) was ineffective. Furthermore, Bcl-2 cliimeric proteins containing transmembrane domains from the mitochondrial protein monoamine oxidase B (HaoB) or the endoplasmic reticulum protein cytochrome b5 showed no effect on transcription factor activity. Subcellular localization studies showed that under conditions of transient transfection, the active Bcl-2 forms (wild type and ΔBH4) were predominantly found in the nuclear fraction, whereas the non-active forms (ΔTM, MaoB, and cytochrome b5) were in the non-nuclear fraction. Additionally, stably expressed Bcl-2 loses its ability to inhibit transcriptional activation and localizes predominantly to the non-nuclear fraction. Expression of FKBP38 (a chaperone that shuttles Bcl-2 to the mitochondria) removes co-expressed Bcl-2 from the nuclear fraction and reverses its effect on transcription factor activity. Finally, using an indticible gene expression system, we show that nuclear compartment-associated Bcl-2 prevents entry of NFκB subunits to the nucleus without affecting NFκB release from its cytosolic inhibitory subunit IκBα. These results suggest that (a) Bcl-2 suppresses transcriptional activity of multiple transcription factors; (b) Bcl-2 does not interfere with NFκB activation but prevents entrance of its active subunits to the nucleus; (c) membrane anchoring is required for this function of Bcl-2; and (d) association of Bcl-2 with the nuclear compartment is also necessary. We speculate that nuclear compartment-associated Bcl-2 may affect nuclear trafficking of multiple factors necessary for transcriptional activity.",
author = "Massaad, {Cynthia A.} and Portier, {Bryce P.} and Giulio Taglialatela",
year = "2004",
month = "12",
day = "24",
doi = "10.1074/jbc.M407659200",
language = "English (US)",
volume = "279",
pages = "54470--54478",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "52",

}

TY - JOUR

T1 - Inhibition of transcription factor activity by nuclear compartment- associated Bcl-2

AU - Massaad, Cynthia A.

AU - Portier, Bryce P.

AU - Taglialatela, Giulio

PY - 2004/12/24

Y1 - 2004/12/24

N2 - Using a reporter gene assay in PC12, HEK293, HeLa, and NIH-3T3 cells, we show that the anti-apoptotic protein Bcl-2 significantly inhibits transcriptional activation of various transcription factors, including NFκB, AP1, CRE, and NFAT. A Bcl-2 mutant lacking its BH4 domain (ΔBH4) also inhibited transcription, whereas a Bcl-2 mutant lacking its transmembrane domain (ΔTM) was ineffective. Furthermore, Bcl-2 cliimeric proteins containing transmembrane domains from the mitochondrial protein monoamine oxidase B (HaoB) or the endoplasmic reticulum protein cytochrome b5 showed no effect on transcription factor activity. Subcellular localization studies showed that under conditions of transient transfection, the active Bcl-2 forms (wild type and ΔBH4) were predominantly found in the nuclear fraction, whereas the non-active forms (ΔTM, MaoB, and cytochrome b5) were in the non-nuclear fraction. Additionally, stably expressed Bcl-2 loses its ability to inhibit transcriptional activation and localizes predominantly to the non-nuclear fraction. Expression of FKBP38 (a chaperone that shuttles Bcl-2 to the mitochondria) removes co-expressed Bcl-2 from the nuclear fraction and reverses its effect on transcription factor activity. Finally, using an indticible gene expression system, we show that nuclear compartment-associated Bcl-2 prevents entry of NFκB subunits to the nucleus without affecting NFκB release from its cytosolic inhibitory subunit IκBα. These results suggest that (a) Bcl-2 suppresses transcriptional activity of multiple transcription factors; (b) Bcl-2 does not interfere with NFκB activation but prevents entrance of its active subunits to the nucleus; (c) membrane anchoring is required for this function of Bcl-2; and (d) association of Bcl-2 with the nuclear compartment is also necessary. We speculate that nuclear compartment-associated Bcl-2 may affect nuclear trafficking of multiple factors necessary for transcriptional activity.

AB - Using a reporter gene assay in PC12, HEK293, HeLa, and NIH-3T3 cells, we show that the anti-apoptotic protein Bcl-2 significantly inhibits transcriptional activation of various transcription factors, including NFκB, AP1, CRE, and NFAT. A Bcl-2 mutant lacking its BH4 domain (ΔBH4) also inhibited transcription, whereas a Bcl-2 mutant lacking its transmembrane domain (ΔTM) was ineffective. Furthermore, Bcl-2 cliimeric proteins containing transmembrane domains from the mitochondrial protein monoamine oxidase B (HaoB) or the endoplasmic reticulum protein cytochrome b5 showed no effect on transcription factor activity. Subcellular localization studies showed that under conditions of transient transfection, the active Bcl-2 forms (wild type and ΔBH4) were predominantly found in the nuclear fraction, whereas the non-active forms (ΔTM, MaoB, and cytochrome b5) were in the non-nuclear fraction. Additionally, stably expressed Bcl-2 loses its ability to inhibit transcriptional activation and localizes predominantly to the non-nuclear fraction. Expression of FKBP38 (a chaperone that shuttles Bcl-2 to the mitochondria) removes co-expressed Bcl-2 from the nuclear fraction and reverses its effect on transcription factor activity. Finally, using an indticible gene expression system, we show that nuclear compartment-associated Bcl-2 prevents entry of NFκB subunits to the nucleus without affecting NFκB release from its cytosolic inhibitory subunit IκBα. These results suggest that (a) Bcl-2 suppresses transcriptional activity of multiple transcription factors; (b) Bcl-2 does not interfere with NFκB activation but prevents entrance of its active subunits to the nucleus; (c) membrane anchoring is required for this function of Bcl-2; and (d) association of Bcl-2 with the nuclear compartment is also necessary. We speculate that nuclear compartment-associated Bcl-2 may affect nuclear trafficking of multiple factors necessary for transcriptional activity.

UR - http://www.scopus.com/inward/record.url?scp=11144222923&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=11144222923&partnerID=8YFLogxK

U2 - 10.1074/jbc.M407659200

DO - 10.1074/jbc.M407659200

M3 - Article

VL - 279

SP - 54470

EP - 54478

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 52

ER -