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The ERBB4/HER4 receptor tyrosine kinase regulates gene expression by functioning as a STAT5A nuclear chaperone.

Williams CC, Allison JG, Vidal GA, Burow ME, Beckman BS, Marrero L, Jones FE - J. Cell Biol. (2004)

Bottom Line: We have identified an intrinsic ERBB4 NLS (residues 676-684) within the ERBB4 intracellular domain (4ICD) that is essential for nuclear accumulation of 4ICD.Together, our results establish a novel molecular mechanism of transmembrane receptor signal transduction involving nuclear cotranslocation of the receptor intracellular domain and associated transcription factor.Subsequent binding of the two proteins at transcription factor target promoters results in activation of gene expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Tulane University Health Sciences Center, Tulane Cancer Center, New Orleans, LA 70112, USA.

ABSTRACT
In the lactating breast, ERBB4 localizes to the nuclei of secretory epithelium while regulating activities of the signal transducer and activator of transcription (STAT) 5A transcription factor essential for milk-gene expression. We have identified an intrinsic ERBB4 NLS (residues 676-684) within the ERBB4 intracellular domain (4ICD) that is essential for nuclear accumulation of 4ICD. To determine the functional significance of 4ICD nuclear translocation in a physiologically relevant system, we have demonstrated that cotransfection of ERBB4 and STAT5A in a human breast cancer cell line stimulates beta-casein promoter activity. Significantly, nuclear localization of STAT5A and subsequent stimulation of the beta-casein promoter requires nuclear translocation of 4ICD. Moreover, 4ICD and STAT5A colocalize within nuclei of heregulin beta 1 (HRG)-stimulated cells and both proteins bind to the endogenous beta-casein promoter in T47D breast cancer cells. Together, our results establish a novel molecular mechanism of transmembrane receptor signal transduction involving nuclear cotranslocation of the receptor intracellular domain and associated transcription factor. Subsequent binding of the two proteins at transcription factor target promoters results in activation of gene expression.

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Related in: MedlinePlus

Activated ERBB4 mediates nuclear translocation of coexpressed STAT5A. MCF-7B cells were cotransfected with (A and B) Red-STAT5A and ERBB4-EGFP or (C and D) Red-STAT5A and ERBB4muNLS-EGFP. Immediately before fixation in 4% PFA at 48 h after transfection, cells were mock stimulated (Mock) or stimulated with 50 ng/ml of HRG for 30 min at RT (HRG). After fixation, cells were stained with Hoechst dye and observed by deconvolution microscopy. Cytoplasmic and nuclear colocalization of ERBB4 (green) and STAT5A (red) of 10 cotransfected cells in the merged image (Merge w/Hoechst) was measured using a pixel correlation algorithm (B and D). Bar, 5 μm.
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fig5: Activated ERBB4 mediates nuclear translocation of coexpressed STAT5A. MCF-7B cells were cotransfected with (A and B) Red-STAT5A and ERBB4-EGFP or (C and D) Red-STAT5A and ERBB4muNLS-EGFP. Immediately before fixation in 4% PFA at 48 h after transfection, cells were mock stimulated (Mock) or stimulated with 50 ng/ml of HRG for 30 min at RT (HRG). After fixation, cells were stained with Hoechst dye and observed by deconvolution microscopy. Cytoplasmic and nuclear colocalization of ERBB4 (green) and STAT5A (red) of 10 cotransfected cells in the merged image (Merge w/Hoechst) was measured using a pixel correlation algorithm (B and D). Bar, 5 μm.

Mentions: Deconvolution microscopy revealed ERBB4 accumulation at the cell membrane of mock-treated cells with a high level of cytoplasmic STAT5A colocalization, probably representing STAT5A associated with constitutively activated ERBB4 protein (Fig. 5 A, top, and Fig. 5 B). When cotransfected cells were stimulated with HRG, pronounced nuclear accumulation and dramatic nuclear colocalization of 4ICD and STAT5A was observed (Fig. 5 A, bottom, and Fig. 5 B). The punctate pattern of 4ICD/STAT5A nuclear colocalization resembles sub-nuclear structures referred to as nuclear speckles. The exact function of nuclear speckles remains controversial but appear to be sites of splicing machinery assembly associated with highly active transcription (Lamond and Spector, 2003).


The ERBB4/HER4 receptor tyrosine kinase regulates gene expression by functioning as a STAT5A nuclear chaperone.

Williams CC, Allison JG, Vidal GA, Burow ME, Beckman BS, Marrero L, Jones FE - J. Cell Biol. (2004)

Activated ERBB4 mediates nuclear translocation of coexpressed STAT5A. MCF-7B cells were cotransfected with (A and B) Red-STAT5A and ERBB4-EGFP or (C and D) Red-STAT5A and ERBB4muNLS-EGFP. Immediately before fixation in 4% PFA at 48 h after transfection, cells were mock stimulated (Mock) or stimulated with 50 ng/ml of HRG for 30 min at RT (HRG). After fixation, cells were stained with Hoechst dye and observed by deconvolution microscopy. Cytoplasmic and nuclear colocalization of ERBB4 (green) and STAT5A (red) of 10 cotransfected cells in the merged image (Merge w/Hoechst) was measured using a pixel correlation algorithm (B and D). Bar, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172499&req=5

fig5: Activated ERBB4 mediates nuclear translocation of coexpressed STAT5A. MCF-7B cells were cotransfected with (A and B) Red-STAT5A and ERBB4-EGFP or (C and D) Red-STAT5A and ERBB4muNLS-EGFP. Immediately before fixation in 4% PFA at 48 h after transfection, cells were mock stimulated (Mock) or stimulated with 50 ng/ml of HRG for 30 min at RT (HRG). After fixation, cells were stained with Hoechst dye and observed by deconvolution microscopy. Cytoplasmic and nuclear colocalization of ERBB4 (green) and STAT5A (red) of 10 cotransfected cells in the merged image (Merge w/Hoechst) was measured using a pixel correlation algorithm (B and D). Bar, 5 μm.
Mentions: Deconvolution microscopy revealed ERBB4 accumulation at the cell membrane of mock-treated cells with a high level of cytoplasmic STAT5A colocalization, probably representing STAT5A associated with constitutively activated ERBB4 protein (Fig. 5 A, top, and Fig. 5 B). When cotransfected cells were stimulated with HRG, pronounced nuclear accumulation and dramatic nuclear colocalization of 4ICD and STAT5A was observed (Fig. 5 A, bottom, and Fig. 5 B). The punctate pattern of 4ICD/STAT5A nuclear colocalization resembles sub-nuclear structures referred to as nuclear speckles. The exact function of nuclear speckles remains controversial but appear to be sites of splicing machinery assembly associated with highly active transcription (Lamond and Spector, 2003).

Bottom Line: We have identified an intrinsic ERBB4 NLS (residues 676-684) within the ERBB4 intracellular domain (4ICD) that is essential for nuclear accumulation of 4ICD.Together, our results establish a novel molecular mechanism of transmembrane receptor signal transduction involving nuclear cotranslocation of the receptor intracellular domain and associated transcription factor.Subsequent binding of the two proteins at transcription factor target promoters results in activation of gene expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Tulane University Health Sciences Center, Tulane Cancer Center, New Orleans, LA 70112, USA.

ABSTRACT
In the lactating breast, ERBB4 localizes to the nuclei of secretory epithelium while regulating activities of the signal transducer and activator of transcription (STAT) 5A transcription factor essential for milk-gene expression. We have identified an intrinsic ERBB4 NLS (residues 676-684) within the ERBB4 intracellular domain (4ICD) that is essential for nuclear accumulation of 4ICD. To determine the functional significance of 4ICD nuclear translocation in a physiologically relevant system, we have demonstrated that cotransfection of ERBB4 and STAT5A in a human breast cancer cell line stimulates beta-casein promoter activity. Significantly, nuclear localization of STAT5A and subsequent stimulation of the beta-casein promoter requires nuclear translocation of 4ICD. Moreover, 4ICD and STAT5A colocalize within nuclei of heregulin beta 1 (HRG)-stimulated cells and both proteins bind to the endogenous beta-casein promoter in T47D breast cancer cells. Together, our results establish a novel molecular mechanism of transmembrane receptor signal transduction involving nuclear cotranslocation of the receptor intracellular domain and associated transcription factor. Subsequent binding of the two proteins at transcription factor target promoters results in activation of gene expression.

Show MeSH
Related in: MedlinePlus