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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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Identification of a minimal ERBB4 sequence that enhances EGFP nuclear accumulation. (A) ERBB4 residues representing putative NLSs fused to the amino terminus of EGFP. (B–E) Localization by deconvolution microscopy of indicated NLS-EGFP constructs 48 h after transfection of HEK 293T cells. (F) Basic residues (red) altered in NLS1-EGFP fusions. (G–J) Localization by deconvolution microscopy of indicated NLS1-EGFP mutants 48 h after transfection of HEK 293T cells.
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fig1: Identification of a minimal ERBB4 sequence that enhances EGFP nuclear accumulation. (A) ERBB4 residues representing putative NLSs fused to the amino terminus of EGFP. (B–E) Localization by deconvolution microscopy of indicated NLS-EGFP constructs 48 h after transfection of HEK 293T cells. (F) Basic residues (red) altered in NLS1-EGFP fusions. (G–J) Localization by deconvolution microscopy of indicated NLS1-EGFP mutants 48 h after transfection of HEK 293T cells.

Mentions: As an experimental approach toward the identification of an intrinsic ERBB4 NLS, we tested the ability of three independent ERBB4 basic residue stretches positioned within 4ICD (Fig. 1 A; Srinivasan et al., 2000) to enhance nuclear translocation of EGFP. When transiently transfected into HEK 293T cells the EGFP (Fig. 1 B) vector control and amino-terminal EGFP fusions of NLS2 (Fig. 1 D) and NLS3 (Fig. 1 E) displayed identical patterns of diffuse cytoplasmic localization accompanied by basal levels of nuclear accumulation. An NLS1-EGFP fusion (Fig. 1 C) on the other hand exhibited distinct nuclear accumulations within subnuclear structures resembling nucleoli. Although these results do not exclude a role for NLS2 and NLS3 in nuclear translocation, consistent with our results, a putative juxtamembrane EGFR NLS, with basic residue clusters similar to the ERBB4 NLS1, was shown to mediate nuclear translocation of β-galactosidase (Lin et al., 2001).


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)

Identification of a minimal ERBB4 sequence that enhances EGFP nuclear accumulation. (A) ERBB4 residues representing putative NLSs fused to the amino terminus of EGFP. (B–E) Localization by deconvolution microscopy of indicated NLS-EGFP constructs 48 h after transfection of HEK 293T cells. (F) Basic residues (red) altered in NLS1-EGFP fusions. (G–J) Localization by deconvolution microscopy of indicated NLS1-EGFP mutants 48 h after transfection of HEK 293T cells.
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Related In: Results  -  Collection

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

fig1: Identification of a minimal ERBB4 sequence that enhances EGFP nuclear accumulation. (A) ERBB4 residues representing putative NLSs fused to the amino terminus of EGFP. (B–E) Localization by deconvolution microscopy of indicated NLS-EGFP constructs 48 h after transfection of HEK 293T cells. (F) Basic residues (red) altered in NLS1-EGFP fusions. (G–J) Localization by deconvolution microscopy of indicated NLS1-EGFP mutants 48 h after transfection of HEK 293T cells.
Mentions: As an experimental approach toward the identification of an intrinsic ERBB4 NLS, we tested the ability of three independent ERBB4 basic residue stretches positioned within 4ICD (Fig. 1 A; Srinivasan et al., 2000) to enhance nuclear translocation of EGFP. When transiently transfected into HEK 293T cells the EGFP (Fig. 1 B) vector control and amino-terminal EGFP fusions of NLS2 (Fig. 1 D) and NLS3 (Fig. 1 E) displayed identical patterns of diffuse cytoplasmic localization accompanied by basal levels of nuclear accumulation. An NLS1-EGFP fusion (Fig. 1 C) on the other hand exhibited distinct nuclear accumulations within subnuclear structures resembling nucleoli. Although these results do not exclude a role for NLS2 and NLS3 in nuclear translocation, consistent with our results, a putative juxtamembrane EGFR NLS, with basic residue clusters similar to the ERBB4 NLS1, was shown to mediate nuclear translocation of β-galactosidase (Lin et al., 2001).

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