ErbB Receptor Cross-talk and Co-localization in Mouse Lung Epithelial Cells

K. Zscheppang; E. Korenbaum; S. M. Ramadurai; H. C. Nielsen; C. E. L. Dammann

doi:10.1055/s-2005-862730

Pneumologie, Table of Contents

ErbB Receptor Cross-talk and Co-localization in Mouse Lung Epithelial Cells

K Zscheppang ¹^,², E Korenbaum ³, SM Ramadurai ⁴, HC Nielsen ⁴, CEL Dammann ¹^,⁴

¹Department of Pediatric Pulmonology and Neonatology, Hannover Medical School, Hannover, Germany
²University of Applied Sciences Lausitz, Senftenberg, Germany
³Department of Biophysical Chemistry, Hannover Medical School, Hannover, Germany and
⁴Department of Pediatrics, Tuft's New England Medical Center, Boston, MA

Abstract

Background: ErbB receptors, critically important for embryonic neuronal, cardiac, and lung development, are expressed in late gestation lung where they play a major role regulating the onset of surfactant synthesis (AJRCCM 167:1711 – 16, 2003). ErbB receptor ligands neuregulin (NRG) and epidermal growth factor (EGF) initiate this process. We previously showed that all four erbB receptors are expressed in both fetal lung type II cells and fibroblasts, and differ in phosphorylation and cellular response after ligand stimulation. Different erbB receptor ligands can cause diverse biologic results by stimulating specific erbB-dimers. It is not known how dimerization, cellular localization, and co-localization of dimers are regulated in fetal type II cells.

Objective: We hypothesized that different ligands initiate different erbB receptor dimerization, localization and co-localization patterns in mouse epithelial type II cells.

Design/Methods: We used MLE-12 cells, a mouse alveolar type II cell line, as a model. Cells were cultured either in culture dishes or on glass cover slips. The cells were stimulated for 2 min with EGF (100ng/ml) or NRG (33 nM). Co-immunoprecipitation (Co-IP) was prepared for each erbB receptor. For fluorescent microscopy, cells were incubated with erbB antibodies, followed by fluorescent-tagged 2^o antibody. Receptor localization and co-localization were determined by confocal microscopy.

Results: Co-IP resulted in dimerization patterns that preferentially included erbB4. ErbB4 was the only receptor that showed a mixed pattern of autophosphorylation and phosphorylation in response to stimulation. Confocal microscopy studies showed a predominant localization of erbB1 in the cytoplasm, while erbB2 and ErbB3 almost exclusively localized to the nucleus. In contrast, erbB4 again exhibited an mixed pattern, between cytoplasm and nucleus. EGF and NRG stimulated a more diffuse staining of erbB1 and 4 in the cytoplasm.

Conclusions: Among all four erbB receptors expressed by fetal lung epithelial cells, erbB4 appears to play a unique role. First, erbB4 might be the universal dimerization partner. Second, erbB4 might be rather mobile, moving between cell membrane and nucleus. Due to these unique capabilities, erbB4 might play a major role in the signalling of fetal surfactant synthesis. (Funded by NIH, Charles Hood and Peabody Fountdation, and HiLF).