The role of ERBB2 and ERBB3 receptors in skin development, homeostasis, and tumor susceptibility

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The epidermal growth factor receptor (EGFR/ERBB1) and its ligands are established regulators of skin and hair follicle homeostasis, and their deregulation rapidly results in pathological alterations. In contrast - although they are also expressed in the skin and have been implicated in skin tumorigenesis - the functions exerted by the related tyrosine kinase receptors ERBB2 and ERBB3 in this tissue remain largely unexplored.We propose to generate mouse lines lacking ERBB2 or ERBB3 specifically in the skin. In particular, we propose to cross mice carrying floxed alleles of the Erbb2 or Erbb3 genes with mice expressing cre recombinase under the control of the bovine keratin 5 promoter (K5cre). Resulting K5cre-Erbb2fl/fl (= K5-Erbb2del) and K5cre-Erbb3fl/fl (= K5-Erbb3del) mice will have an early deletion (starting around embryonic day e15.5) of the corresponding receptors restricted to keratinocytes of the hair follicle and interfollicular epidermis, allowing us to evaluate the role of ERBB2 for skin development and homeostasis. We will evaluate the skin histology (H&E, proliferation, apoptosis), the epidermal differentiation by localizing several differentiation markers, and the kinetics of hair follicle morphogenesis and cycle induction.We also propose to create mice with skin-specific loss of ERBB2 or ERBB3 by employing the K14creERT2 line and tamoxifen treatment and subject them to a two-stage chemical carcinogenesis protocol. In this protocol, a single application of the initiator DMBA is followed by repeated applications of the pro-inflammatory agent TPA, resulting in benign papillomas that regress or progress to squamous cell carcinoma. The number and the size of the tumors will be recorded weekly and, at the end of the experiment (~ 20 weeks after DMBA application), tumors will be fixed for histological analysis or frozen for expression analysis. Thus, the two-stage chemical carcinogenesis provides an opportunity to definitively clarify the influence of ERBB2 and ERBB3 in early and late events in skin cancer development and progression. Finally, several studies showed that ERBB2 is activated by UV irradiation and increases UV-induced skin tumorigenesis. Unfortunately, the employed models did not allow definitive conclusions about the role of ERBB2 in this process. Therefore, we propose to expose ERBB2-deficient mice and control littermates to UVA or UVB irradiation. Twenty-four hours later, we will evaluate parameters such as the epidermal thickness, the epidermal cell proliferation and apoptosis, and the number of sunburn cells.Finally, to allow mechanistic studies, we propose the establishment of permanent keratinocyte cell lines lacking ERBB2 or ERBB3 as well as control lines. With these cell lines, we will evaluate the formation of EGFR/ERBB dimers and the downstream signaling pathways, cell proliferation, adhesion, and anchorage-independent growth, as well as the interaction between ERBB2 and ERBB3 with E-cadherin and LRIG1.