Ets1 Induces Dysplastic Changes When Expressed in Terminally-Differentiating Squamous Epidermal Cells Priyadharsini Nagarajan, Neha Parikh, Lee Ann Garrett-Sinha*, Satrajit Sinha* Department of Biochemistry, State University of New York at Buffalo, Center for Excellence in Bioinformatics and Life Sciences, Buffalo, New York, United States of America Abstract Background: Ets1 is an oncogene that functions as a transcription factor and regulates the activity of many genes potentially important for tumor initiation and progression. Interestingly, the Ets1 oncogene is over-expressed in many human squamous cell cancers and over-expression is highly correlated with invasion and metastasis. Thus, Ets1 is believed to mainly play a role in later stages of the oncogenic process, but not early events. Methodology/Principal Findings: To better define the role of Ets1 in squamous cell carcinogenesis, we generated a transgenic mouse model in which expression of the Ets1 oncogene could be temporally and spatially regulated. Upon Ets1 induction in differentiating cells of stratified squamous epithelium, these mice exhibited dramatic changes in epithelial organization including increased proliferation and blocked terminal differentiation. The phenotype was completely reversed when Ets1 expression was suppressed. In mice where Ets1 expression was re-induced at a later age, the phenotype was more localized and the lesions that developed were more invasive. Many potential Ets1 targets were upregulated in the skin of these mice with the most dramatic being the metalloprotease MMP13, which we demonstrate to be a direct transcriptional target of Ets1. Conclusions/Significance: Collectively, our data reveal that upregulation of Ets1 can be an early event that promotes pre- neoplastic changes in epidermal tissues via its regulation of key genes driving growth and invasion. Thus, the Ets1 oncogene may be important for oncogenic processes in both early and late stages of tumor development. Citation: Nagarajan P, Parikh N, Garrett-Sinha LA, Sinha S (2009) Ets1 Induces Dysplastic Changes When Expressed in Terminally-Differentiating Squamous Epidermal Cells. PLoS ONE 4(1): e4179. doi:10.1371/journal.pone.0004179 Editor: Juha Klefstrom, University of Helsinki, Finland Received June 4, 2008; Accepted November 28, 2008; Published January 14, 2009 Copyright: 2009 Nagarajan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: These studies were supported by grants from the American Cancer Society and the National Institutes of Dental and Craniofacial Research. Neither funder was involved in design or conduct of the study nor in data collection and analysis or manuscript preparation/review/approval. Competing Interests: The authors have declared that no competing interests exist. * E-mail: leesinha@buffalo.edu (LAG-S); ssinha2@buffalo.edu (SS) Introduction The stratified squamous epithelium of the skin forms a barrier between the underlying tissues and the outer milieu to prevent the passage of water and other substances between these compart- ments. Keratinocytes are the principal cell type found in stratified squamous epithelia and generate biomolecules that are necessary for the stability and resistance of the epithelial layer to mechanical stress. The innermost layer of this stratified epithelium, known as the basal layer (stratum basale), consists of a proliferative compartment of undifferentiated cells. Basal cells periodically withdraw from the cell cycle, detach from the basement membrane and migrate outwards to enter the suprabasal compartment. Differentiation of keratinocytes can be monitored by the morphological appearance of the cells and by the expression of particular marker proteins. Based on these criteria, the differentiated layers of the epidermis can be visualized as three separate regions: the spinous or prickle cell layer (stratum spinosum), the granular layer (stratum granulosum) and the cornified layer (stratum corneum). Squamous cell carcinoma (SCC) is a malignant tumor of skin keratinocytes that frequently arises in response to excessive sun exposure or to chronic irritation. Numerous mouse models of squamous cell cancer have been developed including carcinogen- induced SCC, ultraviolet (UV) light-induced SCC and spontane- ous SCC in various transgenic and knockout mouse strains. Several common genetic alterations have been detected in squamous cell tumors, including upregulation of oncogenes and mutation of tumor suppressor genes [1,2]. In addition, several recent studies have reported upregulation of the Ets1 proto- oncogene in human SCC arising in skin [3] and other stratified epithelia [4,5,6,7,8]. Moreover, upregulation of Ets1 expression has also been detected in animal models of oral SCC [9,10]. Expression of high levels of Ets1 is correlated with increased invasiveness and metastatic potential in both human SCC and animal models of SCC. Ets1 is a transcription factor that regulates the expression of many key genes that are involved in cell growth, survival and invasion. Importantly, Ets1 is thought to regulate the expression of proteases such as urokinase plasminogen activator (uPA) and various members of the matrix metalloprotease family (MMP1, MMP2, MMP3, MMP9 and MMP13) [11,12,13,14,15,16]. Ex- pression of these proteases is likely required for tumor cells to degrade the surrounding extracellular matrix, invade nearby tissues and eventually metastasize to distant sites. In addition, to these proteases, Ets1 is also implicated in upregulation of a wide- variety of genes that may promote tumorigenesis including genes that control cellular proliferation or survival (bax, bcl-2, caspase-1, c- PLoS ONE | www.plosone.org 1 January 2009 | Volume 4 | Issue 1 | e4179