The mouse model of experimental skin carcinogenesis has long proposed that epidermal stem cells are a major target of carcinogens and form the latent neoplastic population that ultimately expands into tumors. The hematopoeitic stem and progenitor cell marker, CD34, is uniquely expressed on hair follicle stem cells in mouse skin, and as a cell surface marker, facilitates isolation of this population using Fluorescence Activated Cell Sorting (FACS) of single cell suspensions of keratinocytes. Our previous analysis of stem cells isolated from untreated wild type mice yielded a gene list of about 2000 differentially expressed genes in hair follicle stem cells relative to non-stem basal keratinocytes. The T-box transcription factor, Tbx1, was found to be highly expressed in CD34+ stem cells, with about a 20-fold enrichment in the stem cell population. This gene is a developmentally important gene, essential for cardiac and inner ear development. Tbx1 is also lost in the human chromosome 22 deletion syndrome (Velocardiofacial syndrome and DiGeorge Syndrome), characterized by craniofacial defects and cardiac abnormalities. In addition, Tbx1 has been shown to be expressed in embryonic skin in the emerging hair follicle placode with progressive localization to the outer root sheath. Because Tbx1 regulates cardiac progenitor cell proliferation and is involved in cell fate determination, we were interested if there was a role for this gene in skin tumor development, given the high level of expression in the adult skin stem cell population. To test this, we investigated the effect of ectopic expression of Tbx1 in a mouse spindle cell tumor cell line. When transfected with mouse Tbx1, these cells exhibited reduced growth in culture, loss of anchorage-independent growth, and delayed tumor growth in mice following intradermal injection of transfected cells. These data provide a novel role for Tbx1 as a negative regulator of tumor growth through restoration of contact inhibition (manuscript in preparation: Ectopic expression of the T-box transcription factor, Tbx1, suppresses skin tumor growth through restoration of contact inhibition). In addition, we have data suggesting that Tbx1 may play a unique role in DNA repair through interaction with p53 and Mre11, potentially expanding the role of Tbx1 in skin tumorigenesis. Pilot studies utilizing microarray analysis of CD34-expressing stem cells from acetone- and TPA-treated wild type (WT) and v-Ha-ras transgenic mice has shown that this approach is sensitive enough to discriminate differential responses between mutant and normal hair follicle stem cells isolated from mice exposed to tumor promoting chemicals. Current annotation suggests that mutant (i.e., mutant ras-expressing) stem cells express genes associated with anti-apoptotic pathways, providing a cell survival mechanism in pre-neoplastic stem cells that would facilitate expansion into skin tumors. We are also pursuing a proteomics approach to understanding key signaling pathways affected in initiated stem cells, using the chemical carcinogen exposure of wild type mice, followed by tumor promotion. Our microarray analysis revealed that the cell proliferation-associated gene, Stathmin 1 (Stmn1), was uniquely up-regulated in stem cells harboring mutant ras (TPA-treated mice). We have subsequently demonstrated that there is a high level of expression of Stmn1 in actively growing hair follicle localized to the hair matrix cells which is lost as hair follicles regress into the resting phase. Therefore, Stmn1 may be a novel marker of hair follicle growth in both mouse and human skin. We have also shown Stmn1 expression in basal and squamous cell carcinomas (BCC and SCC, respectively) of human origin, and SCC of mouse origin, which has not been described before, and may be an indicator of a potential therapeutic target in the treatment of skin disease. We have obtained Stmn1 KO mice and will be using this model to investigate the role of stathmin in hair follicle cycling and skin tumorigenesis. Taken together, our current studies demonstrate that mouse hair follicle stem cells can be isolated from mice following chemical exposure and can be used in subsequent gene and protein assays to provide unique insight into stem cell response to environmental exposures, which may lead to novel targets for intervention and prevention of skin cancers in humans.
| Madson, Justin G; Lynch, David T; Svoboda, Jessica et al. (2009) Erbb2 suppresses DNA damage-induced checkpoint activation and UV-induced mouse skin tumorigenesis. Am J Pathol 174:2357-66 |
| Wei, Sung-Jen; Williams, Jason G; Dang, Hong et al. (2008) Identification of a specific motif of the DSS1 protein required for proteasome interaction and p53 protein degradation. J Mol Biol 383:693-712 |
| Garantziotis, Stavros; Zudaire, Enrique; Trempus, Carol S et al. (2008) Serum inter-alpha-trypsin inhibitor and matrix hyaluronan promote angiogenesis in fibrotic lung injury. Am J Respir Crit Care Med 178:939-47 |
