We use mouse skin epidermis and epidermal squamous cell carcinoma (SCC) as models of tissue growth in development and disease. Skin epidermis is a particularly suitable system for our investigation for the following reasons:
Well-defined physiological system: Skin is composed of two compartments, the epithelial epidermis and mesenchymal dermis, separated by a basement membrane. Epidermal growth is achieved through continuous cycles of progenitor cell self-renewal and differentiation under the control of cell extrinsic signals coming from the surrounding mesenchyme.
Implications for human health: SCC of the skin is the second most common cancer in people, with an estimated 700,000 new cases in the US each year. Fortunately, most lesions are detected early and surgically removed, accounting for disease’s high survival rates. Importantly, ontogeny of epidermal SCC parallels cancers associated with high mortality, including the SCC of the head and neck and the lung SCC.
Tools for analysis of gene function: In addition to standard methods of creating transgenic animals, mouse epidermis can be efficiently and stably targeted through in utero injection of lentivirus. Using lentivirus to express shRNA, for RNAi-mediated gene knockdown, or ORF, for gene overexpression, we can rapidly assess gene function and complex genetic interactions in skin morphogenesis and disease in vivo.
Epidermal progenitors (K5) at the epithelial-mesenchymal interface (Nido) fuel the establishment of the differentiated layers (K10)
In utero injection of GFP-expressing lentivirus results
in uniform transduction of the epidermis
Candidate-based analysis of gene function in regulation of epidermal tissue growth.
Following completion of in vivo RNAi screens of ~16,000 mouse genes for regulators of epidermal tissue growth during embryonic development and oncogenic hyperplasia, we have started the exciting phase of validating individual candidates. We are also probing the precise cellular and molecular mechanisms behind the observed growth effects, with a focus on novel genes that specifically operate within the physiological environment by altering the balance between stem cell renewal and differentiation.
Large-scale investigation of modifiers of epidermal tumor initiation.
We have successfully combined the pooled-format lentiviral-mediated RNAi and quantitative Illumina sequencing into a rapid, comprehensive, and relatively low-cost approach to genome-wide gene function analysis during development. We are currently extending the use of this approach to adult epidermis, with a goal of identifying bona fide enhancers of tumor initiation and progression. The complexity of our lentiviral pools vary from patient-specific to genome-wide.
Development of a general model of epithelial growth and tumorigenesis.
Small modifications in the in utero injection method result in efficient lentiviral transduction of other tissues, including the oral, mammary and airway epithelium. These are distinct from the epidermis in their organization, physiological environment, and rate of developmental and regenerative growth, and carcinomas in these epithelia are the leading cause of tumor-associated deaths worldwide. Using RNAi-mediated gene knockdown we are testing general applicability of molecular mechanisms uncovered in our studies of epidermal growth and tumorigenesis.