IDENTIFICATION OF INVASIVE AND METASTATIC POTENTIAL OF TPL-2 KNOCKOUT KERATINOCYTES

Tpl-2/MAP3K8 is a serine/threonine protein kinase that can activate the transcription of target genes involved in pathways associated with mitosis, cellular differentiation, apoptosis, and inflammation. Our laboratory has previously identified that mice missing the Tpl-2 gene (Tpl-2 knockout mice) are more susceptible to the development of skin cancer--the most prevalent cancer in the United States. However, it is unknown whether the squamous skin cancers that develop in Tpl-2 knockout mice have more potential to become invasive than those skin tumors that form in normal mice. Therefore, in this thesis, we compared the invasive and metastatic potential of skin cells (keratinocytes) from normal and Tpl-2 knockout mice with stimulation and genetic alterations like constitutively-activated Ras. Using various techniques including microarray, real-time polymerase chain reaction, and zymography we found MMP-9, a well-known protease associated with metastatic cells, to be up-regulated at the RNA level. Also, while Tpl-2+/+ keratinocytes migrate faster than Tpl-2-/- in an in vitro scratch assay with H-Ras-infection, Tpl-2-/- keratinocytes have a higher malignant conversion rate and better potential for angiogenic ability when the Ras oncogene is constitutively activated. These experiments provide evidence that Tpl-2-/- keratinocytes, in conjunction with mutant Ras, show potential to become aggressive and invasive.