Procathepsin L is the precursor to the lysosomal cysteine protease cathepsin L. It is secreted in large amounts by malignantly transformed mouse cells, and by cells treated with tumor promoters or growth factors. The function of procathepsin L in malignancy is not known, but involvement in tumor invasion and metastasis and in suppression of the immune response to tumors has been suggested. In normal tissues, procathepsin L is secreted by the liver, osteoclasts, and Sertoli cells, indicating possible involvement in bone resorption and sperm maturation. To study the biological function of procathepsin L, we are attempting to inactivate this gene by insertional mutagenesis into embryonic stem (ES) cells with the goal of establishing transgenic mice lacking cathepsin L function. We have cloned cathepsin L DNA isogenic with ES cell DNA into two targeting vectors (pPNT(s) and pSSC9) which carry different promoters and different numbers of Herpes Simplex Virus (HSV) thymidine kinase (tk) genes. The pSSC9 vector was constructed to allow for several unique restriction sites during cloning and to enhance the positive-negative selection strategy by incorporating two HSV-tk genes. The pPNT based vector which has been used successfully by other labs has been modified to allow for easier linearization. Attempts to inactivate cathepsin L in ES cells are in progress.
