Bladder epithelium or urothelium is involved in major urinary tract diseases including bladder cancer, urinary tract infection and interstitial cystitis. However, research into the molecular bases of urothelial diseases has been hampered by the lack of effective model systems. Utilizing uroplakin II (UPII) gene promoter, we have succeeded over the last several years in targeting gene expression specifically into the urothelium of transgenic mice. The existing system, however, does not allow gene expression and inactivation to occur in a temporally controlled fashion, thus precluding many biological questions from being addressed. The overall goals of the current project are to develop and validate the second-generation transgenic systems that will allow urothelium-specific and inducible gene expression as well as knockout;and to utilize these novel systems to study the in vivo roles of several critical genes in urothelial growth, differentiation and tumorigenesis. Toward these goals, we plan to carry out two series of studies. In the first, we will generate transgenic lines in which the UPII promoter drives the urothelium-specific expression of a reverse tetracycline transactivator (UPll/rtTA). We will then test the feasibility and parameters of urothelium-specific and inducible gene expression by crossing UPll/rtTA mice with TRE/LacZ reporter mice so that LacZ expression is under the control of tetracycline response elements (TRE) and only occurs upon doxycycline treatment. In addition, we will establish a urothelium-specific and inducible knockout system, by crossing the UPll/rtTA mice with TRE/Cre mice. We will then test the functionality of this system by crossing the UPll/rtTA-TRE/Cre bi-transgenic mice with Rosa26 reporter mice in which the reporter genes are transcriptionally activated upon Cre expression. This system will allow inducible knockout of any genes of interest in the urothelium. In the second series of studies, we will inducibly express an activated fibroblast growth factor receptor as well as inducibly delete the retinoblastoma gene in the urothelium and test the hypothesis that the two genetic alterations induce low-grade superficial papillary tumors and high-grade invasive tumors, respectively. We will also examine the signaling pathways that these genetic alterations exploit in transforming the urothelium. These studies will generate powerful experimental tools for studying bladder biology and diseases and offer molecular insights regarding urothelial growth and tumorigenesis.
