PUB - Publikationen an der Universität Bielefeld

It has been known, MT1-, 2-, 3-, 5-MMP have a transmembrane (TM) domain and MT4-, 6-MMP contain a GPI anchor, both of which are supposed to localize these proteins on cell surface. However, a truncated MT1-MMP that is deleted its TM domain still retains on the surface of transfected COS cells. MMP-2, particularly active MMP-2, has been regarded as a central player for angiogenesis, cellular tumor invasion as well as metastases in both animal model and clinic samples. One of important functions for MT-MMPs is to process or activate pro-MMP-2 into an active form; however, controversial results were reported on the ability of MT2-, -3, -4, -6-MMP to process the pro-MMP-2. In here, we reported that human MT1-MMP and mouse MT3-MMP were found to localize on the plasma membrane of transfected COS cells; in contrast, human MT2-MMP was not detected on transfected COS cell surface. Human MT1-MMP is soluble in Triton X-100 buffer, but human MT2-MMP and mouse MT3-MMP displayed insolubility. Mouse MT3-MMP and human MT2-MMP did not efficiently process the activation of pro-MMP-2 in COS cells. Through a series of chimeric construction of MT2-MMP or MT3-MMP, localization and activity of those chimeric proteins were investigated by immunofluorescence microscopy and gelatine zymography. The failure of human MT2-MMP to activate pro-MMP-2 is due to its poor localization at COS cell surface. In this event, the cytoplasmic tail of human MT2-MMP plays an important role since the substitution of its cytoplasmic and transmembrane domain with the corresponding part of FcRIIa receptor not only localized human MT2-MMP ectodomains on the cell plasma membrane but also showed the activity to process pro-MMP-2. The cytoplasmic tail of mouse MT3-MMP was able to localize either MT1-MMP or MT3-MMP ectodomains onto the cell surface and interestingly it only facilitated MT1-MMP but not MT3-MMP ectodomains to ecert their activities to process pro-MMP-2. This suggests that there is a different mechanism for mouse MT3-MMP to process the pro-MMP-2 low efficiently compared to human MT2-MMP. Stable double transgenic mice were established via crossing response transgenic mice carrying full-length mouse MT3-MMP with regulatory transgenic mice containing reverse tetracycline regulatory transactivator (rtTA). Tg-MT3-MMP was found in most of the studied organs such as heart, kidney, muscle, tongue and occasionally in brain in CR4 double transgenic mice under induction condition. The expression of the transgenes (MT3-MMP and Lac Z) displayed a Dox-HCl dose and time-dependent type. CRx3001 displayed a faster and more efficient response to doxycycline than other mouse lines did. More active-form MT3-MMP proteins were found in the kidney of CR4x3001 double transgenic mice treated by doxycycline compared to the negative control mice. In general, there were no significant differences among wild type, single and double transgenic mice at the later stages of development. Tumor that was occasionally found in transgenic mice seem not due to the overexpression of transgenic MT3-MMP.