CYSTEINE PROTEASE ACTIVATED BY EXPRESSION OF HIV-1 PROTEASE IN TRANSGENIC MICE - MIP26 (AQUAPORIN-0) CLEAVAGE AND CATARACT FORMATION IN-VIVO AND EX-VIVO

Transgenic mice, homozygous for HIV-1 protease expression in the eye l ens, display degradation of some lens crystallins and cytoskeletal pro teins prior to cataract formation on postnatal days 23-25, Alterations to the internal lens hydration state also occur; therefore, the statu s of the aquaporin protein MIP26 was examined over postnatal days 16-2 5 to determine if it was altered during cataractogenesis. The MTP was identical in transgenic and control lenses until day 21. By postnatal day 25 (frank cataract), in the lenses obtained from transgenic animal s, the 26-kDa band was absent and there was a concurrent increase in t he proportion of MIP23, Immunoblotting demonstrated cleavage at the C terminus, Lenses were also maintained in an organ culture system to de monstrate that the cataractogenic process is inherent to the isolated lens and to determine the contribution of cysteine protease action, Or gan culture experiments revealed a similar progression to nuclear cata ract formation as seen in vivo. Two-dimensional gel analysis of the so luble lens crystallin fraction of organ cultured lenses revealed the s ame cleavage pattern as occurs in vivo, Organ culture of transgenic le nses with E64, a cysteine protease inhibitor, dramatically delayed cat aractogenesis and prevented proteolytic cleavage of both MIP26 and cry stallins, HIV-1 protease, while the trigger of cataract formation, doe s not appear to be the protease responsible for cleavage of MIP or len s crystallins. These results suggest that activation of endogenous cys teine protease activity is involved in the cleavage of these proteins and occurs downstream of HIV-1 protease action.