예쁜꼬마선충을 이용한칼시규린 조절인자(RCAN-1)와 티오레독신 환원효소(TRXR-1 and TRXR-2)의 유전적 연구

Abstract

Calcineurin is a calcium-dependent serine-threonine phosphatase. In mammalian calcineurin activates T cell by dephosphorylate NFAT. Overexpression of RCAN-1, the only ortholog in C. elegans, resulted in calcineurin loss-of-function like phenotypes, such as small body size, cuticle defects, small brood size, slow growth and resistance to serotonin mediated egg laying. This thesis mainly use genetic method and behavior analysis to study how rcan-1 is involved in regulate thermotaxis. The rcan-1 mutants were cryophilic. The RCAN-1 regulate thermotaxis by inhibiting calcineurin function. By yeast two hybrid system, the PxIxIT calcineurin motif was conserved in RCAN-1. The PxIxIT motif was important to rescue the thermotaxis behavior. My result also showed that CREB (cAMP response element-binding protein) pathway was also involve in calcineurin pathway to regulate thermotaxis. The two phosphorylation sites in RCAN-1 was found to be important for RCAN-1 full inhibitory activity. It was previously reported that RCAN1 -/- showed resistant to oxidative stress. Both rcan-1 overexpression and oxidative stress was found to aggravate degenerative neuronal disease such as Alzheimer’s disease. Thioredoxin reductase (TRXR) exists in two major isoenzymes in higher eukaryotic cells; cytosolic trxr-1 and mitochondrial trxr-2. In mammals, trxr-2 protects mitochondria from oxidative stress, where reactive oxidative species are mainly generated, while trxr-1 plays a role to maintain optimal oxido-reductive status in cytosol. In this study, I report differential physiological functions of these two TRXRs in C. elegans. trxr-1 is highly expressed in pharynx, vulva and intestine, whereas trxr-2 is mainly expressed in pharyngeal and body wall muscles where it is localized in mitochondria. Deficiency of trxr-2 caused defects in longevity and delayed development, while deletion mutation of trxr-1resulted in interference in acidification of lysosomal compartment in intestine. Both trxr-1 and trxr-2 were up-regulated when worms were challenged by heat shock stress, but neither paraquat treatment nor daf-2 mutant background. These results suggest that trxr-1 and trxr-2 function differently at organismal level presumably by their differential sub-cellular localization in C. elegans.