신경계 칼시뉴린에 의한 성장조절에 관여하는 유전자 연구

Abstract

칼시뉴린은 칼슘/칼모듈린에 의존하는 세린/스레오닌 탈인산화효소이며, 예쁜꼬마선충에서 몸 길이를 조절하는데 기능이 있다고 알려 져 있다. 하지만 어떻게 칼시뉴린이 성장을 조절하는지는 명확하게 밝혀 져 있지 않다. 이 연구에서는 몸 길이의 조절하는데 있어서 칼시뉴린의 신경에서의 역할을 연구하였다. 칼시뉴린과 상호작용을 하는 인산화효소를 찾기 위해 RNAi 스크리닝을 수행하였고 34개의 인산화효소가 킬시뉴린 돌연변이의 작은 몸길이를 되돌렸다. Positive 후보군 중에서 Dual-specificity Yak1-related Kianse(DYRK) 패밀리 단백질인 MBK-2를 찾았다. mbk-2의 타겟 유전자중 하나인 oma-1과 oma-2 또한 칼시뉴린 돌연변이의 작은 몸길이를 정상의 수준으로 회복시켰다. 이는 칼시뉴린은 MBK-2와 함께 OMA-1 전사요소를 통해 몸 길이가 조절된다고 할 수 있다. 칼시뉴린 돌연변이의 작은 몸 길이가 신경전달물질에 의해 영향을 받는지 알아보기 위해 다양한 신경전달물질을 예쁜꼬마선충에 처리를 하였다. 흥미롭게도 아세틸콜린 만이 칼시뉴린 돌연변이의 몸길이를 길게 변하게 하였지만 다른 신경전달물질들은 영향이 없었다. 또한 칼시뉴린의 cholinergic 신경에 발현시켰을 경우, 칼시뉴린 돌연변이의 몸길이가 완전히 회복되었다. 이 결과들은 칼시뉴린은 아세틸콜린의 분비를 조절하여 몸 길이 사이즈를 조절한다고 할 수 있다. 칼시뉴린 돌연변이에서는 몸 길이를 조절하는 잘 알려져 있는 TGF-β 신호경로가 억제되는 것을 확인하였다. 따라서 칼시뉴린은 선택적으로 아세틸콜린의 분비를 조절하여 TGF-β 신호경로에 영향을 미쳐 개체의 성장 및 몸 길이를 조절한다고 할 수 있다.|Calcineurin, Ca2+/calmodulin-dependent serine/threonine phosphatase, has been reported to function in body size regulation in C. elegans. However, how calcineurin regulates growth is largely unknown. In this study, I studied the neuronal function of calcineurin in the regulation of body size. RNAi screening to identify kinases which counteract with calcineurin activity revealed 34 kinases that rescued the phenotype of small body size in calcineurin tax-6(lf) background. Among the positive candidates, I found that mbk-2, ortholog of the Dual-specificity Yak1-Related Kinase (DYRK) family proteins partially rescued the body size of tax-6(lf). One of the target genes for mbk-2, oma-1 and oma-2 which encode a zinc finger proteins and both are paralogous to each other, reversed small body length of tax-6(lf) as well as mbk-2 significantly suggesting that calcineurin may work with MBK-2 through OMA-1. To see which neurotransmitter is affected in small body size of tax-6 calcineurin loss-of-function mutant, I treated worms with various neurotransmitters. Interestingly exogenous acetylcholine reversed small the body size in tax-6 loss-of-function mutants, while other tested neurotransmitters did not affect the body growth. In addition, cholinergic neuronal expression of tax-6 also recovered small body length of tax-6 mutants. These results suggest that calcineurin modulates acetylcholine release to control worm growth. In addition, loss-of-function mutation in calcineurin tax-6 suppresses TGF-β pathway, which leads to reduced body size. Therefore, calcineurin pathway may regulate organismal growth by selectively controlling acetylcholine neurotransmission that affects TGF-β signaling.; Calcineurin, Ca2+/calmodulin-dependent serine/threonine phosphatase, has been reported to function in body size regulation in C. elegans. However, how calcineurin regulates growth is largely unknown. In this study, I studied the neuronal function of calcineurin in the regulation of body size. RNAi screening to identify kinases which counteract with calcineurin activity revealed 34 kinases that rescued the phenotype of small body size in calcineurin tax-6(lf) background. Among the positive candidates, I found that mbk-2, ortholog of the Dual-specificity Yak1-Related Kinase (DYRK) family proteins partially rescued the body size of tax-6(lf). One of the target genes for mbk-2, oma-1 and oma-2 which encode a zinc finger proteins and both are paralogous to each other, reversed small body length of tax-6(lf) as well as mbk-2 significantly suggesting that calcineurin may work with MBK-2 through OMA-1. To see which neurotransmitter is affected in small body size of tax-6 calcineurin loss-of-function mutant, I treated worms with various neurotransmitters. Interestingly exogenous acetylcholine reversed small the body size in tax-6 loss-of-function mutants, while other tested neurotransmitters did not affect the body growth. In addition, cholinergic neuronal expression of tax-6 also recovered small body length of tax-6 mutants. These results suggest that calcineurin modulates acetylcholine release to control worm growth. In addition, loss-of-function mutation in calcineurin tax-6 suppresses TGF-β pathway, which leads to reduced body size. Therefore, calcineurin pathway may regulate organismal growth by selectively controlling acetylcholine neurotransmission that affects TGF-β signaling.