예쁜꼬마선충을 이용한 단백질 인산화효소의 유전자 연구

Abstract

칼시뉴린은 칼슘/칼모듈린에 의존적인 세린/트레오닌 단백질 탈인산화효소로 뉴런에서 높게 발현되며 많은 기능을 가지고 있다고 알려져 있다. 그런 기능 중에서 우리는 몸길이를 조절하는 것에 초점을 맞추었다. 칼시뉴린 돌연변이체는 작은 몸길이를 가지며 뉴런특이적으로 tax-6의 발현을 처리해주면 몸길이가 원래의 상태로 되돌아간다는 연구 결과가 있다. 이러한 연구결과에 따라 우리는 예쁜꼬마선충에서 칼시뉴린에 관여하는 인산화효소를 연구하였다. 몸길이를 조절하는데 칼시뉴린과 인산화효소 사이의 관계를 확인하기 위해서 이전에 우리는 RNAi 스크리닝을 이용하여 positive 인산화효소 후보군을 찾았다. 이번 연구에서 우리는 F13C5.2, PAK-, PKC-1 and MBK-2 의 4가지 인산화효소 후보군을 택하여 이들의 돌연변이체들을 주문하였다. 그 후 tax-6 loss-of-funtion 돌연변이체와 이중 돌연변이체를 만들고 몸길이를 측정하였다. 몸길이 측정의 결과 4종류의 이중 돌연변이체가 작은 몸길이 표현형을 회복하였다. |Calcineurin is a Ca2+/calmodulin�dependent serine/threonine protein phosphatase and highly expressed in neuron and is well known to have various biological functions. Among these functions, we focused on the regulation of body length. The previous reports showed that calcineurin loss-of-function mutants have short body length and are rescued by the neuron specific overexpression of TAX-6. In order to identify kinases that counteract calcineurin in the regulation of body length, we screened positive kinase candidates using RNAi that rescued tax-6 RNAi feeding worm�s short body size. In this study, we selected 4 positive kinase candidates (F13C5.2, PAK-, PKC-1 and MBK-2) and investigated 9 mutants. We crossed kinase and tax-6 loss-of-function to make double mutants and the body length was measured. The double mutants of pak-2(tm867); tax-6(ok2065), pkc-1(ok563); tax-6(ok2065), pkc-1(tm540); tax-6(ok2065) and mbk-2(ne992); tax-6(ok2065) rescued tax-6 small body size phenotype.; Calcineurin is a Ca2+/calmodulin�dependent serine/threonine protein phosphatase and highly expressed in neuron and is well known to have various biological functions. Among these functions, we focused on the regulation of body length. The previous reports showed that calcineurin loss-of-function mutants have short body length and are rescued by the neuron specific overexpression of TAX-6. In order to identify kinases that counteract calcineurin in the regulation of body length, we screened positive kinase candidates using RNAi that rescued tax-6 RNAi feeding worm�s short body size. In this study, we selected 4 positive kinase candidates (F13C5.2, PAK-, PKC-1 and MBK-2) and investigated 9 mutants. We crossed kinase and tax-6 loss-of-function to make double mutants and the body length was measured. The double mutants of pak-2(tm867)