Screening for regulatory proteins of osteoclastogenesis with lentiviral shRNA library

2015 Fiscal Year Final Research Report

• Project/Area Number: 24592806
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Functional basic dentistry
• Research Institution: Saga University
• Principal Investigator: Kukita Akiko 佐賀大学, 医学部, 准教授 (30153266)
• Co-Investigator(Kenkyū-buntansha): Kukita Toshio 九州大学, 大学院歯学研究院, 教授 (70150464)
• Co-Investigator(Renkei-kenkyūsha): Nishioka Kenichi 佐賀大学, 医学部, 講師 (80370120) ; Kuhara Satoru 九州大学, 大学院農学研究院, 教授 (00153320)
• Project Period (FY): 2012-04-01 – 2016-03-31
• Keywords: 破骨細胞分化 / shRNAライブラリー / スクリーニング

Outline of Final Research Achievements

We performed the screening for proteins which have essential roles in osteoclast differentiation and function by using shRNA lentivirus library that target signaling pathway in mouse. First, we prepared a reporter cell line RAW-ctsk-Venus1 which induce the expression of fluorescent protein Venus under the control of cathepsin K promoter. RANKL dose dependently induced Venus expression in RAW-ctsk-Venus1. Surface expression of population of Venus-positive cells was analyzed. RANK expression of Venus　low-positive cells was lower than that of Venus high-positive cells. RAW-ctsk-Venus 1 cells were infected by shRNA lentivirus library and stimulated with RANKL. The fraction of Venus low-positive cells affected by infection were collected by cell sorter. Analyzing of enriched population of shRNA in Venus low-positive cells revealed that several candidates for regulators of osteoclastogenesis including some signaling molecules Map3k and Vav3.

Free Research Field

医歯薬学