Quantitative analysis of cellular phosphorylation signaling using electrospray ionization-mass spectrometry with an air current-regulating system

• Project/Area Number: 25513006
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: オミクス計測科学
• Research Institution: Yokohama City University
• Principal Investigator: Kawakami Takao 横浜市立大学, 生命医科学研究科, 客員准教授 (40366117)
• Project Period (FY): 2013-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000); Fiscal Year 2015: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2014: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2013: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
• Keywords: リン酸化 / プロテオミクス / 質量分析 / プロテオーム / 分析科学

Outline of Final Research Achievements

Quantitative and comprehensive information of posttranslational modifications of proteins is indispensable for elucidating precisely the cellular control mechanism. The optimized data analysis software improved our label-free LC-MS/MS proteomics system. We applied this system into phosphoproteomics of tumor stem cells from breast cancer. siRNA-knockdown of atypical protein kinase C lambda and zeta resulted in suppression of growth and survival of the cells, together with significant reduction of phosphorylation levels of a variety of signaling protein molecules as well as the target kinases themselves.

Research Products

• [Journal Article] Augmentation of multiple protein kinase activities associated with secondary imatinib resistance in gastrointestinal stromal tumors as revealed by quantitative phosphoproteome analysis (2015)
  • Author(s): Nagata K, Kawakami T, Kurata Y, Kimura Y, Suzuki Y, Nagata T, Sakuma Y, Miyagi Y, Hirano H.
  • Journal Title: Journal of Proteomics Volume: 115 Pages: 132-142
  • DOI: 10.1016/j.jprot.2014.12.012
  • Peer Reviewed / Acknowledgement Compliant
• [Presentation] LC-MS/MSを用いたバイオマーカー開発の方法論と実際 (2015)
  • Author(s): 川上隆雄
  • Organizer: 第55回日本臨床化学会年次学術集会（シンポジウム発表）
  • Place of Presentation: 大阪大学コンベンションセンター（大阪府吹田市）
  • Year and Date: 2015-10-30
• [Presentation] リン酸化プロテオミクスを用いた乳癌ALDH1high細胞におけるAtypical Protein Kinase C(aPKC)の機能解析 (2015)
  • Author(s): 165.片山鈴花、中根裕美、稲田将大、小野寺佑佳、神保美穂、原泰志、安部良、田沼靖一、川上隆雄、秋本和憲
  • Organizer: 第59回日本薬学会関東支部大会
  • Place of Presentation: 日本大学薬学部（千葉県船橋市）
  • Year and Date: 2015-09-12
• [Presentation] LC-MS/MSのデータ比較解析ソフトウェアi-RUBYの開発と腫瘍リン酸化プロテオミクスへの応用 (2015)
  • Author(s): 神保美穂、上田尚学、高畑良雄、片山鈴花、秋本和憲、川上隆雄
  • Organizer: 日本プロテオーム学会2015年会
  • Place of Presentation: くまもと森都心プラザ（熊本県熊本市）
  • Year and Date: 2015-07-23
• [Presentation] Proteomics of axons during presynapse formation using the novel culture method "neuron ball culture" (2013)
  • Author(s): Yukio Sasaki, Akiyo Ishikawa, Takao Kawakami, Hisashi Hirano, Yoshio Goshima
  • Organizer: HUPO 12th Annual World Congress
  • Place of Presentation: Pacifico Yokohama Japan
• [Presentation] Quantitative variation of protein components and their phosphorylations in the SWI/SNF chromatin remodeling complex associated with high malignancy of ovarian clear cell adenocarcinoma (2013)
  • Author(s): Ayuko Kimura, Ayako Nomura, Takao Kawakami, Noriaki Arakawa, Hisashi Hirano
  • Organizer: HUPO 12th Annual World Congress
  • Place of Presentation: Pacifico Yokohama Japan
• [Presentation] Performance of the label-free differential LC-MS/MS incorporating peptide identification items to associate peptide detection signals among measurements (2013)
  • Author(s): Takao Kawakami, Makiko Otsuji, Keiko Nagasaka
  • Organizer: HUPO 12th Annual World Congress
  • Place of Presentation: Pacifico Yokohama Japan
• [Book] プロテオミクス辞典 (2013)
  • Author(s): 日本プロテオーム学会（分担執筆）
  • Total Pages: 127
  • Publisher: 講談社