Functional analyses of fibulin-4 on matrix assembly

• Project/Area Number: 26461661
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Dermatology
• Research Institution: Oita University
• Principal Investigator: Sasaki Takako 大分大学, 医学部, 助教 (30133193)
• Co-Investigator(Renkei-kenkyūsha): FUJIWARA Sakuhei 大分大学, 医学部, 教授 (90181411) ; UTANI Atsushi 長崎大学, 医学部, 教授 (10292707)
• Project Period (FY): 2014-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000); Fiscal Year 2016: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2015: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2014: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
• Keywords: 細胞外マトリックス / fibulin-4 / 変異 / 弾性線維 / アセンブリー / 遺伝子変異

Outline of Final Research Achievements

Fibulin-4 is a 60kDa calcium binding glycoprotein that has an important role in development and integrity of extracellular matrices. Several mutations in the fibulin-4 gene are known in patients whose major symptoms are vascular abnormalities and cutis laxa. In order to elucidate the pathogenetic mechanisms, we expressed fibulin-4 mutants recombinantly in HEK293 cells, purified the proteins in native forms and analyzed alterations in secretion, matrix assembly, and interaction with other proteins. Our studies show that different mutations affect these properties in multiple ways, resulting in fibulin-4 deficiency and/or impaired ability to form elastic fibers. The substitution C267Y impaired secretion of the protein. The A397T mutation introduced an extra O-glycosylation site and deleted binding to LTBP1s. The binding of fibulin-4 to the LOX propeptide was strongly reduced by the mutation E57K. These findings show that different mutations result in different molecular defects.

Research Products

• [Int'l Joint Research] University of Erlangen-Nuernberg/University of Regensburg/University of Koeln(ドイツ)
• [Int'l Joint Research] Thomas Jefferson University(米国)
• [Int'l Joint Research] University of Liverpool(英国)
• [Int'l Joint Research] University of Erlangen-Nuernberg/University of Regensburg(ドイツ)
• [Int'l Joint Research] Thomas Jefferson University(米国)
• [Int'l Joint Research] University of Liverpool(英国)
• [Int'l Joint Research] University of Erlangen-Nuernberg/University of Regensburg(ドイツ)
• [Int'l Joint Research] Thomas Jefferson University/University of South Florida/Washington University(米国)
• [Int'l Joint Research] TNO(オランダ)
• [Journal Article] Expression and purification of recombinant fibulins in mammalian cells. (2018)
  • Author(s): Hanada K, Sasaki T
  • Journal Title: Methods Cell Biol. Volume: 143 Pages: 247-259
  • DOI: 10.1016/bs.mcb.2017.08.014
  • Peer Reviewed
• [Journal Article] Fibulin-4 deposition requires EMILIN-1 in the extracellular matrix of osteoblasts. (2017)
  • Author(s): Schiavinato A, Keene DR, Imhof T, Doliana R, Sasaki T, Sengle G
  • Journal Title: Sci Rep. Volume: 7 Issue: 1 Pages: 5526-5526
  • DOI: 10.1038/s41598-017-05835-7
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Functional consequence of fibulin-4 missense mutations associated with vascular and skeletal abnormalities and cutis laxa (2016)
  • Author(s): Sasaki T, Hanisch FG, Deutzmann R, Sakai LY, Sakuma T, Miyamoto T, Yamamoto T, Hannappel E, Chu ML, Lanig H, von der Mark K
  • Journal Title: Matrix Biol Volume: 56 Pages: 132-149
  • DOI: 10.1016/j.matbio.2016.06.003
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Loss of fibulin-4 results in abnormal collagen fibril assembly in bone, caused by impaired lysyl oxidase processing and collagen cross-linking. (2016)
  • Author(s): Sasaki T, Stoop R, Sakai T, Hess A, Deutzmann R, Schlötzer-Schrehardt U, Chu M-L, von der Mark K
  • Journal Title: Matrix Biol. Volume: 50 Pages: 53-66
  • DOI: 10.1016/j.matbio.2015.12.002
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Forelimb Contractures and Abnormal Tendon Collagen Fibrillogenesis in Fibulin-4 Null Mice (2016)
  • Author(s): Markova D, Pan, T-C, Zhang R-Z, Zhang G, Sasaki T, Arita M, Birk D, Chu M-L
  • Journal Title: Cell Tissue Res. Volume: 印刷中 Issue: 3 Pages: 637-646
  • DOI: 10.1007/s00441-015-2346-x
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Fibulin-4 E57K Knock-in Mice Recapitulate Cutaneous, Vascular and Skeletal Defects of Recessive Cutis Laxa 1B with both Elastic Fiber and Collagen Fibril Abnormalities. (2015)
  • Author(s): Igoucheva O, Alexeev V, Halabi CM, Adams SM, Stoilov I, Sasaki T, Arita M, Donahue A, Mecham RP, Birk DE, Chu ML
  • Journal Title: J. Biol. Chem. Volume: 290 Issue: 35 Pages: 21443-21459
  • DOI: 10.1074/jbc.m115.640425
  • Peer Reviewed / Int'l Joint Research
• [Presentation] Loss of fibulin-4 results in abnormal collagen fibril assembly in bone, caused by impaired lysyl oxidase processing and collagen cross-linking (2017)
  • Author(s): Sasaki T, Stoop R, Sakai T, Hess A, Deutzmann R, Schloetzer-Schrehardt U, Chu ML, von der Mark K.
  • Organizer: Gordon Research Conference on Collagen
• [Presentation] fibulin-4欠損による骨コラーゲン線維形成異常 (2016)
  • Author(s): 佐々木隆子、Reinout Stoop, Takao Sakai, Andreas Hess, Rainer Deutzmann, Ursula Schloetzer-Schrehardt,Mon-Li, Dhu and Klaus von der Mark
  • Organizer: 第４８回結合組織学会学術大会
  • Place of Presentation: 長崎県長崎市
  • Year and Date: 2016-06-24
• [Presentation] 血管異常ならびに皮膚脆弱症を呈する患者で同定されたfibulin-4ミスセンス変異の及ぼす構造と機能への影響 (2015)
  • Author(s): 佐々木隆子、Franz-Georg Hanisch, Rainer Deutzmann, Lynn Sakai, Ewald Hannappel, Mon-Li Chu, Harald Lanig, Klaus von der Mark
  • Organizer: 日本結合組織学会
  • Place of Presentation: コクヨホール（東京都港区）
  • Year and Date: 2015-05-15 – 2015-05-16