Qiao Hongjiang, Shibaki Akihiko, Long Heather A, Wang Gang, Li Qiang, Nishie Wataru, Abe Riichiro, Akiyama Masashi, Shimizu Hiroshi, McMillan James R
Creative Research Initiative, Hokkaido University, Sapporo, Japan.
J Invest Dermatol. 2009 Sep;129(9):2288-95. doi: 10.1038/jid.2009.20. Epub 2009 Feb 26.
Collagen XVII (COL17) participates in keratinocyte adhesion and possibly migration, as COL17 defects disrupt keratinocyte-basal lamina adhesion and underlie the disease non-Herlitz junctional epidermolysis bullosa. Using small interference RNA (siRNA) to knock down COL17 expression in HaCaT cells, we assessed cell characteristics, including adhesion, migration, and signaling. Control and siRNA-transfected keratinocytes showed no difference in adhesion on plastic dishes after incubation for 8 hours in serum-free keratinocyte-growth medium; however, when grown on collagen IV alone or BD matrigel (containing collagen IV and laminin isoforms), COL17-deficient cells showed significantly reduced adhesion compared with controls (P<0.01), and mitogen-activated protein kinase (MAPK)/ERK kinase (MEK)1/2 and MAPK showed reduced phosphorylation. Furthermore, COL17-deficient HaCaT cells plated on plastic exhibited reduced motility that was p38MAPK-dependent (after addition of the p38MAPK inhibitor SB203580). Together, these results suggest that COL17 has significantly wider signaling roles than were previously thought, including the involvement of COL17 in keratinocyte adhesion to collagen IV, in p38MAPK-dependent cell migration, and multiple cell signaling events pertaining to MEK1/2 phosphorylation.
ⅩⅦ型胶原蛋白(COL17)参与角质形成细胞的黏附,可能还参与其迁移,因为COL17缺陷会破坏角质形成细胞与基底膜的黏附,是导致非赫利茨型交界性大疱性表皮松解症的病因。我们利用小干扰RNA(siRNA)敲低HaCaT细胞中COL17的表达,评估细胞特性,包括黏附、迁移和信号传导。在无血清角质形成细胞生长培养基中孵育8小时后,对照和经siRNA转染的角质形成细胞在塑料培养皿上的黏附情况没有差异;然而,当单独在IV型胶原或BD基质胶(含IV型胶原和层粘连蛋白异构体)上生长时,与对照相比,COL17缺陷细胞的黏附显著降低(P<0.01),丝裂原活化蛋白激酶(MAPK)/细胞外信号调节激酶(ERK)1/2和MAPK的磷酸化水平降低。此外,接种在塑料上的COL17缺陷型HaCaT细胞的运动能力降低,且这种降低依赖于p38MAPK(添加p38MAPK抑制剂SB20358之后)。这些结果共同表明,COL17具有比之前认为的更为广泛的信号传导作用,包括COL17参与角质形成细胞与IV型胶原的黏附、p38MAPK依赖的细胞迁移以及与MEK1/2磷酸化相关的多个细胞信号事件。