Effects of transfection of ICAP-1α and its mutants on adhesion and migration of 2H-11 cells.
This study examined the effect of integrin cytoplasmic domain-associated protein 1α (ICAP-1α) and its mutatants T38A and I138A on the adhesion, migration and tube formation of 2H-11 cells. rAAV-ICAP-1α, rAAV-T38A and rAAV-I138A were constructed. After infection, the expression of ICAP-1α and p-ERK1/2, p-c-Jun protein was measured by Western blotting. Adhesion ability was evaluated by using MTT. Cell migration was determined by using Boyden chamber method. Tube formation test was conducted on Matrigel. The results showed that in ICAP-1α, T38A and I138A groups, ICAP-1α protein expression was increased. In T38A and I138A groups, phospho-ERK1/2, phospho-c-Jun protein expressions were significantly increased as compared with the control group and the GFP group. ICAP-1α group protein expression was obviously decreased when compared with the control group and the GFP group. Cell adhesion ratio was 0.1429±0.0080 in control group, 0.1434±0.0077 in GFP group and the ratio in T38A and I138A groups increased to 0.3210±0.0082 and 0.3250±0.0079, respectively. In ICAP-1α group, the ratio was decreased to 0.1005±0.0073. In T38A and I138A groups, the number of migrating 2H-11 cells was increased to 31.45±3.20 and 33.10±5.40 against 18.51±2.80 in control group and 20.47±3.12 in GFP group. In ICAP-1α group, the number was decreased to 12.06±1.72. The number of tube-like structures was increased to 20.41±2.54 in T38A and to 22.26±3.07 in I138A groups as compared to those of control group 12.45±1.84 and GFP group 13.63±2.71. In ICAP-1α group, the number of tube-like structures was decreased to 8.32±1.24. It was suggested that rAAV-T38A and rAAV-I138A transfection can substantially increase 2H-11 cell adhesion, migration and angiogenisis, while rAAV-ICAP-1α can greatly inhibit the effect. These effects might be correlated with ERK1/2 and c-Jun protein phosphorylation.
Department of Cardiology, Huazhong University of Science and Technology, Wuhan, China. zhjie_2001@163.com
Jie
J
Zhang
Wangcui
W
Luo
Zhengxiang
Z
Liu
Jingyang
J
Lin
Zhongliang
Z
Cheng
D048868:Adaptor Proteins, Signal Transducing; D002448:Cell Adhesion / Q000187:drug effects*; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects*; D000229:Dependovirus / Q000235:genetics / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000096:biosynthesis* / Q000235:genetics; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D050505:Mutant Proteins / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D005353:Fibronectins; C108257:ITGB1BP1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D050505:Mutant Proteins; D011994:Recombinant Proteins