Expression of the extracellular domain of OB-cadherin as an Fc fusion protein using bicistronic retroviral expression vector.
Osteoblast cadherin (OB-cadherin, also known as cadherin-11) is a Ca(2+)-dependent homophilic cell adhesion molecule that is expressed mainly in osteoblasts. OB-cadherin is expressed in prostate cancer and may be involved in the homing of metastatic prostate cancer cells to bone. The extracellular domain of OB-cadherin may be used to inhibit the adhesion between prostate cancer cells and osteoblasts. In this report, we describe the expression of the extracellular domain of OB-cadherin as an Fc fusion protein (OB-CAD-Fc) in human embryonic kidney 293FT cells using a bicistronic retroviral vector. Coexpression of GFP and OB-CAD-Fc through the bicistronic vector permitted enrichment of OB-CAD-Fc-expressing cells by fluorescence-activated cell sorting. Recombinant OB-CAD-Fc proteins were secreted into cell medium, and about 0.85 mg of purified OB-CAD-Fc protein was purified from 1l of the conditioned medium using immobilized protein A-affinity chromatography. The purified OB-CAD-Fc was biologically active because it supported the adhesion of PC3 cells and L cells transduced with OB-cadherin. The availability of OB-CAD-Fc offers opportunities to test whether OB-CAD-Fc can be used to inhibit OB-cadherin-mediated prostate cancer bone metastasis in vivo or to generate antibodies for inhibiting the adhesion between prostate cancer cells and osteoblasts.
Lira
Cristina B B
CB
Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, Unit 89, 1515 Holcombe Boulevard, Houston, TX 77030, USA. liracbb@gmail.com
Chu
Khoi
K
Lee
Yu-Chen
YC
Hu
Mickey C-T
MC
Lin
Sue-Hwa
SH
D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics*; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line; D002846:Chromatography, Affinity / Q000379:methods; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors
D015820:Cadherins; D017077:Culture Media, Conditioned; D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins; C086960:osteoblast cadherin