Demonstration of the specific binding of bovine transferrin to the human transferrin receptor in k562 cells: Evidence for interspecies transferrin internalization

✍ Scribed by Larisa Tsavaler; Barry S. Stein; Howard H. Sussman

• Publisher: John Wiley and Sons
• Year: 1986
• Tongue: English
• Weight: 804 KB
• Volume: 128
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.1041280102

✦ Synopsis

Specific binding of ferric bovine transferrin to the human transferrin receptor was investigated using K562 cells propagated in serum-free medium without transferrin supplemented with elemental iron. Affinit chromatography of solubilized extracts of K562 cells surface-labeled with 5l was performed using bovine transferrin-and human transferrin-Sepharose 48 resins. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of resin eluates reveal that bovine transferrin specifically binds a M, = 188,000 protein which dissociates into a M, = 94,000 protein under reducing conditions, a finding identical to what is seen with human transferrin. The M, = 94,000 reduced protein isolated by bovine transferrin resin shows an identical one-dimensional partial proteolytic digestion map with that of t h e human transferrin receptor.

Unlabeled bovine transferrin was shown to specifically compete 1251-Iabeled human transferrin from the human transferrin receptor on the surface of K562 cells at 4OC in a similar manner as unlabeled human transferrin; however, approximately a 2,000-fold higher concentration of bovine ligand was required to achieve comparable competition (50% inhibition of binding).

Indirect immunofluorescence cytolocalization of bovine transferrin in K562 cells grown in serum-free medium supplemented with ferric bovine transferrin reveal patterns similar to those seen for human transferrin (both focal perinuclear and diffuse cytoplasmic fluorescence). Monensin treatment re- sults in a dramatic accumulation of bovine ligand in perinuclear aggregates, suggesting that it is recycled through the Golgi, as is human transferrin. K562 cells grown in serum-free medium supplemented with either 300 pg/ml of ferric human or ferric bovine +ransferrin were found to demonstrate superimposable growth curves.