Study of Intracellular Localization and Traffic of Newly Synthesized Ceruloplasmin Receptor in Cultured Human Fibroblasts

L. K. Sasina, L. V. Puchkova^*, and V. S. Gaitskhoki

Research Institute for Experimental Medicine, Russian Academy of Medical Sciences, ul. Akademika Pavlova 12, St. Petersburg, 197376 Russia; fax: (812) 234-9489; E-mail: ludmila@usr8.iem.ras.spb.ru

^* To whom correspondence should be addressed.

Received October 27, 1997; Revision received January 13, 1998

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Ceruloplasmin (Cp) receptor in cells of non-hepatocyte lineage (human HT-1080 fibroblasts) is synthesized by membrane-bound polyribosomes and then becomes a resident of the plasma membrane. The intracellular traffic of [¹⁴C]Cp receptor was followed in pulse-chase experiments using specific antibodies. It was shown that pulse-labeled Cp receptor, after reaching the place of its residence in the plasma membrane, is retained there for 90 min and then appears in the cytosol. Immunoactive 20-kD fragments of Cp receptor were found in the culture medium 1.5 h later. The intracellular traffic of ¹²⁵I-labeled Cp bound to the fibroblast cell surface was traced in parallel chase experiments. It was shown that the internalized Cp receptor was recovered from the floating fraction of the cytosol. Comparison of the dynamics of the retention of internalized [¹⁴C]Cp receptor and ¹²⁵I-labeled Cp in the subcellular compartments demonstrated that the traffic of both proteins within the fibroblasts is coordinated in time and proceeds via a common route. The role of Cp receptor in copper uptake by non-hepatocyte cells is discussed.

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KEY WORDS: copper metabolism, ceruloplasmin receptor, ceruloplasmin

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