In situ labeling of the adipocyte lipid binding protein with 3-[125I]iodo-4-azido-N-hexadecylsalicylamide. Evidence for a role of fatty acid binding proteins in lipid uptake.

Abstract

Differentiating 3T3-L1 cells have been used to investigate the process of fatty acid uptake, its cellular specificity, and the involvement of cytoplasmic carrier proteins. The profile of fatty acid uptake in both differentiated and undifferentiated cells was biphasic, consisting of an initial rapid phase (0-20 s) followed by a second slower phase (60-480 s). In both cell types the initial phase of fatty acid (FA) uptake was temperature-insensitive whereas the rate of uptake during the second phase decreased 4-fold when measurements were made at 4 degrees C. The rate of [9,10-3H]oleate uptake in 3T3-L1 adipocytes was 10-fold greater than in the fibroblastic precursor cells. The acquisition of a differentially expressed cytoplasmic fatty acid binding protein (adipocyte lipid binding protein (ALBP] occurs coincident with the increased ability of these cells to take up FAs. Uptake experiments with 3-[125I]iodo-4-azido-N-hexadecylsalicylamide demonstrated that this photoactivatable FA analogue accumulated intracellularly in a time-, temperature-, and cell-specific fashion. Moreover, when 3T3-L1 adipocytes were presented with 3-[125I]iodo-4-azido-N-hexadecylsalicylamide and then irradiated, a single cytoplasmic 15-kDa protein was labeled. The in situ-labeled 15-kDa protein was identified as ALBP by its ability to be immunoprecipitated with anti-ALBP antisera. Taken together these results indicate that fatty acids traverse the plasma membrane and are bound by ALBP in the cytoplasmic compartment. It is likely that lipid uptake in other cell systems, such as liver, heart, intestine, and nerve tissue, proceeds by a similar process and that this represents a general mechanism for cell-specific FA uptake and utilization.