TRANSPORT OF CHOLESTEROL FROM THE ENDOPLASMIC-RETICULUM TO THE PLASMA-MEMBRANE IS CONSTITUTIVE IN CACO-2 CELLS AND DIFFERS FROM THE TRANSPORT OF PLASMA-MEMBRANE CHOLESTEROL TO THE ENDOPLASMIC-RETICULUM

The transport of newly synthesized cholesterol from its site of synthe sis, the endoplasmic reticulum, to the plasma membrane was studied in CaCo-2 cells. The appearance of newly synthesized cholesterol on the c ell surface was rapid. By 30 min, 50% of the total labeled cholesterol was observed in the plasma membrane. The arrival of cholesterol at th e plasma membrane was independent of new protein synthesis, a function al Golgi apparatus, or microtubular function. Progesterone, verapamil, and trifluoperazine, inhibitors of p-glycoprotein which are known to inhibit cholesterol transport from the plasma membrane to the endoplas mic reticulum, reduced the amount of newly synthesized cholesterol rea ching the plasma membrane. The p-glycoprotein inhibitors, however, cau sed the accumulation of sterol intermediates in the plasma membrane, s uggesting that sterol trafficking to the plasma membrane remained inta ct, but that trafficking from the plasma membrane to the endoplasmic r eticulum was disrupted. In contrast, nigericin, another potent inhibit or of cholesterol movement from the plasma membrane to the endoplasmic reticulum, did not alter the transport of newly synthesized cholester ol to the plasma membrane. Moreover promoting cholesterol transport fr om the plasma membrane to the endoplasmic reticulum by sphingomyelin h ydrolysis or by micellar cholesterol influx did not alter the percent of newly synthesized cholesterol transported to the plasma membrane. L ikewise, preventing plasma membrane cholesterol from reaching the endo plasmic reticulum by incubating cells with lysophosphatidylcholine, fi lipin, or digitonin did not alter the arrival of newly synthesized cho lesterol to the plasma membrane. The results suggest that the amount o f cholesterol moving to the plasma membrane from the endoplasmic retic ulum is constitutive and regulated at the level of cholesterol synthes is and not at the level of the transport process The pathways of chole sterol transport to and from the plasma membrane are distinct.