Effect of various hepatic membrane fractions on microtubule assembly-with special emphasis on the role of membrane phospholipids.

Abstract

This report describes an interaction between rat brain microtubule protein and various hepatic fractions in vitro. Purified preparations of Golgi membranes, plasma membrane, rough and smooth endoplasmic reticulum, nuclear membranes, and mitochondria were obtained from the livers of 200-g rats. Several concentrations of fresh or sonicated frozen membranes were incubated with twice-cycled rat brain microtubule protein in a microtubule assembly buffer for 60 min at 30 degrees C. Changes in microtubule assembly were assessed either by quantitative electron microscopy on negatively stained samples or by spectrophotometric methods. The results show that all the tested membranes "bound" microtubule protein, preventing assembly: Golgi and plasma membranes, as well as mitochondria, were especially potent in this regard. To identify the membrane-associated components responsible for microtubule protein binding, the membranes were extracted with methanol-chloroform, and liposomes were prepared from the resulting lipids. Microtubule protein incubated with these liposomes showed a differential ability to assemble that was similar to the effect obtained with intact membranes. Membrane-extracted phospholipids were identified as the lipid component responsible for these changes, with the negatively charged phospholipids (cardiolipin and phosphatidylserine) being uniquely active. These findings indicate that hepatic membranes differentially interact with brain microtubule protein; this interaction may be dependent on membrane phospholipids.