Clathrin coated vesicles in Neurospora crassa.

Electropherograms of Neurospora crassa homogenates showed a polypeptide with a mobility slightly lower than that of a standard sample of clathrin (from bovine brain). Subcellular fractionation of the homogenate resulted in a 20-fold enrichment of the putative N. crassa clathrin in the microsomal fraction. Further fractionation of the microsomal fraction by glass bead permeation chromatography yielded a fraction enriched about 150-fold relative to the homogenate. Coated vesicles (42.5 +/- 2.5 nm diameter) were found in this preparation by electron microscopy of negatively stained specimens. Ribosomes were virtually absent from this sample. N. crassa clathrin remained associated with the coated vesicles after repeated centrifugation and homogenization steps, even in the presence of 0.4 M-NaCl, but was released by treatment with Tris buffer pH 8.5. However the polypeptide was again sedimentable after dialysis against Mes buffer pH 6.5. Under the electron microscope this sediment resembled the empty coats of higher eukaryotes. The results taken together indicate that a clathrin-like protein occurs in wild type cells of N. crassa.