Examination of directly frozen rough endoplasmic reticulum (ER) of retinal pigment epithelial cells by freeze-fracture and freeze-substitution revealed distinct paired transmembrane proteins associated with membrane ribosomes. Ribosomal subunits on intact ER membrane are directly visualized for the first time, providing a global view of the structure of the ribosome and the corresponding structures on the ER membrane, The ribosomal intersubunit cleft appears to be continuous with a cleft between paired transmembrane proteins that extends into the lumen of the ER, This continuous cleft may be the path taken by nascent polypeptides. (C) 1996 Academic Press, Inc.