CLONING AND FUNCTIONAL-CHARACTERIZATION OF MAMMALIAN HOMOLOGS OF THE COPII COMPONENT SEC23

We screened a human cDNA library with a probe derived from a partial S EC23 mouse homologue and isolated two different cDNA clones (hSec23A a nd hSec23B) encoding proteins of a predicted molecular mass of 85 kDa. hSec23Ap and hSec23Bp were 85% identical and shared 48% identity with the yeast Sec23p. Affinity-purified anti-hSec23A recognized a protein of similar to 85 kDa on immunoblots of human, mouse, and rat cell ext racts but did not recognize yeast Sec23p. Cytosolic hSec23Ap migrated with an apparent molecular weight of 350 kDa on a gel filtration colum n, suggesting that it is part of a protein complex. By immunoelectron microscopy, hSec23Ap was found essentially in the ribosome-free transi tional face of the endoplasmic reticulum (ER) and associated vesicles. hSec23Ap is a functional homologue of the yeast Sec23p as the hSec23A isoform complemented the temperature sensitivity of the Saccharomyces cerevisiae sec23-1 mutation at a restrictive temperature of 34 degree s C. RNase protection assays indicated that both hSec23 isoforms are c oexpressed in various human tissues, although at a variable ratio. Our data demonstrate that hSec23Ap is the functional human counterpart of the yeast COPII component Sec23p and suggest that it plays a similar role in mammalian protein export from the ER. The exact function of hS ec23Bp remains to be determined.