Expression cloning of LDLB, a gene essential for normal Golgi function and assembly of the IdICp complex

The Chinese hamster ovary (CHO) cell mutants IdIC and IdIB, which exhibit almost identical pheno-types, define two genes required for multiple steps in the normal medial and trans Golgi-associated processing of glycoconjugates. The LDLC gene encodes IdICp, an ≈ 80-kDa protein, which in wild-type, but not IdIB, cells associates reversibly with the cytoplasmic surface of the Golgi apparatus. Here, we have used a retrovirus-based expression cloning system to clone a murine cDNA, LDLB, that corrects the pleiotropic mutant phenotypes of IdIB cells. The corresponding mRNA was not detected in IdIB mutants. LDLB encodes an ≈110-kDa protein, IdIBp, which lacks homology to known proteins and contains no common structural motifs. Database searches identified short segments of homology to sequences from Drosophila melanogaster, Arabidopsis thaliana, and Caenorhabditis elegans, and the essentially full-length homologous human sequence (82% identity); however, as was the case for IdICp, no homologue was identified in Saccharomyces cerevisiae. We have found that in wild-type cell cytosols, IdICp is a component of an ≈950-kDa "IdICp complex," which is smaller, ≈700 kDa, in IdIB cytosols. Normal assembly of this complex is IdlBp-dependent and may be required for Golgi association of IdICp and for the normal activities of multiple luminal Golgi processes.