Molecular cloning and expression of cDNAs encoding human alpha-mannosidase II and a previously unrecognized alpha-mannosidase IIx isozyme.

Abstract

Golgi a-mannosidase II (a-Mll) is an enzyme involved in the processing of N-linked glycans. Using a previously isolated murine cDNA clone as a probe, we have isolated cDNA clones encompassing the human a-MII cDNA open reading frame and initiated isolation of human genomic clones. During the isolation of genomic clones, genes related to that encoding a-MIl were isolated. One such gene was found to encode an isozyme, designated acMIIx. A 5-kb cDNA clone encoding a-MIIX was then isolated from a human melanoma cDNA library. However, comparison between a-MIIX and a-MIl cDNAs suggested that the cloned cDNA encodes a truncated polypeptide with 796 amino acid residues, while a-MI consists of 1144 amino acid residues. To reevaluate the sequence of a-MIIx cDNA, polymerase chain reaction (PCR) was-performed with lymphocyte mRNAs. Comparison of the sequence of PCR products with the a-MIIX genomic sequence revealed that alternative splicing of the a-MIIX transcript can result in an additional transcript encoding a 1139-amino acid polypeptide. Northern analysis showed transcription of a-MIIX in various tissues, suggesting that the a-MIIX gene is a housekeeping gene. COS cells transfected with c-MIIX cDNA containing the full-length open reading frame showed an increase of a-mannosidase activity. The ca-MIIX gene was mapped to human chromosome 15q25, whereas the a-MIl gene was mapped to 5q21-22.a-Mannosidase (a-M) activities are involved in both biosynthesis and catabolism of N-linked glycans (1, 2). These enzyme activities are present in cells ranging from yeast to human. There are different forms of a-Ms: lysosomal a-Ms are soluble and involved in degradation of N-glycans, endoplasmic reticulum (ER) and Golgi a-Ms are involved in processing of newly synthesized N-glycans, and cytoplasmic a-Ms may be involved in degradation of dolichol intermediates that are not needed for protein glycosylation or oligosaccharides derived from glycoprotein turnover in the ER (1).. Substrate specificities of these a-Ms differ from each other, and Golgi a-MIl specifically hydrolyzes two peripheral mannosyl residues from Manal1-6(Manal --3)Manal ->6(GlcNAcf31--2Mana1 ->3)[Man/31 ->4GlcNAc,lB --4GlcNAc31 ->]asparagine structure. Several a-Ms have been cloned to date. These include Golgi a-MIT (3, 4), ER/cytosolic a-MI (5), two isozymes of Golgi a-MI (6-8), lysosomal a-M (9), Dictyostelium a-M (10), and yeast a-M (11) The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Conserved oligomeric Golgi complex subunit 1 deficiency reveals a previously uncharacterized congenital disorder of glycosylation type II

“…Furthermore, there was a dramatic loss of Golgi localization of ␤-1,4 galactosyltransferase I immunoreactivity observed in the patient's cells that could be reversed by reintroducing wild-type COG1 cDNA. Golgi mannosidase II levels were substantially decreased in the patient's cells, although no hybrid N-glycan structures were detected, perhaps because of the compensatory activity of other Golgi mannosidases, such as mannosidase IIx (30). Thus, multiple abnormalities in the proteins associated with the glycosylation machinery of the Golgi apparatus are probably responsible for the global galactosylation and sialylation defects in the patient's cells and therefore the CDG-II phenotype.…”

Human STX Polysialyltransferase Forms the Embryonic Form of the Neural Cell Adhesion Molecule

“…It is, on the other hand, noteworthy that hPST is located on chromosome 5, band q21. An isozyme of ␣-mannosidase II, ␣-manII x , was also located to chromosome 5, band q21, while ␣-mannosidase II resides on chromosome 15, band q25 (47). Moreover, a tumor suppressor gene, the adenomatous polyposis coli (APC) gene, is located on 5q21 (48).…”

Essential and mutually compensatory roles of α-mannosidase II and α-mannosidase IIx in N -glycan processing in vivo in mice

“…Previous studies suggest that human MX is catalytically active and plays a role in N-glycan biosynthesis (16)(17)(18). When mouse Man2a2, the orthologue of human MAN2A2, was disrupted, MX-nulls were apparently normal, except that mutant males were subfertile (18).…”

“…Human MX (16) is the product of the MAN2A2 gene and is homologous to human MII, which is encoded by MAN2A1. Previous studies suggest that human MX is catalytically active and plays a role in N-glycan biosynthesis (16)(17)(18).…”