Mutation of Nogo-B Receptor, a Subunit of cis-Prenyltransferase, Causes a Congenital Disorder of Glycosylation

Abstract

Summary Dolichol is an obligate carrier of glycans for N-linked protein glycosylation, O-mannosylation, and GPI anchor biosynthesis. Cis-prenyltransferase (cis-PTase) is the first enzyme committed to the synthesis of dolichol. However, the proteins responsible for mammalian cis-PTase activity have not been delineated. Here we show that Nogo-B receptor (NgBR) is a subunit required for dolichol synthesis in yeast, mice and man. Moreover, we describe a family with a congenital disorder of glycosylation caused by a loss of function mutation in the conserved C terminus of NgBR-R290H and show that fibroblasts isolated from patients exhibit reduced dolichol profiles and enhanced accumulation of free cholesterol identically to fibroblasts from mice lacking NgBR. Mutation of NgBR-R290H in man and orthologs in yeast proves the importance of this evolutionarily conserved residue for mammalian cis-PTase activity and function. Thus, these data provides a genetic basis for the essential role of NgBR in dolichol synthesis and protein glycosylation.

A Lettuce (Lactuca sativa) Homolog of Human Nogo-B Receptor Interacts with cis-Prenyltransferase and Is Necessary for Natural Rubber Biosynthesis

“…These biochemical data support our proposed role of CPTL1 as a "CPT-scaffolding protein on ER" that can modulate CPT activity, and it becomes apparent that both CPTL1 and CPT1 are required for efficient cis-polyisoprene biosynthesis. This finding is further supported by recent CPT activity assays, co-expressing human NgBR/CPT or yeast NUS1/CPT (35). Regrettably, in CPTL2/CPT3 assays, isoprene polymers larger than the regular dolichols could not be synthesized.…”

“…Because CPTL is clearly required in eukaryotes for cis-polyisoprene synthesis (whether it is dolichol or NR) (Ref. 35 and this work), the rubber tree should also require CPTL for dolichol and NR biosynthesis. We propose that the single copy of CPTL in the rubber tree has a dual role in supporting both the NR synthesis in latex and the ubiquitous dolichol synthesis in other cells.…”

“…Lacking all enzymatic activity, NgBR is able to interact with human CPT (24). Recently, NgBR and its homologs in mice and yeast (NUS1) were identified as necessary subunits for CPT activities in respective organisms (35). NgBR also interacts with Nogo-B and NPC2 for entirely unrelated physiological events, such as remodeling vascular epithelial cells and controlling intracellular trafficking of cholesterol, respectively (23,25).…”

cis-Prenyltransferase: New Insights into Protein Glycosylation, Rubber Synthesis, and Human Diseases

“…In the context of the crystal structure of homo-dimeric UPPS, Arg 242 in the -RXG-motif is involved IPP binding (26), and the very C terminus of one monomer of decaprenyl diphosphate synthase in M. tuberculosis interacts with the active site of the other subunit (52). Interestingly, patients harboring an R290H mutation in NgBR, and the corresponding mutations in ScNus1 and SpNus1, show reduced cis-PT activity and chain length of the polyprenol diphosphate (47). Finally C-terminal tagging of L. sativa CPT-like (CPTL-NgBR ortholog) (50) severely reduces cis-PTs activity, thereby supporting the idea that the C terminus is highly conserved and important for cis-PT activity.…”

“…However, recent data have challenged this paradigm because mammalian, fungal, and long-chain plant cis-PTs are composed of two subunits critical for enzymatic activity. The two-component system is composed of NgBR (Nogo-B receptor) and hCIT in mammals (47); Nus1 and Rer2 or Nus1 and Srt1 in Saccharomyces cerevisiae; SpNus1 and SpRer2 in Schizosaccharomyces pombe (47); SlCPT3 and SlCPTBP in tomato (48); Lew1 and At2g17570 in Arabidopsis (49); and LsCPTL1 and LsCPT1 in lettuce (50), respectively. Rubber synthase of Taraxacum brevicorniculatum and Lactuca sativa are also heteromeric enzymes composed of at least two subunits (50,51).…”