MAN1B1-CDG: Three new individuals and associated biochemical profiles

Abstract

Congenital disorders of glycosylation (CDG) constitute an ever-growing group of genetic diseases affecting the glycosylation of proteins. CDG individuals usually present with severe multisystem disorders. MAN1B1-CDG is a CDG with nonspecific clinical symptoms such as intellectual deficiency and developmental delay. Although up to 40 affected individuals were described so far, its final diagnosis is not straightforward using common biochemical methods due to the trace-level accumulation of defective glycan structures. In this study, we present three unreported MAN1B1-CDG individuals and propose a decision tree to reach diagnosis using a panel of techniques ranging from exome sequencing to gel electrophoresis and mass spectrometry. The occurrence of MAN1B1-CDG in patients showing unexplained intellectual disability and development delay, as well as a particular transferrin glycosylation profile, can be ascertained notably using matrix assisted laser desorption/ionization - time of flight (MALDI-TOF) mass spectrometry analysis of endo-β-acetylglucosaminidase H-released serum N-glycans. In addition to reporting new pathogenic variants and additional clinical signs such as hypersialorrhea, we highlight particular biochemical features of MAN1B1-CDG with potential glycoprotein-specific glycosylation defects.

Keywords: 2-DE, two-dimensional electrophoresis; A1AT, α1-antitrypsin; ApoC-III, apolipoprotein C-III; BMI, body mass index; CDG; CDG, congenital disorder(s) of glycosylation; CE, capillary electrophoresis; DD, developmental delay; DWI, Diffusion-weighted imaging; ER, endoplasmic reticulum; ESI-QTOF, electrospray ionization – quadrupole time of flight; Endo H, endo-ß-N-acetylglucosaminidase H; FLAIR, fluid-attenuated inversion recovery; HPLC, high performance liquid chromatography; Hpt, haptoglobin; Hypersialorrhea; ID, intellectual disability; Intellectual disability; M6, Man6GlcNAc2; M8A/B/C, Man8GlcNAc2 lacking the first/middle/third terminal mannose; M9, Man9GlcNAc2; MALDI-TOF, matrix assisted laser desorption/ionization – time of flight; MAN1B1; MRI, magnetic resonance imaging; MS, mass spectrometry; Man, mannose; N-glycan mass spectrometry; PNGase F, peptide-N-glycosidase F; Trf, transferrin; WES, whole exome sequencing.

Fig. 1

Involvement of the MAN1B1 enzyme in the secretory pathway. A) After the calnexin/calreticulin (CNX/CRT) cycle, MAN1B1 removes the Man residue of the middle branch of the GlcNAc2Man9 (“M9”) glycan linked to proteins. Glycoproteins harboring the resulting “M8B” structure are preferentially directed towards sorting and secretion. B) In case of MAN1B1 deficiency, the glycoproteins harboring the “M8A” or the “M8C” glycan structures are more likely degraded but could also result (via the schematized “M6” moiety) in glycoproteins harboring abnormal hybrid-type oligosaccharides.

Fig. 2

Capillary electrophoresis (CE) transferrin patterns of control and MAN1B1-CDG patients' sera. A) Compared to the control, CE transferrin (Trf) profiles of the three patients (P1, P2, P3) shared an important and isolated increase of the 3-sialo Trf fraction. These profiles are suggestive of a Trf protein variant (left rectangle). B) After neuraminidase treatment, CE Trf profiles of the patients showed one 0-sialo Trf peak, excluding a protein variant (showing two 0-sialo Trf peaks after neuraminidase treatment, as illustrated in the right rectangle). The observed shift of the asialo Trf peak of patient 2 probably results from a homozygous Trf protein variant and/or a misinterpretation of the Phoresys software.

Fig. 3

MALDI-TOF mass spectra of permethylated N-glycans released from serum samples from a healthy subject and the three MAN1B1-CDG patients following A) PNGase F, and B) Endo H treatment. Measurements were performed in the positive-ion mode and all ions are present in sodiated form. Green circles, mannose; yellow circles, galactose; blue squares, N-acetyl glucosamine; red triangles, fucose; purple diamonds, sialic acid. Structures of hybrid-type N-glycans at m/z 2390.2 (PNGase F) and m/z 2145.1 (Endo H) were suggested following the work of Messina et al. [17].