Cell surface glycan engineering reveals that matriglycan alone can recapitulate dystroglycan binding and function

Abstract

Abstractα-Dystroglycan (α-DG) is uniquely modified on O-mannose sites by a repeating disaccharide (-Xylα1,3-GlcAβ1,3-)n termed matriglycan, which is a receptor for laminin-G domain-containing proteins and employed by old-world arenaviruses for infection. Using chemoenzymatically synthesized matriglycans printed as a microarray, we demonstrate length-dependent binding to Laminin, Lassa virus GP1, and the clinically-important antibody IIH6. Utilizing an enzymatic engineering approach, an N-linked glycoprotein was converted into a IIH6-positive Laminin-binding glycoprotein. Engineering of the surface of cells deficient for either α-DG or O-mannosylation with matriglycans of sufficient length recovers infection with a Lassa-pseudovirus. Finally, free matriglycan in a dose and length dependent manner inhibits viral infection of wildtype cells. These results indicate that matriglycan alone is necessary and sufficient for IIH6 staining, Laminin and LASV GP1 binding, and Lassa-pseudovirus infection and support a model in which it is a tunable receptor for which increasing chain length enhances ligand-binding capacity.

Structural conservation of Lassa virus glycoproteins and recognition by neutralizing antibodies

“…This array presents chemoenzymatically-generated matriglycan oligosaccharides of defined length and shows length-dependent binding of LIV GPC-I53-50A to matriglycan, consistent with previous observations of GP1 and pseudovirus binding (Fig. S7E, Sheikh et al, 2022). Whereas LIV GPC-I53-50A showed strong binding to the microarray with 24 repeating disaccharide units, the same protein complexed with 12.1F bound markedly less.…”

“…To elucidate the mechanism of binding and neutralization for these mAbs, we performed nano differential scanning fluorimetry (nanoDSF) experiments, a matriglycan microarray competition assay (Sheikh et al, 2022), and BLI-based LAMP-1 competition experiments (Fig. 3C, D, and E).…”

“…In contrast, 12.1F and 19.7E had only marginal effects on GPC thermostability, suggesting these GP1-A mAbs likely do not neutralize by stabilizing the prefusion state of GPC. To probe the interaction between GPC and the matriglycan moieties of β-dystroglycan, we used a synthetic matriglycan microarray (Sheikh et al, 2022). This array presents chemoenzymatically-generated matriglycan oligosaccharides of defined length and shows length-dependent binding of LIV GPC-I53-50A to matriglycan, consistent with previous observations of GP1 and pseudovirus binding (Fig.…”

“…The synthesis of matriglycan compounds were reported previously (Sheikh et al, 2022). All compounds were printed on NHS-ester activated glass slides (NEXTERION® Slide H, Schott Inc.) using a Scienion sciFLEXARRAYER S3 non-contact microarray equipped with a Scienion PDC80 nozzle (Scienion Inc.).…”

“…LASV exploits two host cell receptors to infect human cells. Host cell attachment is mediated by matriglycan moieties on α-dystroglycan which interact with residues on the GPC trimer apex (Acciani et al, 2017; Katz et al, 2022; Sheikh et al, 2022; Willard et al, 2018). Upon macropinocytosis and trafficking of LASV through the endosomal compartments (Oppliger et al, 2016), GPC undergoes a pH-dependent switch allowing binding to endosomal receptor lysosomal-associated membrane protein 1 (LAMP-1).…”