Urinary excretion of nonsulfated HNK-1 glycans is associated with renal dysfunction

Nonsulfated human natural killer-1 (nsHNK-1) glycan is a trisaccharide biosynthesized by β-1,3-glucuronyltransferase 2 (GlcAT-S) and expressed in the mouse kidney, especially the apical membrane of proximal tubules. Although such apical membrane proteins are known to shed into urine, the presence and significance of urinary nsHNK-1 glycans have remained unexplored. Here, we demonstrated that nsHNK-1 glycan is detectable in mouse urine and identified its major carrier as meprin A subunit alpha (MEP1A), a metalloprotease enriched at the apical membrane of proximal tubules and known to be excreted into urine. Mass spectrometry revealed nsHNK-1 glycosylation at four sites on MEP1A, with the highest occupancy at Asn221 in the catalytic domain. In GlcAT-S knockout mice, urinary MEP1A lacked the nsHNK-1 glycan and exhibited reduced enzymatic activity, suggesting nsHNK-1 glycosylation has a functional contribution. Induced tubular injury decreased urinary MEP1A-associated nsHNK-1 levels, following a similar pattern to reduced MEP1A-associated nsHNK-1 levels in the kidney. In contrast, glomerular injury resulted in increased MEP1A-associated nsHNK-1 expression in the kidney, accompanied by elevated urinary nsHNK-1 levels. These findings suggest that urinary nsHNK-1 levels differentially reflect renal dysfunction and may serve as a novel biomarker of kidney function.