A Macro View of a Micronutrient: Revealing the Genetics and Elucidating the Impact of Serum Sulfate on Human Health in the Old Order Amish

Abstract

Sulfate (SO42-) is integral in the biotransformation of multiple compounds through sulfotransferase-mediated sulfate conjugation (sulfation). These compounds include xenobiotics, hormones, glycosaminoglycans, and neurotransmitters. Two non-linked, nonsense variants (R12X and W48X) in SLC13A1, which encodes the sodium-sulfate cotransporter, NaS1, responsible for sulfate (re)absorption in the intestines and kidneys, were found to be enriched in an Old Order Amish (Amish) cohort compared to outbred populations. Serum sulfate was measured in 977 Amish subjects and established to be heritable (h2=0.40). Both SLC13A1 R12X and W48X were independently associated with a 27.6% (P=2.7x10-8) and 27.3% (P=6.9x10-14) decrease in serum sulfate, respectively (P=8.8x10-20 for carriers of either SLC13A1 nonsense variant). We further performed the first exome- and genome-wide association study (ExWAS/GWAS) of serum sulfate and identified a missense variant (L348P) in SLC26A1, also associated with decreased serum sulfate (P=4.4x10-12) and enriched in this Amish cohort. SLC26A1 encodes the basolateral sulfate-anion transporter (Sat1), indicative of this locus as a true determinant of serum sulfate. Consistent with sulfate's role in xenobiotic detoxification and protection against acetaminophen-induced hepatotoxicity, SLC13A1 nonsense variant carriers had higher aminotransferase levels. Contrary to our hypothesis that sulfate-lowering variants would be associated with decreased DHEA-S and DHEA-S/DHEA ratio, we instead observed a 16% decrease in DHEA amongst SLC13A1 nonsense variant carriers, compared to non-carriers (P=0.01). SLC26A1 L348P was associated with lower whole-body bone mineral density and higher serum calcium, consistent with the osteochondrodysplasia exhibited by dogs and sheep with naturally-occurring, homozygous, loss-of-function mutations in Slc13a1, and the nephrocalcinosis and increased calcium oxalate kidney stone formation in Slc26a1-knockout mice. However, serum sulfate was not associated directly with any of these clinical phenotypes. In addition, we created and validated a novel slc13a1-knockdown zebrafish model, resulting in slc13a1 morphants that appear phenotypically analogous to the phenotypes observed in Slc13a1/NaS1-null mammals, particularly in regard to their gross, tail, and gastrointestinal abnormalities and decreased body length. These findings provide important and novel insights into the genetic regulation of serum sulfate and its biological and clinical implications in humans while simultaneously demonstrating the power and translational potential of systematic identification and characterization of rare nonsense variants in founder populations.