Background— In a whole-genome scan, a single nucleotide polymorphism (SNP) (rs7566605) upstream of the insulin-induced gene 2 ( INSIG2 ) was shown to influence body mass index and obesity in the Framingham Heart Study, with replication of these results in an additional 4 of 5 studies. However, other studies could not replicate the association. Because INSIG2 plays an important role in cholesterol biosynthesis, we hypothesized that human INSIG2 variants might play a role in the regulation of plasma lipid and lipoprotein levels. Methods and Results— We selected tagging SNPs spanning >100 kb of INSIG2 locus and sequenced 18 434 base pairs to discover novel SNPs. Thirty-two SNPs were genotyped in 645 individuals from the Quebec Family Study. Two SNPs (rs10490626 and rs12464355) were associated with plasma low-density lipoprotein cholesterol (LDL-C) ( P <0.0015) and total apolipoprotein B (apoB) levels ( P <0.014), whereas no association was found between any SNP and body mass index. We replicated the finding of rs10490626 for both LDL-C and total apoB in additional study samples, including 758 individuals from Saguenay–Lac St. Jean, Quebec ( P =0.040 for LDL-C, P =0.044 for apoB), 3247 Europeans ( P =0.028 for LDL-C, P =0.030 for apoB), and 1695 South Asians ( P =0.0036 for LDL-C, P =0.034 for apoB) from the INTERHEART study (for LDL-C, the combined 2-sided P =6.2×10 −5 and for total apoB, P =0.0011). Furthermore, we identified a variant in the human sorbin and SH 3 -domain–containing-1 gene that was associated with INSIG2 mRNA levels, and this SNP was shown to act in combination with rs10490626 to affect LDL-C ( P =0.022) in the Quebec Family Study and in INTERHEART South Asians ( P =0.019) and Europeans ( P =0.052). Conclusion— These results suggest that INSIG2 genetic variants may have a more direct role in lipid and lipoprotein metabolism than in obesity.