Cubilin and megalin expression and their interaction in the rat intestine:effect of thyroidectomy

Cubilin is a 460-kDa multipurpose, multidomain receptor that contains an NH 2-terminal 110-residue segment followed by 8 epidermal growth factor (EGF)- like repeats and a contiguous stretch (representing nearly 88% of its mass) of 27 CUB (initially found in complement components C1r/C1s, Uegf, and bon e morphogenic protein-1) domains. Cubilin binds to intrinsic factor (IF)-co balamin (cbl, vitamin B-12) complex and promotes the ileal transport of cbl . The 460-kDa form of cubilin is the predominant form present in the apical brush-border membranes of rat intestine, kidney, and yolk sac, but a 230-k Da form of cubilin is also noted in the intestinal membranes. In thyroidect omized (TDX) rats, levels of intestinal brush-border IF-[Co-57]-labeled cbl binding, 460-kDa cubilin protein levels and tissue (kidney) accumulation o f cbl were reduced by similar to 70%. Immunoblot analysis using cubilin ant iserum of intestinal total membranes from TDX rats revealed cubilin fragmen ts with molecular masses of 200 and 300 kDa. Both of these bands, along wit h the 230-kDa band detected in the total membranes of control rats and unli ke the 460-kDa form, failed to react with antiserum to EGF. Mucosal membran e cubilin associated with megalin was reduced from similar to 12% in contro l to similar to4% in TDX rats, and this decreased association was not due t o altered megalin levels. Thyroxine treatment of TDX rats resulted in rever sal of all of these effects, including an increase to nearly 24% of cubilin associated with megalin. In vitro, megalin binding to cubilin occurred wit h the NH2-terminal region that contained the EGF-like repeats and CUB domai ns 1 and 2 but not with a downstream region that contained CUB domains 2-10 . These studies indicate that thyroxine deficiency in rats results in decre ased uptake and tissue accumulation of cbl caused mainly by destabilization and deficit of cubilin in the intestinal brush border.