Shedding of klotho by ADAMs in the kidney

Abstract

The anti-aging gene klotho plays an important role in Ca 2ϩ and phosphate homeostasis. Membrane-bound klotho is an essential coreceptor for fibroblast growth factor-23 and can be cleaved by proteases, including a disintegrin and metalloproteinase (ADAM)10 and ADAM17. Cleavage of klotho occurs at a site directly above the plasma membrane (␣-cut) or between the KL1 and KL2 domain (␤-cut), resulting in soluble full-length klotho or KL1 and KL2 fragments, respectively. The aim of the present study was to gain insights into the mechanisms behind klotho cleavage processes in the kidney. Klotho shedding was demonstrated using a Madin-Darby canine kidney cell line stably expressing klotho and human embryonic kidney-293 cells transiently transfected with klotho. Here, we report klotho expression on both the basolateral and apical membrane, with a higher abundance of klotho at the apical membrane and in the apical media. mRNA expression of ADAM17 and klotho were enriched in mouse distal convoluted and connecting tubules. In vitro ADAM/matrix metalloproteinase inhibition by TNF484 resulted in a concentration-dependent inhibition of the ␣-cut, with a less specific effect on ␤-cut shedding. In vivo TNF484 treatment in wild-type mice did not change urinary klotho levels. However, ADAM/matrix metalloproteinase inhibition did increase renal and duodenal mRNA expression of phosphate transporters, whereas serum phosphate levels were significantly decreased. In conclusion, our data show that renal cells preferentially secrete klotho to the apical side and suggest that ADAMs are responsible for ␣-cut cleavage.klotho; a disintegrin and metalloproteinase 10; a disintegrin and metalloproteinase 17; klotho shedding; a disintegrin and metalloproteinase/matrix metalloproteinase inhibitor CA 2ϩ AND PHOSPHATE HOMEOSTASIS is a tightly regulated process involving several hormones. Klotho is emerging as a key factor in this process (5,18,51). Initially, the klotho gene was discovered as an anti-aging gene, since lack of klotho in mice resulted in early aging and a premature death (23, 26), whereas overexpression of klotho led to an increased lifespan (28).Klotho is a single-pass type 1 transmembrane protein consisting of two extracellular domains, which are the KL1 and KL2 subunits. Full-length klotho has potential enzymatic activity (30, 49), and in the body, it exists in two forms, namely, membrane-bound and soluble klotho. Membrane-bound klotho is predominantly present in the renal distal convolution, consisting of the distal convoluted tubule (DCT) and connecting tubule (CNT), parathyroid glands, and choroid plexus in the brain (26,31). A key function of membrane-bound klotho is to act as an obligate cofactor for the fibroblast growth factor (FGF) receptor, thereby enabling binding and downstream signaling of FGF23 via the ERK1/2 MAPK pathway (27, 51). In the kidney, binding of FGF23 to the receptor complex results in enhanced phosphate excretion via reduced expression of the Na ϩ -phosphate transporters NaPi-IIa and NaPiIIc (12). Moreov...

Role of αKlotho and FGF23 in regulation of type II Na-dependent phosphate co-transporters

“…The extracellular domain of membrane αKlotho can be shed by secretases and released into the circulation and urine [8, 15, 42, 44, 97]. This cleaved αKlotho protein is hereinafter referred as soluble αKlotho.…”

Tissue expression and source of circulating αKlotho

“…Klotho is also released from the cell membrane by proteolytic cleavage, a process mediated by the alpha-secretases ADAM10 and 17 [4]. A first cut near the cell membrane (alpha-cut) produces a 130 kDa circulating protein, and a second cut (beta-cut) between the KL1 and KL2 domains produces two smaller proteins with molecular sizes of approximately 60-70 kDa [5]. In addition, a truncated Klotho variant containing only the KL1 domain may be produced through alternative splicing, although the physiological relevance of this splicing variant is unclear [6].…”

Recombinant α-Klotho may be prophylactic and therapeutic for acute to chronic kidney disease progression and uremic cardiomyopathy

“…2 α-Klotho is a single-pass transmembrane protein highly expressed in the kidney. 1 It is proteolytically cleaved (cleaved α-Klotho), 3–5 and its transcript is proposed to be alternatively spliced to generate a soluble form (secreted α-Klotho). 1 These extracellular polypeptides of α-Klotho constitute “soluble α-Klotho,” which functions as an endocrine substance with far-reaching effects.…”