Renal aplasia in humans is associated with RET mutations

Abstract

In animal models, kidney formation is known to be controlled by the proteins RET, GDNF, and GFRA1; however, no human studies to date have shown an association between abnormal kidney development and mutation of these genes. We hypothesized that stillborn fetuses with congenital renal agenesis or severe dysplasia would possess mutations in RET, GDNF, or GFRA1. We assayed for mutations in these genes in 33 stillborn fetuses that had bilateral or unilateral renal agenesis (29 subjects) or severe congenital renal dysplasia (4 subjects). Mutations in RET were found in 7 of 19 fetuses with bilateral renal agenesis (37%) and 2 of 10 fetuses (20%) with unilateral agenesis. In two fetuses, there were two different RET mutations found, and a total of ten different sequence variations were identified. We also investigated whether these mutations affected RET activation; in each case, RET phosphorylation was either absent or constitutively activated. A GNDF mutation was identified in only one fetus with unilateral agenesis; this subject also had two RET mutations. No GFRA1 mutations were seen in any fetuses. These data suggest that in humans, mutations in RET and GDNF may contribute significantly to abnormal kidney development.

Figure 1

RET and GDNF Mutations in Fetuses with Bilateral or Unilateral Renal Agenesis Eight RET mutations were seen in 7/19 fetuses (37%) with bilateral renal agenesis (BA), and three RET mutations were discovered in 2/10 fetuses (20%) with unilateral renal agenesis (UA). One GDNF mutation occurred in 1/10 fetuses (10%) with unilateral renal agenesis.

Figure 2

Schematic Illustration of the Identified Mutations in RET and GDNF in Stillborn Fetuses with Either Bilateral Renal Agenesis or Unilateral Agenesis Triangles signify the location and type of mutations. RET: S, signaling domain; CLD, cadherin-like domain; CYS, cysteine-rich region; TM, transmembrane region; TK, tyrosine kinase domain. GDNF: S, signaling domain; CS, cleavage site.

Figure 3

Western Blots of Normal and Mutant RET51 Expressed in SK-N-MC Neuroblastoma Cells ± GDNF Blots were sequentially probed for phospho-RET51 (Y1062) (pRET), total RET51 (RET), and GFRA1. Top gels: cell lysates with no GDNF added to the media. KP7, KP16, KP17, and KP18 are constitutively phosphorylated. Bottom gels: cell lysates treated with GDNF. Normal RET51 is phosphorylated and KP7, KP16, KP17, and KP18 seem further phosphorylated. The RET mutations in KP14, KP19, KP25, and KP29 appear to inactivate RET51 (prevent autophosphorylation in response to GDNF).