Shedding light on the mechanisms of cystinuria

Calcium ions tie cystine transporters for correct trafficking and function

A team from the Max Planck Institute of Biophysics (Frankfurt am Main), The Jikei University School of Medicine (Tokyo) and Nara Medical University (Nara) in Japan explored the molecular mechanisms of a genetic disease called cystinuria by studying the structure and function of the responsible amino acid transporter known as system b0,+ or b0,+AT-rBAT. Electron cryo-microscopy revealed the structure of ovine b0,+AT-rBAT in lipid nanodiscs: it shows the super-dimeric (b0,+AT-rBAT)2 complex in a curved lipid bilayer. The structures further reveal a previously uncharacterized calcium ion, which appears to play key roles in stabilizing the super-dimeric complex. The results explain how some of the known mutations lead to protein trafficking deficiencies in cystinuria.

Cystinuria is an inherited disorder in which defective amino acid reabsorption in the kidney causes over-excretion of cystine and dibasic amino acids into urine. Since cystine tends to crystallize in aqueous solution, the patients suffer from kidney stones and sometimes result in kidney failure. Two proteins are implicated in the disease, rBAT and b0,+AT, the mutations of which exhibit different phenotypes, called type I and non-type I. Although positions of disease-causing mutations have been spotted by the decades of studies, the molecular mechanisms by which those mutations lead to cystinuria remained elusive.

Using electron cryo-microscopy and multiple biochemical and cell biological methods, the group found that calcium ions in the extracellular domain of rBAT tie two molecules of the rBAT-b0,+AT complexes together, forming a protein complex called the super-dimer. Calcium binding promotes the trafficking of rBAT- b0,+AT from the endoplasmic reticulum to the plasma membrane, thereby allowing the transporter to function at the correct location. One of the common cystinuria mutants, T261M, were found to disrupt the super-dimer and thereby prevented plasma membrane localization. This was the first demonstration that defects in super-dimerization can cause the disease.

"The finding was unexpected in the beginning, " said Yongchan, “but as we dug into the role of calcium and super-dimerization by using multiple biochemical methods, we got gradually convinced that these two factors play key roles in the biogenesis and disease-related mutations of this transporter.” Although the study did not directly show how we can cure the disease, it does provide a new perspective in understanding cystinuria.

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