LACK OF FUNCTION OF AN N-ETHYLMALEIMIDE-SENSITIVE THIOL PROTEIN IN ERYTHROCYTE-MEMBRANE OF AUTOSOMAL-DOMINANT POLYCYSTIC KIDNEY-DISEASE

The polycystic kidney disease 1 (PKD1) gene product polycystin has bee n predicted to be an integral membrane protein involved in cell-cell a nd cell-matrix interactions. The erythrocyte membrane fluidity in auto somal dominant polycystic kidney disease (ADPKD) patients is increased , and this may be due to a membrane cytoskeletal abnormality. The abno rmal erythrocyte sodium-lithium countertransport kinetics in ADPKD are related to an altered thiol protein in the cytoskeleton. The possibil ity that a similar thiol protein abnormality causes the increased eryt hrocyte membrane fluidity in ADPKD was investigated. The membrane flui dity of intact erythrocytes from 12 ADPKD patients and 12 healthy cont rol subjects was assessed from the fluorescence anisotropies of 1,6-di phenyl-1,3,5-hexatriene (DPH) and trimethylammonium-diphenylhexatriene (TMA-DPH). The effect on membrane fluidity of N-ethylmaleimide (NEM), cytochalasin D, heating at 48 degrees C for 20 min, or more specifica lly, liposomes containing antibodies to actin or ankyrin, was determin ed. In erythrocytes from healthy control subjects, the fluorescence an isotropy of DPH (mean +/- SEM: 0.223 +/- 0.001) was decreased after tr eatment with NEM (0.200 +/- 0.003, P < 0.001), cytochalasin D (0.206 /- 0.006, P < 0.001), heating (0.199 +/- 0.002, P < 0.001), and antibo dies to actin (0.194 +/- 0.002, P < 0.001) or ankyrin (0.196 +/- 0.002 , P < 0.001). The TMA-DPH anisotropy (0.279 +/- 0.001) was also decrea sed after treatment with NEM (0.264 +/- 0.001, P < 0.001), cytochalasi n D (0.264 +/- 0.001, P < 0.001), heating (0.265 +/- 0.001, P < 0.001) , and antibodies to actin (0.262 +/- 0.002, P < 0.001) or ankyrin (0.2 62 +/- 0.002, P < 0.001). NEM had no additional effect on the other tr eatments, suggesting that its target thiol protein was associated with the cytoskeleton. In untreated erythrocytes from ADPKD patients, fluo rescence anisotropies of both DPH and TMA-DPH were reduced, and none o f the treatments altered the anisotropy of either DPH or TMA-DPH. In A DPKD, a cytoskeletal thiol protein is abnormal and possibly explains a bnormal lipid bilayer properties and transport protein function in ery throcytes in this disease.