STUDY OF THE CI-MPR IN LIVER OF NEWBORN AND ADULT RATS

JOFRE GF, CARVELLI L, BIANCO F, SARTOR T, SOSA MA

Merlo, San Luis (Argentina)

Congreso; XXII Annual Scientific Meeting of the Cuyo Biology Society; 2004

Sociedad de Biología de Cuyo

  The cation-independent mannose-6-phosphate receptor (CI-MPR) has been widely studied since it can recognize several types of ligands. A body of evidence demonstrated that CI-MPR can also exist as homodimer in cell membranes, although it is still contro-versial if this feature is related to its binding capability. We previ-ously demonstrated that CI-MPR expression in rat liver decreases progressively from fetuses to adults. However, the Kd values indi-cated that the affinity for mannose-6-phosphate bearing ligands is higher at late stages of development. Since that, we wondered if this fact is related to the dimer/monomer (D/M) ratio in the mem-branes at different ages. In this study we explored this hypothesis by stabilizing the pre-existing dimers in the membranes at 4°C, using the irreversible cross-linker Disuccinimidyl suberate, in new-born and adult rats. The membrane proteins were then solubilized with TritonX-100 and loaded to a phosphomannan affinity matrix. The CI-MPR bound to the column were eluted with 5 mM of man-nose- 6-phosphate, and analyzed by SDS-PAGE. We observed that dimers and monomers were bound to the column and the ratio D/M was similar at the ages studied. We also confirmed that dimeric forms are stabilized by disulfide bonds. Interestingly, we found two bands around 300 kDa (monomeric) in both ages, wich are stabi-lized by the cross-linker, indicating that CI-MPR may interact with other small membrane proteins (judged by the vicinity of the bands). We concluded that the difference in CI-MPR kDa values between adult and newborn rats is not due to the D/M ratio, but possibly to the presence of neighbor proteins in the membranes.