Regulation of Nutrient Transport in Quiescent, Lactating, and Neoplastic MammaryEpithelia

摘要

We studied developmental changes in glucose transporter (GT) targeting in mammarygland, a prerequisite for the understanding GT targeting in breast cancer. Previously, we established that GLUT1 is targeted to Golgi during lactation. To understand the regulation of this process, we carried out subcellular fractionation and density gradient centrifugation, Western blotting, and immunofluorescence in mammary glands of mothers whose pups were prematurely weaned. We conclude: (1) There is enrichment of Golgi by GLUT1 during lactation. This is lost by 5h of weaning. (2) Enrichment can be restored by returning the pups to the mother for 5h. (3) At 10h, total cellular content of GLUT1 begins to decrease. (4) A 72 kD protein recognized by the GLUT1 antibody showed even more striking Golgi enrichment than GLUT1. (5) Intermediate MW forms at 50 and 65 kD were also observed. These each demonstrate specific patterns of appearance, disappearance, and subcellular localization, and can be deglycosylated to give aglyco GLUT1. (6) The 72 kD protein was resistant to deglycosylation. Based on the kinetics of its appearance and disappearance, its physicochemical properties which suggest it is not a GT, and based on its subcellular localization in the Golgi, p72 is an excellent candidate for a protein involved in sequestering GTs within the Golgi. (7) Ubiquitin appears to play an important role in the rapid degradation of GLUT1 and p72 during premature weaning. Differential display analysis of gene expression regulated by prolactin and dexamethasone has identified five novel genes, and full length clones are being isolated. One of these may encode a protein with a motif similar to the trans Golgi targeting motif of the GLUT4 GT.