Proteoglycan biosynthesis in murine monocytic leukemic (M1) cells before and after differentiation.
Murine monocytic leukemic (M1) cells were cultured in the presence of [3H]glucosamine and [35S]sulfate. Labeled proteoglycans were purified by anion exchange chromatography and characterized by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with chemical and enzymatic degradation. M1 cells synthesize a single predominant species of proteoglycan which distributes almost equally between the cell and medium after 17 h labeling. The cell-associated proteoglycan has an overall size of about 135 kDa and contains three to five chondroitin sulfate chains (28-31 kDa each) attached to a chondroitinase-generated core protein of 28 kDa. The synthesis and subsequent secretion of this proteoglycan was enhanced 4-5-fold in cells induced to differentiate into macrophages. This was not a phenomenon of arrest in the G0/G1 stage of the cell cycle, since density inhibited undifferentiated cells arrested at this stage did not increase proteoglycan synthesis. The chondroitin sulfate chains contained exclusively chondroitin 4- and 6-sulfate; however, the ratio of these two disaccharides differed between the medium- and cell-associated proteoglycans, and changed during progression of the cells into a fully differentiated phenotype. Pulse-chase kinetics indicate the presence of two distinct pools of proteoglycan; one that is secreted very rapidly from the cell after a approximately 1-h lag, and a second pool that is turned over in the cell with a half-time of approximately 3.5 h. Subtle differences in the glycosylation patterns of the medium- and cell-associated species are consistent with synthesis of two pools. Papain digestion suggests that the chondroitin sulfate chains are clustered on a small protease resistant peptide. The data suggest that this proteoglycan is similar to the serglycin proteoglycan family.
McQuillan
D J
DJ
Bone Research Branch, National Institute of Dental Research, Bethesda, Maryland 20892.
Yanagishita
M
M
Hascall
V C
VC
Bickel
M
M
D000818:Animals; D002236:Carbohydrate Conformation; D002454:Cell Differentiation; D002460:Cell Line; D006868:Hydrolysis; D007700:Kinetics; D007951:Leukemia, Myeloid / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008970:Molecular Weight; D010447:Peptide Hydrolases; D011509:Proteoglycans / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism; D013462:Sulfur Radioisotopes; D014316:Tritium; D014407:Tumor Cells, Cultured / Q000378:metabolism* / Q000473:pathology
D011509:Proteoglycans; D013431:Sulfates; D013462:Sulfur Radioisotopes; D014316:Tritium; D010447:Peptide Hydrolases