Intracellular trafficking and degradation of unassociated proalpha2 chains of collagen type I.
Procollagen I is a trimer consisting of two proalpha1(I) chains and one proalpha 2(I) chain. In certain cases of mild osteogenesis imperfecta, abnormal proalpha1(I) chains are degraded very soon after synthesis. As a consequence, the cells produce excess proalpha2(I) chains, which cannot form trimers and are not secreted. The objective of this work was to determine the intracellular fate of unassociated proalpha2(I) chains. Mov13 mouse fibroblasts, which do not synthesize proalpha1(I) mRNA, but do produce proalpha2(I) mRNA, were incubated with radioactive amino acids using pulse-chase protocols, and proteins were analyzed by gel electrophoresis, autoradiography, and Western blotting. Mov13 cells produced proalpha2(I) chains that were degraded intracellularly within 30 min. Degradation was inhibited when cells were treated with brefeldin-A, which blocks transit from endoplasmic reticulum to Golgi. Fixed cells exposed to various immunofluorescence markers and imaged by confocal laser scanning microscopy showed that proalpha2(I) chains colocalized with Golgi and lysosome markers. Degradation was inhibited and chains were secreted when cells were treated with wortmannin, which blocks trafficking to lysosomes. These results demonstrate that unassociated proalpha2(I) chains leave the endoplasmic reticulum, transit the Golgi, and enter lysosomes where they are degraded.
Gotkin
Marilyn G
MG
Program in Biology, Graduate Center of the City University of New York, New York 11016, USA.
Ripley
Catherine R
CR
Lamande
Shireen R
SR
Bateman
John F
JF
Bienkowski
Robert S
RS
D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002461:Cell Line, Transformed; D003094:Collagen / Q000378:metabolism; D024042:Collagen Type I / Q000737:chemistry / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism / Q000648:ultrastructure; D006056:Golgi Apparatus / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D021122:Protein Subunits / Q000378:metabolism*; D021381:Protein Transport
D024042:Collagen Type I; D021122:Protein Subunits; D020126:Brefeldin A; D003094:Collagen