Quality control of ER synthesized proteins: an exposed thiol group as a three-way switch mediating assembly, retention and degradation.
Plasma cells secrete IgM only in the polymeric form: the C-terminal cysteine of the mu heavy chain (Cys575) is responsible for both intracellular retention and assembly of IgM subunits. Polymerization is not quantitative, and part of IgM is degraded intracellularly. Neither chloroquine nor brefeldin A (BFA) inhibits degradation, suggesting that this process occurs in a pre-Golgi compartment. Degradation of IgM assembly intermediates requires Cys575: the monomeric IgMala575 mutant is stable also when endoplasmic reticulum (ER) to Golgi transport is blocked by BFA. Addition of the 20 C-terminal residues of mu to the lysosomal protease cathepsin D is sufficient to induce pre-Golgi retention and degradation of the chimeric protein: the small amounts of molecules which exit from the ER are mostly covalent dimers. By contrast, when retained by the KDEL sequence, cathepsin D is stable in the ER, indicating that retention is not sufficient to cause degradation. Replacing the C-terminal cysteine with serine restores transport through the Golgi. As all chimeric cathepsin D constructs display comparable protease activity in vitro, their different fates are not determined by gross alterations in folding. Thus, also out of its normal context, the mu chain Cys575 plays a crucial role in quality control, mediating assembly, retention and degradation.
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DIBIT-HSR, Milano, Italy.
Fagioli
C
C
Finazzi
D
D
Sitia
R
R
Alberini
C M
CM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002402:Cathepsin D / Q000378:metabolism; D002460:Cell Line; D003545:Cysteine / Q000378:metabolism; D004247:DNA; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D000882:Haplorhini; D007075:Immunoglobulin M / Q000096:biosynthesis* / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D010950:Plasma Cells / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism*
D007075:Immunoglobulin M; D013438:Sulfhydryl Compounds; D004247:DNA; D002402:Cathepsin D; D003545:Cysteine