Proteolytic cleavage of the disease-related lysosomal membrane glycoprotein CLN7.
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Proteolytic cleavage of the disease-related lysosomal membrane glycoprotein CLN7. / Steenhuis, Pieter; Froemming, Joshua; Reinheckel, Thomas; Storch, Stephan.
In: Biochim Biophys Acta, Vol. 1822, No. 10, 10, 2012, p. 1617-1628.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - Proteolytic cleavage of the disease-related lysosomal membrane glycoprotein CLN7.
AU - Steenhuis, Pieter
AU - Froemming, Joshua
AU - Reinheckel, Thomas
AU - Storch, Stephan
PY - 2012
Y1 - 2012
N2 - CLN7 is a polytopic lysosomal membrane glycoprotein of unknown function and is deficient in variant late infantile neuronal ceroid lipofuscinosis. Here we show that full-length CLN7 is proteolytically cleaved twice, once proximal to the used N-glycosylation sites in lumenal loop L9 and once distal to these sites. Cleavage occurs by cysteine proteases in acidic compartments and disruption of lysosomal targeting of CLN7 results in inhibition of proteolytic cleavage. The apparent molecular masses of the CLN7 fragments suggest that both cleavage sites are located within lumenal loop L9. The known disease-causing mutations, p.T294K and p.P412L, localized in lumenal loops L7 and L9, respectively, did not interfere with correct lysosomal targeting of CLN7 but enhanced its proteolytic cleavage in lysosomes. Incubation of cells with selective cysteine protease inhibitors and expression of CLN7 in gene-targeted mouse embryonic fibroblasts revealed that cathepsin L is required for one of the two proteolytic cleavage events. Our findings suggest that CLN7 is inactivated by proteolytic cleavage and that enhanced CLN7 proteolysis caused by missense mutations in selected luminal loops is associated with disease.
AB - CLN7 is a polytopic lysosomal membrane glycoprotein of unknown function and is deficient in variant late infantile neuronal ceroid lipofuscinosis. Here we show that full-length CLN7 is proteolytically cleaved twice, once proximal to the used N-glycosylation sites in lumenal loop L9 and once distal to these sites. Cleavage occurs by cysteine proteases in acidic compartments and disruption of lysosomal targeting of CLN7 results in inhibition of proteolytic cleavage. The apparent molecular masses of the CLN7 fragments suggest that both cleavage sites are located within lumenal loop L9. The known disease-causing mutations, p.T294K and p.P412L, localized in lumenal loops L7 and L9, respectively, did not interfere with correct lysosomal targeting of CLN7 but enhanced its proteolytic cleavage in lysosomes. Incubation of cells with selective cysteine protease inhibitors and expression of CLN7 in gene-targeted mouse embryonic fibroblasts revealed that cathepsin L is required for one of the two proteolytic cleavage events. Our findings suggest that CLN7 is inactivated by proteolytic cleavage and that enhanced CLN7 proteolysis caused by missense mutations in selected luminal loops is associated with disease.
KW - Animals
KW - Humans
KW - Mice
KW - COS Cells
KW - Cercopithecus aethiops
KW - Mutation, Missense
KW - HEK293 Cells
KW - Fibroblasts/metabolism
KW - Proteolysis
KW - Cell Line, Transformed
KW - Cathepsin L/metabolism
KW - Cysteine Proteases/metabolism
KW - Lysosome-Associated Membrane Glycoproteins/genetics/metabolism
KW - Lysosomes/genetics/metabolism
KW - Membrane Transport Proteins/genetics/metabolism
KW - Animals
KW - Humans
KW - Mice
KW - COS Cells
KW - Cercopithecus aethiops
KW - Mutation, Missense
KW - HEK293 Cells
KW - Fibroblasts/metabolism
KW - Proteolysis
KW - Cell Line, Transformed
KW - Cathepsin L/metabolism
KW - Cysteine Proteases/metabolism
KW - Lysosome-Associated Membrane Glycoproteins/genetics/metabolism
KW - Lysosomes/genetics/metabolism
KW - Membrane Transport Proteins/genetics/metabolism
M3 - SCORING: Journal article
VL - 1822
SP - 1617
EP - 1628
JO - Biochim Biophys Acta
JF - Biochim Biophys Acta
SN - 0006-3002
IS - 10
M1 - 10
ER -
