The alternative use of exons 2 and 3 in cathepsin B mRNA controls enzyme trafficking and triggers nuclear fragmentation in human cells.
Pathological overexpression and trafficking of the cysteine peptidase cathepsin B depend in part on the composition of its mRNA. To investigate the roles of the alternatively spliced exons 2 and 3 in the 5'-untranslated region of cathepsin B mRNA we produced constructs of cathepsin B fused to green fluorescent protein. Expression and trafficking of the fluorescent chimeric products was followed in living human immortalized chondrocytes and HeLa cells. Although synthesized at different rates, proteins encoded by the full transcript and by that missing exon 2 followed a classic route, with the endosomal-lysosomal compartment as the final target. The point-mutated variant missing the glycosylation site for lysosomal targeting followed the secretory pathway. A truncated form of cathepsin B lacking the signal peptide and part of the propeptide, and encoded by the construct missing exons 2 and 3, was neither found in the Golgi apparatus nor in vesicles, but rather in the cytoplasm as patches associated with membranous and short fibrillar elements. This particular form of truncated cathepsin B produced nuclear damage and shrinking of the trans Golgi network and of the acidic compartment. The C-terminal, six-amino acid-long propeptide of cathepsin B did not exhibit a discernible role in protein trafficking.
Müntener
Kathrin
K
Institute of Biochemistry, University of Zurich, Winterthurerstrasse 190, Switzerland.
Zwicky
Roman
R
Csucs
Gabor
G
Baici
Antonio
A
D017398:Alternative Splicing; D002401:Cathepsin B / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D002461:Cell Line, Transformed; D002467:Cell Nucleus; D019902:Chondrocytes / Q000378:metabolism; D053938:DNA Fragmentation; D017931:DNA Primers / Q000737:chemistry; D005091:Exons; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015854:Up-Regulation
D017931:DNA Primers; D008164:Luminescent Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D002401:Cathepsin B