Elevated mRNA expression and defective processing of cathepsin D in HeLa cells lacking the mannose 6‐phosphate pathway

Disruption of the mannose 6‐phosphate (M‐6‐P) pathway in HeLa cells by inactivation of the GNPTAB gene, which encodes the α/β subunits of GlcNAc‐1‐phosphotransferase, results in missorting of newly synthesized lysosomal acid hydrolases to the cell culture media instead of transport to the endolysosomal system. We previously demonstrated that the majority of the lysosomal aspartyl protease, cathepsin D, is secreted in these GNPTAB−/− HeLa cells. However, the intracellular content of cathepsin D in these cells was still greater than that of WT HeLa cells which retained most of the protease, indicating a marked elevation of cathepsin D expression in response to abrogation of the M‐6‐P pathway. Here, we demonstrate that HeLa cells lacking GlcNAc‐1‐phosphotransferase show a fivefold increase in cathepsin D mRNA expression over control cells, accounting for the increase in cathepsin D at the protein level. Further, we show that this increase at the mRNA level occurs independent of the transcription factors TFEB and TFE3. The intracellular cathepsin D can still be trafficked to lysosomes in the absence of the M‐6‐P pathway, but fails to undergo proteolytic processing into the fully mature heavy and light chains. Uptake experiments performed by feeding GNPTAB−/− HeLa cells with various phosphorylated cathepsins reveal that only cathepsin B is capable of partially restoring cleavage, providing evidence for a role for cathepsin B in the proteolytic processing of cathepsin D..

Medienart:

E-Artikel

Erscheinungsjahr:

2021

Erschienen:

2021

Enthalten in:

Zur Gesamtaufnahme - volume:11

Enthalten in:

FEBS Open Bio - 11(2021), 6, Seite 1695-1703

Sprache:

Englisch

Beteiligte Personen:

Lin Liu [VerfasserIn]
Balraj Doray [VerfasserIn]

Links:

doi.org [kostenfrei]
doaj.org [kostenfrei]
doi.org [kostenfrei]
Journal toc [kostenfrei]

Themen:

Biology (General)
Cathepsin B
Cathepsin D
Cathepsin L
GlcNAc‐1‐phosphotransferase
Lysosomes
Mannose 6‐phosphate pathway

doi:

10.1002/2211-5463.13169

funding:

Förderinstitution / Projekttitel:

PPN (Katalog-ID):

DOAJ056613342