title: Calpain mobilizes Atg9/Bif-1 vesicles from Golgi stacks upon autophagy induction by thapsigargin.
abstract: CAPNS1 is essential for stability and function of the ubiquitous calcium-dependent proteases micro- and milli-calpain. Upon inhibition of the endoplasmic reticulum Ca2+ATPase by 100 nM thapsigargin, both micro-calpain and autophagy are activated in human U2OS osteosarcoma cells in a CAPNS1-dependent manner. As reported for other autophagy triggers, thapsigargin treatment induces Golgi fragmentation and fusion of Atg9/Bif-1-containing vesicles with LC3 bodies in control cells. By contrast, CAPNS1 depletion is coupled with an accumulation of LC3 bodies and Rab5 early endosomes. Moreover, Atg9 and Bif-1 remain in the GM130-positive Golgi stacks and Atg9 fails to interact with the endocytic route marker transferrin receptor and with the core autophagic protein Vps34 in CAPNS1-depleted cells. Ectopic expression of a Bif-1 point mutant resistant to calpain processing is coupled to endogenous p62 and LC3-II accumulation. Altogether, these data indicate that calpain allows dynamic flux of Atg9/Bif-1 vesicles from the Golgi toward the budding autophagosome.
authors.affiliation: C.I.B. National Laboratory, AREA Science Park, Padriciano 99, Trieste 34149, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Marcassa
authors.affiliation: C.I.B. National Laboratory, AREA Science Park, Padriciano 99, Trieste 34149, Italy.
authors.forename: Marzia
authors.initials: M
authors.lastname: Raimondi
authors.affiliation: Department of Biosciences, University of Helsinki, PO Box 56, Helsinki 00014, Finland.
authors.forename: Tahira
authors.initials: T
authors.lastname: Anwar
authors.affiliation: Department of Biosciences, University of Helsinki, PO Box 56, Helsinki 00014, Finland.
authors.forename: Eeva-Liisa
authors.initials: EL
authors.lastname: Eskelinen
authors.affiliation: International Centre for Genetic Engineering and Biotechnology, AREA Science Park - Padriciano 99, Trieste 34149, Italy.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Myers
authors.affiliation: International Centre for Genetic Engineering and Biotechnology, AREA Science Park - Padriciano 99, Trieste 34149, Italy.
authors.forename: Gianluca
authors.initials: G
authors.lastname: Triolo
authors.affiliation: C.I.B. National Laboratory, AREA Science Park, Padriciano 99, Trieste 34149, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Schneider
authors.affiliation: C.I.B. National Laboratory, AREA Science Park, Padriciano 99, Trieste 34149, Italy francesca.demarchi@lncib.it.
authors.forename: Francesca
authors.identifier: http://orcid.org/0000-0003-1565-3162
authors.initials: F
authors.lastname: Demarchi
keywords: Autophagy; Bif-1; CAPNS1; Calpain; Endophilin B1; Thapsigargin