title: TAILS N-Terminomics and Proteomics Show Protein Degradation Dominates over Proteolytic Processing by Cathepsins in Pancreatic Tumors.
abstract: Deregulated cathepsin proteolysis occurs across numerous cancers, but in vivo substrates mediating tumorigenesis remain ill-defined. Applying 8-plex iTRAQ terminal amine isotopic labeling of substrates (TAILS), a systems-level N-terminome degradomics approach, we identified cathepsin B, H, L, S, and Z in vivo substrates and cleavage sites with the use of six different cathepsin knockout genotypes in the Rip1-Tag2 mouse model of pancreatic neuroendocrine tumorigenesis. Among 1,935 proteins and 1,114 N termini identified by TAILS, stable proteolytic products were identified in wild-type tumors compared with one or more different cathepsin knockouts (17%-44% of 139 cleavages). This suggests a lack of compensation at the substrate level by other cathepsins. The majority of neo-N termini (56%-83%) for all cathepsins was consistent with protein degradation. We validated substrates, including the glycolytic enzyme pyruvate kinase M2 associated with the Warburg effect, the ER chaperone GRP78, and the oncoprotein prothymosin-alpha. Thus, the identification of cathepsin substrates in tumorigenesis improves the understanding of cathepsin functions in normal physiology and cancer.
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Anna
authors.initials: A
authors.lastname: Prudova
authors.affiliation: Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York City, NY 10021, USA.
authors.forename: Vasilena
authors.initials: V
authors.lastname: Gocheva
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Auf dem Keller
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Eckhard
authors.affiliation: Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York City, NY 10021, USA.
authors.forename: Oakley C
authors.initials: OC
authors.lastname: Olson
authors.affiliation: Ludwig Institute for Cancer Research, 1066 Lausanne, Switzerland; Department of Oncology, University of Lausanne, 1066 Lausanne, Switzerland.
authors.forename: Leila
authors.initials: L
authors.lastname: Akkari
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Georgina S
authors.initials: GS
authors.lastname: Butler
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Fortelny
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Philipp F
authors.initials: PF
authors.lastname: Lange
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: Mark
authors.affiliation: Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York City, NY 10021, USA; Ludwig Institute for Cancer Research, 1066 Lausanne, Switzerland; Department of Oncology, University of Lausanne, 1066 Lausanne, Switzerland. Electronic address: johanna@joycelab.org.
authors.forename: Johanna A
authors.initials: JA
authors.lastname: Joyce
authors.affiliation: Centre for Blood Research, Life Sciences Institute and Faculty of Dentistry, Department of Oral Biological and Medical Sciences, University of British Columbia, Vancouver, BC V6T 1Z3, Canada. Electronic address: chris.overall@ubc.ca.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Overall
mesh_terms: D000818:Animals; D063646:Carcinogenesis / Q000378:metabolism; D002403:Cathepsins / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins / Q000378:metabolism; D008822:Mice, Transgenic; D015513:Oncogene Proteins / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D013379:Substrate Specificity / Q000502:physiology
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; C000717248:Hspa5 protein, mouse; D015513:Oncogene Proteins; D020543:Proteome; D002403:Cathepsins
keywords: ECM; TAILS degradomics; cysteine cathepsins; degradation; lysosomal hydrolases; pancreatic neuroendocrine cancer; proteases; proteolytic processing; proteomics; substrate discovery