Cathepsin Inhibition Prevents Autophagic Protein Turnover and Downregulates Insulin Growth Factor-1 Receptor-Mediated Signaling in Neuroblastoma

Abstract

Inhibition of the major lysosomal proteases, cathepsins B, D, and L, impairs growth of several cell types but leads to apoptosis in neuroblastoma. The goal of this study was to examine the mechanisms by which enzyme inhibition could cause cell death. Cathepsin inhibition caused cellular accumulation of fragments of the insulin growth factor 1 (IGF-1) receptor. The fragments were located in dense organelles that were characterized as autophagosomes. This novel discovery provides the first clear link between lysosomal function, autophagy, and IGF-1-mediated cell proliferation. A more in-depth analysis of the IGF1 signaling pathway revealed that the mitogen-activated protein kinase (MAPK) cell-proliferation pathway was impaired in inhibitor treated cells, whereas the Akt cell survival pathway remained functional. Shc, an adapter protein that transmits IGF-1 signaling through the MAPK pathway, was sequestered in autophagosomes; whereas IRS-2, an adapter protein that transmits IGF-1 signaling through the Akt pathway, was unaffected by cathepsin inhibition. Furthermore, Shc was sequestered in autophagosomes as its active form, indicating that autophagy is a key mechanism for downregulating IGF-1-induced cell proliferation. Cathepsin inhibition had a greater effect on autophagic sequestration of the neuronal specific adapter protein, Shc-C, than ubiquitously expressed Shc-A, providing mechanistic support for the enhanced sensitivity of neuronally derived tumor cells. We also observed impaired activation of MAPK by epidermal growth factor treatment in inhibitor-treated cells. The Shc adapter proteins are central to transducing proliferation signaling by a range of receptor tyrosine kinases; consequently, cathepsin inhibition may become an important therapeutic approach for treating neuroblastoma and other tumors of neuronal origin.

Control of Rab7a activity and localization through endosomal type Igamma PIP 5‐kinase is required for endosome maturation and lysosome function

“…These hydrolases are synthesized in the rough endoplasmic reticulum and travel along the secretory pathway until they reach the TGN, where they are sorted and delivered to late endosomes/lysosomes . The inhibition of lysosomal hydrolases, such as cathepsins, can induce lysosome dysfunction and prevent autophagic degradation . To determine whether PIPKIγi5 and/or Rab7a control the sorting of hydrolases to lysosomes, the subcellular distribution of the lysosomal hydrolase cathepsin D (CTSD) was monitored.…”

“…5,41 The inhibition of lysosomal hydrolases, such as cathepsins, can induce lysosome dysfunction and prevent autophagic degradation. 42,43 To determine whether PIPKIγi5 and/or Rab7a control the sorting of hydrolases to lysosomes, the subcellular distribution of the lysosomal hydrolase cathepsin D (CTSD) was monitored. The loss of PIPKIγi5 significantly decreased the colocalization of CTSD with the late endosome/lysosome marker LAMP1 ( Figure 5A,B), suggesting there was a decrease in sorting to late endosomes/lysosomes.…”

Inducing hypertrophic effects of type I skeletal muscle fibers: A hypothetical role of time under load in resistance training aimed at muscular hypertrophy

“…Despite this modestly increased protein synthesis capacity, protein degradation mechanisms, such as autophagy, are known to be increased in the oxidative fibers [7]. This is supported by findings that cathepsins, important factors in lysosomal proteolysis that are usually abundant in tissues with high protein turnover, are present in higher concentrations in muscle fibers with a high oxidative capacity [11,12]. Due to a greater oxidative capacity of type I muscle fibers, higher accumulation of reactive oxygen species and metabolites are expected to occur, lowering the biological potential for hypertrophy due to activation of the pathways responsible for the protein degradation, acting as a quality control system [13,14].…”

The trophic effect of nerve growth factor in primary cultures of rat hippocampal neurons is associated to an anti‐inflammatory and immunosuppressive transcriptional program

“…In particular, Panx1 is involved in neuronal cell death regulation (Nishida et al, ), while Bgn and Fn1 have neuroprotective effects (Koops et al, ; Yanqing, Yu‐Min, Jian, Bao‐Guo, & Chuan‐Zhen, ). Shc is known to act downstream of the TrkA receptor and of the Akt survival signaling‐pathway (Soori, Lu, & Mason, ), whereas Anxa2 induces apoptotic signals (Bozzo, Bellomo, Silengo, Tarone, & Altruda, ; Han, Zhang, Lu, & Xu, ; Jin et al, ). Following Ngf treatment, our analysis also revealed the down‐regulation of Btk, Ship , Pik3c γ, Lyn, and Nalp3 , which encode for enzymes that regulate immune cell signaling and increase neuronal viability (Campbell, Raheem, Malemud, & Askari, ; Ha, Kwon, & Park, ; Ho, Drego, Hakimian, & Masliah, ; Huang, Hunter, Kim, Indik, & Schreiber, ; Islam & Smith, ; Kawakami et al, ; Zhang et al, ).…”