title: Dynein function and protein clearance changes in tumor cells induced by a Kunitz-type molecule, Amblyomin-X.
abstract: Amblyomin-X is a Kunitz-type recombinant protein identified from the transcriptome of the salivary glands of the tick Amblyomma cajennense and has anti-coagulant and antitumoral activity. The supposed primary target of this molecule is the proteasome system. Herein, we elucidated intracellular events that are triggered by Amblyomin-X treatment in an attempt to provide new insight into how this serine protease inhibitor, acting on the proteasome, could be comparable with known proteasome inhibitors. The collective results showed aggresome formation after proteasome inhibition that appeared to occur via the non-exclusive ubiquitin pathway. Additionally, Amblyomin-X increased the expression of various chains of the molecular motor dynein in tumor cells, modulated specific ubiquitin linkage signaling and inhibited autophagy activation by modulating mTOR, LC3 and AMBRA1 with probable dynein involvement. Interestingly, one possible role for dynein in the mechanism of action of Amblyomin-X was in the apoptotic response and its crosstalk with autophagy, which involved the factor Bim; however, we observed no changes in the apoptotic response related to dynein in the experiments performed. The characteristics shared among Amblyomin-X and known proteasome inhibitors included NF-κB blockage and nascent polypeptide-dependent aggresome formation. Therefore, our study describes a Kunitz-type protein that acts on the proteasome to trigger distinct intracellular events compared to classic known proteasome inhibitors that are small-cell-permeable molecules. In investigating the experiments and literature on Amblyomin-X and the known proteasome inhibitors, we also found differences in the structures of the molecules, intracellular events, dynein involvement and tumor cell type effects. These findings also reveal a possible new target for Amblyomin-X, i.e., dynein, and may serve as a tool for investigating tumor cell death associated with proteasome inhibition.
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Mario T F
authors.initials: MT
authors.lastname: Pacheco
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Carolina M
authors.initials: CM
authors.lastname: Berra
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil; Department of Biochemistry, Federal University of São Paulo, São Paulo, Brazil.
authors.forename: Kátia L P
authors.initials: KL
authors.lastname: Morais
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Juliana M
authors.initials: JM
authors.lastname: Sciani
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Vania G
authors.initials: VG
authors.lastname: Branco
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Rosemary V
authors.initials: RV
authors.lastname: Bosch
authors.affiliation: Biochemistry and Biophysics Laboratory, Butantan Institute, São Paulo, Brazil.
authors.forename: Ana M
authors.initials: AM
authors.lastname: Chudzinski-Tavassi
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D060829:Arthropod Proteins; D001343:Autophagy / Q000187:drug effects; D045744:Cell Line, Tumor; D004398:Dyneins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D026863:Ixodidae / Q000737:chemistry / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects; D061988:Proteasome Inhibitors / Q000008:administration & dosage* / Q000378:metabolism; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D012471:Salivary Proteins and Peptides / Q000008:administration & dosage* / Q000235:genetics / Q000378:metabolism
chemical_list: C559369:Amblyomin-X protein, Amblyomma cajennense; D060829:Arthropod Proteins; D061988:Proteasome Inhibitors; D011994:Recombinant Proteins; D012471:Salivary Proteins and Peptides; D046988:Proteasome Endopeptidase Complex; D004398:Dyneins