Chimeric tRNAs as tools to induce proteome damage and identify components of stress responses.
Misfolded proteins are caused by genomic mutations, aberrant splicing events, translation errors or environmental factors. The accumulation of misfolded proteins is a phenomenon connected to several human disorders, and is managed by stress responses specific to the cellular compartments being affected. In wild-type cells these mechanisms of stress response can be experimentally induced by expressing recombinant misfolded proteins or by incubating cells with large concentrations of amino acid analogues. Here, we report a novel approach for the induction of stress responses to protein aggregation. Our method is based on engineered transfer RNAs that can be expressed in cells or tissues, where they actively integrate in the translation machinery causing general proteome substitutions. This strategy allows for the introduction of mutations of increasing severity randomly in the proteome, without exposing cells to unnatural compounds. Here, we show that this approach can be used for the differential activation of the stress response in the Endoplasmic Reticulum (ER). As an example of the applications of this method, we have applied it to the identification of human microRNAs activated or repressed during unfolded protein stress.
Geslain
Renaud
R
Institute for Research in Biomedicine, Omnia Molecular, Barcelona Science Park, Instituto de Biología Molecular de Barcelona, CSIC, Parc Cientific de Barcelona, c/Baldiri Reixac 15-21, Barcelona 08028, Spain.
Cubells
Laia
L
Bori-Sanz
Teresa
T
Alvarez-Medina
Roberto
R
Rossell
David
D
Martí
Elisa
E
Ribas de Pouplana
Lluís
L
D000818:Animals; D048708:Cell Growth Processes; D002460:Cell Line; D002470:Cell Survival; D002642:Chick Embryo; D003627:Data Interpretation, Statistical; D006801:Humans; D035683:MicroRNAs / Q000145:classification / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D014176:Protein Biosynthesis; D020543:Proteome / Q000235:genetics*; D012362:RNA, Transfer, Ser / Q000737:chemistry* / Q000378:metabolism; D056811:Unfolded Protein Response / Q000235:genetics*
D035683:MicroRNAs; D020543:Proteome; D012362:RNA, Transfer, Ser