title: Perturbation of cellular proteostasis networks identifies pathways that modulate precursor and intermediate but not mature levels of frataxin.
abstract: Friedreich's Ataxia is a genetic disease caused by expansion of an intronic trinucleotide repeat in the frataxin (FXN) gene yielding diminished FXN expression and consequently disease. Since increasing FXN protein levels is desirable to ameliorate pathology, we explored the role of major cellular proteostasis pathways and mitochondrial proteases in FXN processing and turnover. We targeted p97/VCP, the ubiquitin proteasome pathway (UPP), and autophagy with chemical inhibitors in cell lines and patient-derived cells. p97 inhibition by DBeQ increased precursor FXN levels, while UPP and autophagic flux modulators had variable effects predominantly on intermediate FXN. Our data suggest that these pathways cannot be modulated to influence mature functional FXN levels. We also targeted known mitochondrial proteases by RNA interference and discovered a novel protease PITRM1 that regulates intermediate FXN levels. Treatment with the aforementioned chemical and genetic modulators did not have a differential effect in patient cells containing lower amounts of FXN. Interestingly, a number of treatments caused a change in total amount of FXN protein, without an effect on mature FXN. Our results imply that regulation of FXN protein levels is complex and that total amounts can be modulated chemically and genetically without altering the absolute amount of mature FXN protein.
authors.affiliation: Rare Disease Research Unit, Worldwide Research and Development, Pfizer, 610 Main Street, Cambridge, MA 02139, USA.
authors.forename: Joseph F
authors.initials: JF
authors.lastname: Nabhan
authors.affiliation: Rare Disease Research Unit, Worldwide Research and Development, Pfizer, 610 Main Street, Cambridge, MA 02139, USA.
authors.forename: Renea L
authors.initials: RL
authors.lastname: Gooch
authors.affiliation: Worldwide Medicinal Chemistry, Pfizer, 610 Main Street, Cambridge, MA 02139, USA.
authors.forename: Eugene L
authors.initials: EL
authors.lastname: Piatnitski Chekler
authors.affiliation: Worldwide Medicinal Chemistry, Pfizer, Eastern Point Road, Groton, CT 06340, USA.
authors.forename: Betsy
authors.initials: B
authors.lastname: Pierce
authors.affiliation: Rare Disease Research Unit, Worldwide Research and Development, Pfizer, 610 Main Street, Cambridge, MA 02139, USA.
authors.forename: Christine E
authors.initials: CE
authors.lastname: Bulawa
mesh_terms: D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D005621:Friedreich Ataxia / Q000235:genetics / Q000378:metabolism; D006801:Humans; D033862:Iron-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D024101:Mitochondrial Proteins / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D060066:Protein Interaction Maps; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D059748:Proteolysis; D011799:Quinazolines / Q000494:pharmacology; D015398:Signal Transduction; D019680:Trinucleotide Repeat Expansion; D025801:Ubiquitin / Q000378:metabolism; D000074405:Valosin Containing Protein; D000096923:Frataxin
chemical_list: D000251:Adenosine Triphosphatases; D018797:Cell Cycle Proteins; D033862:Iron-Binding Proteins; D024101:Mitochondrial Proteins; D046988:Proteasome Endopeptidase Complex; D011498:Protein Precursors; D011799:Quinazolines; D025801:Ubiquitin; D000074405:Valosin Containing Protein; D000096923:Frataxin; C558357:N2,N4-dibenzylquinazoline-2,4-diamine; C000614178:VCP protein, human