2. Academic Validation
  3. Individual and combined presenilin 1 and 2 knockouts reveal that both have highly overlapping functions in HEK293T cells

Individual and combined presenilin 1 and 2 knockouts reveal that both have highly overlapping functions in HEK293T cells

  • J Biol Chem. 2019 Jul 19;294(29):11276-11285. doi: 10.1074/jbc.RA119.008041.
Christian B Lessard 1,  Edgardo Rodriguez 2,  Thomas B Ladd 1,  Lisa M Minter 3,  Barbara A Osborne 3,  Lucio Miele 4,  Todd E Golde 5,  Yong Ran 6
Affiliations

Affiliations

  • 1 Department of Neuroscience and Neurology, Center for Translational Research in Neurodegenerative Disease, and McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, Florida 32610.
  • 2 Department of Pharmacology and Therapeutics, Center for Translational Research in Neurodegenerative Disease, and McKnight Brain Institute, University of Florida, Gainesville, Florida 32610.
  • 3 Department of Veterinary and Animal Sciences, Center for Bioactive Delivery, Institute for Applied Life Sciences, and Program in Molecular and Cellular Biology, University of Massachusetts, Amherst, Massachusetts 01003.
  • 4 Department of Genetics and Stanley S. Scott Cancer Center, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70112.
  • 5 Department of Neuroscience and Neurology, Center for Translational Research in Neurodegenerative Disease, and McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, Florida 32610 tgolde@ufl.edu.
  • 6 Department of Neuroscience and Neurology, Center for Translational Research in Neurodegenerative Disease, and McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, Florida 32610 yran@ufl.edu.
PMID: 31167792 DOI: 10.1074/jbc.RA119.008041
Abstract

Presenilins 1 and 2 (PS1 and 2) are the catalytic subunits of γ-secretase, a multiprotein Protease that cleaves amyloid protein precursor and other type I transmembrane proteins. Previous studies with mouse models or cells have indicated differences in PS1 and PS2 functions. We have recently reported that clinical γ-secretase inhibitors (GSIs), initially developed to manage Alzheimer's disease and now being considered for other therapeutic interventions, are both pharmacologically and functionally distinct. Here, using CRISPR/Cas9-based gene editing, we established human HEK 293T cell lines in which endogenous PS1, PS2, or both have been knocked out. Using these knockout lines to examine differences in PS1- and PS2-mediated cleavage events, we confirmed that PS2 generates more intracellular β-amyloid than does PS1. Moreover, we observed subtle differences in PS1- and PS2-mediated cleavages of select substrates. In exploring the question of whether differences in activity among clinical GSIs could be attributed to differential inhibition of PS1 or PS2, we noted that select GSIs inhibit PS1 and PS2 activities on specific substrates with slightly different potencies. We also found that endoproteolysis of select PS1 FAD-linked variants in human cells is more efficient than what has been previously reported for mouse cell lines. Overall, these results obtained with HEK293T cells suggest that selective PS1 or PS2 inhibition by a given GSI does not explain the previously observed differences in functional and pharmacological properties among various GSIs.

Keywords

NOTCH; Notch protein; amyloid protein precursor; amyloid-beta (AB); aspartyl protease; cancer; inhibitor; neurodegeneration; presenilin; γ-secretase.

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