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A Novel NIR-FRET Biosensor for Reporting PS/γ-Secretase Activity in Live Cells.
Houser, Mei Cq; Hou, Steven S; Perrin, Florian; Turchyna, Yuliia; Bacskai, Brian J; Berezovska, Oksana; Maesako, Masato.
Afiliación
  • Houser MC; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Hou SS; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Perrin F; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Turchyna Y; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Bacskai BJ; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Berezovska O; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
  • Maesako M; Alzheimer Research Unit, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital, Harvard Medical School, 114, 16th street, Charlestown, MA 02129, USA.
Sensors (Basel) ; 20(21)2020 Oct 22.
Article en En | MEDLINE | ID: mdl-33105735
Presenilin (PS)/γ-secretase plays a pivotal role in essential cellular events via proteolytic processing of transmembrane proteins that include APP and Notch receptors. However, how PS/γ-secretase activity is spatiotemporally regulated by other molecular and cellular factors and how the changes in PS/γ-secretase activity influence signaling pathways in live cells are poorly understood. These questions could be addressed by engineering a new tool that enables multiplexed imaging of PS/γ-secretase activity and additional cellular events in real-time. Here, we report the development of a near-infrared (NIR) FRET-based PS/γ-secretase biosensor, C99 720-670 probe, which incorporates an immediate PS/γ-secretase substrate APP C99 with miRFP670 and miRFP720 as the donor and acceptor fluorescent proteins, respectively. Extensive validation demonstrates that the C99 720-670 biosensor enables quantitative monitoring of endogenous PS/γ-secretase activity on a cell-by-cell basis in live cells (720/670 ratio: 2.47 ± 0.66 (vehicle) vs. 3.02 ± 1.17 (DAPT), ** p < 0.01). Importantly, the C99 720-670 and the previously developed APP C99 YPet-Turquoise-GL (C99 Y-T) biosensors simultaneously report PS/γ-secretase activity. This evidences the compatibility of the C99 720-670 biosensor with cyan (CFP)-yellow fluorescent protein (YFP)-based FRET biosensors for reporting other essential cellular events. Multiplexed imaging using the novel NIR biosensor C99 720-670 would open a new avenue to better understand the regulation and consequences of changes in PS/γ-secretase activity.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Técnicas Biosensibles / Transferencia Resonante de Energía de Fluorescencia / Secretasas de la Proteína Precursora del Amiloide / Presenilinas Límite: Humans Idioma: En Revista: Sensors (Basel) Año: 2020 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Suiza
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Técnicas Biosensibles / Transferencia Resonante de Energía de Fluorescencia / Secretasas de la Proteína Precursora del Amiloide / Presenilinas Límite: Humans Idioma: En Revista: Sensors (Basel) Año: 2020 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Suiza