Supplemental Data Nicastrin Functions as a γ-Secretase-Substrate Receptor

Center for Basic Neuroscience, The University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.
Cell (Impact Factor: 32.24). 09/2005; 122(3):435-47. DOI: 10.1016/j.cell.2005.05.022
Source: PubMed


gamma-secretase catalyzes the intramembrane cleavage of amyloid precursor protein (APP) and Notch after their extracellular domains are shed by site-specific proteolysis. Nicastrin is an essential glycoprotein component of the gamma-secretase complex but has no known function. We now show that the ectodomain of nicastrin binds the new amino terminus that is generated upon proteolysis of the extracellular APP and Notch domains, thereby recruiting the APP and Notch substrates into the gamma-secretase complex. Chemical- or antibody-mediated blocking of the free amino terminus, addition of purified nicastrin ectodomain, or mutations in the ectodomain markedly reduce the binding and cleavage of substrate by gamma-secretase. These results indicate that nicastrin is a receptor for the amino-terminal stubs that are generated by ectodomain shedding of type I transmembrane proteins. Our data are consistent with a model where nicastrin presents these substrates to gamma-secretase and thereby facilitates their cleavage via intramembrane proteolysis.

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    • "As such, we generated adipocyte-specific Nicastrin (A-Nicastrin) knockout mice, which demonstrated lower Nicastrin mRNA and protein levels in both eWAT and iWAT (Figure 5A, B) with specific reductions in the adipocyte fractions of adipose tissue from these mice (Figure 5C). Expectedly, given the necessity of Nicastrin for g-secretase stability and activity[28,29], C-terminal fragment levels of Presenilin 1 and 2 were lower in A-Nicastrin mice (not shown and Figure 5B). A- Nicastrin mice were born at Mendelian frequency, without gross developmental phenotype and unchanged body weight and adiposity with chow-(Supplemental Figure 4A, B) or HFD-feeding (Figure 5DeF), but in contrast to A-Rbpj mice, A-Nicastrin mice showed a trend towards reduced glucose intolerance and insulin sensitivity as compared to Cre-controls (Supplemental Figure 4C, G, H). "
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    ABSTRACT: As the obesity pandemic continues to expand, novel molecular targets to reduce obesity-related insulin resistance and Type 2 Diabetes (T2D) continue to be needed. We have recently shown that obesity is associated with reactivated liver Notch signaling, which, in turn, increases hepatic insulin resistance, opening up therapeutic avenues for Notch inhibitors to be repurposed for T2D. Herein, we tested the systemic effects of γ-secretase inhibitors (GSIs), which prevent endogenous Notch activation, and confirmed these effects through creation and characterization of two different adipocyte-specific Notch loss-of-function mouse models through genetic ablation of the Notch transcriptional effector Rbp-Jk (A-Rbpj) and the obligate γ-secretase component Nicastrin (A-Nicastrin).
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    • "Nicastrin is a type I transmembrane glycoprotein considered to be the scaffolding protein within the γ-secretase complex. Nicastrin may also act as the γ-secretase receptor [144]. The seven-transmembrane APH-1 interacts with nicastrin to form a stable intermediate in an early assembly stage of the γ-secretase complex [140,141], whereas the two-pass transmembrane component PEN2 regulates PS endoproteolysis [145,146]. "
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    • "Such a two-step process has been observed for other substrates of γ-secretase including L1 [26], protein-tyrosine kinase 7 (PTK7) [55] and the epidermal growth factor-like betacellulin precursor [56]. This sequential processing appears to be a general mechanism for γ-secretase substrates with permissive cytoplasmic and transmembrane domains [57], allowing for nicastrin, a member of the γ-secretase complex, to recognize RIP substrates [58]. "
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