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SNX17 regulates Notch pathway and pancreas development through the retromer-dependent recycling of Jag1.

Yin W, Liu D, Liu N, Xu L, Li S, Lin S, Shu X, Pei D - Cell Regen (Lond) (2012)

Bottom Line: SNX17 is a sorting nexin family protein implicated in vesicular trafficking and we find it is specifically required in the ligand-expressing cells for Notch signaling.Mechanistically, SNX17 regulates the protein level of Jag1a on plasma membrane by binding to Jag1a and facilitating the retromer-dependent recycling of the ligand.In zebrafish, inhibition of this SNX17-mediated Notch signaling pathway results in defects in neurogenesis as well as pancreas development.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Regenerative Biology, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530 China.

ABSTRACT

Background: Notch is one of the most important signaling pathways involved in cell fate determination. Activation of the Notch pathway requires the binding of a membrane-bound ligand to the Notch receptor in the adjacent cell which induces proteolytic cleavages and the activation of the receptor. A unique feature of the Notch signaling is that processes such as modification, endocytosis or recycling of the ligand have been reported to play critical roles during Notch signaling, however, the underlying molecular mechanism appears context-dependent and often controversial.

Results: Here we identified SNX17 as a novel regulator of the Notch pathway. SNX17 is a sorting nexin family protein implicated in vesicular trafficking and we find it is specifically required in the ligand-expressing cells for Notch signaling. Mechanistically, SNX17 regulates the protein level of Jag1a on plasma membrane by binding to Jag1a and facilitating the retromer-dependent recycling of the ligand. In zebrafish, inhibition of this SNX17-mediated Notch signaling pathway results in defects in neurogenesis as well as pancreas development.

Conclusions: Our results reveal that SNX17, by acting as a cargo-specific adaptor, promotes the retromer dependent recycling of Jag1a and Notch signaling and this pathway is involved in cell fate determination during zebrafish neurogenesis and pancreas development.

No MeSH data available.


Related in: MedlinePlus

SNX17 regulates the protein level of plasma membrane associated Jag1a. (A) Knockdown of SNX17 reduced the total as well as plasma membrane associated Jag1a protein levels while over-expression of SNX17 enhanced the protein levels of Jag1a. Plasma membrane proteins were labeled with biotin and isolated with the Pierce Cell Surface Protein Isolation Kit and detected by western blot. GAPDH and TFR were the loading control. (B) SNX17 was not required for the endocytosis of Jag1a. Plasma membrane proteins were labeled with biotin and allowed to be endocytosed for 1 h. The remaining cell surface biotin was stripped and the internalized biotin-labeled proteins were immunoprecipitated then detected by western blot. (C) Subcellular distribution of Jag1a. The majority of intracellular Jag1a was detected at the Hrs-positive vesicles.
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Fig4: SNX17 regulates the protein level of plasma membrane associated Jag1a. (A) Knockdown of SNX17 reduced the total as well as plasma membrane associated Jag1a protein levels while over-expression of SNX17 enhanced the protein levels of Jag1a. Plasma membrane proteins were labeled with biotin and isolated with the Pierce Cell Surface Protein Isolation Kit and detected by western blot. GAPDH and TFR were the loading control. (B) SNX17 was not required for the endocytosis of Jag1a. Plasma membrane proteins were labeled with biotin and allowed to be endocytosed for 1 h. The remaining cell surface biotin was stripped and the internalized biotin-labeled proteins were immunoprecipitated then detected by western blot. (C) Subcellular distribution of Jag1a. The majority of intracellular Jag1a was detected at the Hrs-positive vesicles.

Mentions: Previous studies indicate that SNX17 is able to regulate the levels of plasma membrane proteins either through the endocytosis or the recycling process. We investigated whether SNX17 regulated the homeostasis of Jag1a by comparing the protein levels of total as well as the plasma membrane associated Jag1a between the control and siSNX17 treated samples. The plasma membrane proteins were first labeled by biotin then isolated with the Pierce Cell Surface Protein Isolation Kit and Jag1a level was determined by western blot. As shown in Figure 4A, Jag1a level in either whole cell lysate or membrane fraction was clearly reduced when SNX17 was knocked-down by siRNAs. On the other hand, over-expression of SNX17 increased Jag1a protein level. As a control, the protein level of the plasma membrane associated transferrin receptor (TFR) was not affected by either knockdown or over-expression of SNX17.Figure 4


SNX17 regulates Notch pathway and pancreas development through the retromer-dependent recycling of Jag1.

Yin W, Liu D, Liu N, Xu L, Li S, Lin S, Shu X, Pei D - Cell Regen (Lond) (2012)

SNX17 regulates the protein level of plasma membrane associated Jag1a. (A) Knockdown of SNX17 reduced the total as well as plasma membrane associated Jag1a protein levels while over-expression of SNX17 enhanced the protein levels of Jag1a. Plasma membrane proteins were labeled with biotin and isolated with the Pierce Cell Surface Protein Isolation Kit and detected by western blot. GAPDH and TFR were the loading control. (B) SNX17 was not required for the endocytosis of Jag1a. Plasma membrane proteins were labeled with biotin and allowed to be endocytosed for 1 h. The remaining cell surface biotin was stripped and the internalized biotin-labeled proteins were immunoprecipitated then detected by western blot. (C) Subcellular distribution of Jag1a. The majority of intracellular Jag1a was detected at the Hrs-positive vesicles.
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Related In: Results  -  Collection

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Fig4: SNX17 regulates the protein level of plasma membrane associated Jag1a. (A) Knockdown of SNX17 reduced the total as well as plasma membrane associated Jag1a protein levels while over-expression of SNX17 enhanced the protein levels of Jag1a. Plasma membrane proteins were labeled with biotin and isolated with the Pierce Cell Surface Protein Isolation Kit and detected by western blot. GAPDH and TFR were the loading control. (B) SNX17 was not required for the endocytosis of Jag1a. Plasma membrane proteins were labeled with biotin and allowed to be endocytosed for 1 h. The remaining cell surface biotin was stripped and the internalized biotin-labeled proteins were immunoprecipitated then detected by western blot. (C) Subcellular distribution of Jag1a. The majority of intracellular Jag1a was detected at the Hrs-positive vesicles.
Mentions: Previous studies indicate that SNX17 is able to regulate the levels of plasma membrane proteins either through the endocytosis or the recycling process. We investigated whether SNX17 regulated the homeostasis of Jag1a by comparing the protein levels of total as well as the plasma membrane associated Jag1a between the control and siSNX17 treated samples. The plasma membrane proteins were first labeled by biotin then isolated with the Pierce Cell Surface Protein Isolation Kit and Jag1a level was determined by western blot. As shown in Figure 4A, Jag1a level in either whole cell lysate or membrane fraction was clearly reduced when SNX17 was knocked-down by siRNAs. On the other hand, over-expression of SNX17 increased Jag1a protein level. As a control, the protein level of the plasma membrane associated transferrin receptor (TFR) was not affected by either knockdown or over-expression of SNX17.Figure 4

Bottom Line: SNX17 is a sorting nexin family protein implicated in vesicular trafficking and we find it is specifically required in the ligand-expressing cells for Notch signaling.Mechanistically, SNX17 regulates the protein level of Jag1a on plasma membrane by binding to Jag1a and facilitating the retromer-dependent recycling of the ligand.In zebrafish, inhibition of this SNX17-mediated Notch signaling pathway results in defects in neurogenesis as well as pancreas development.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Regenerative Biology, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530 China.

ABSTRACT

Background: Notch is one of the most important signaling pathways involved in cell fate determination. Activation of the Notch pathway requires the binding of a membrane-bound ligand to the Notch receptor in the adjacent cell which induces proteolytic cleavages and the activation of the receptor. A unique feature of the Notch signaling is that processes such as modification, endocytosis or recycling of the ligand have been reported to play critical roles during Notch signaling, however, the underlying molecular mechanism appears context-dependent and often controversial.

Results: Here we identified SNX17 as a novel regulator of the Notch pathway. SNX17 is a sorting nexin family protein implicated in vesicular trafficking and we find it is specifically required in the ligand-expressing cells for Notch signaling. Mechanistically, SNX17 regulates the protein level of Jag1a on plasma membrane by binding to Jag1a and facilitating the retromer-dependent recycling of the ligand. In zebrafish, inhibition of this SNX17-mediated Notch signaling pathway results in defects in neurogenesis as well as pancreas development.

Conclusions: Our results reveal that SNX17, by acting as a cargo-specific adaptor, promotes the retromer dependent recycling of Jag1a and Notch signaling and this pathway is involved in cell fate determination during zebrafish neurogenesis and pancreas development.

No MeSH data available.


Related in: MedlinePlus