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The adhesion force of Notch with Delta and the rate of Notch signaling.

Ahimou F, Mok LP, Bardot B, Wesley C - J. Cell Biol. (2004)

Bottom Line: Notch signaling is repeatedly used during animal development to specify cell fates.Reduced turnover or Delta pulling accelerate this loss.These data suggest that strong adhesion between Notch and Delta might serve as a booster for initiating Notch signaling at a high rate.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, The University of Vermont, VT 05405, USA.

ABSTRACT
Notch signaling is repeatedly used during animal development to specify cell fates. Using atomic force microscopy on live cells, chemical inhibitors, and conventional analyses, we show that the rate of Notch signaling is linked to the adhesion force between cells expressing Notch receptors and Delta ligand. Both the Notch extracellular and intracellular domains are required for the high adhesion force with Delta. This high adhesion force is lost within minutes, primarily due to the action of Presenilin on Notch. Reduced turnover or Delta pulling accelerate this loss. These data suggest that strong adhesion between Notch and Delta might serve as a booster for initiating Notch signaling at a high rate.

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Presenilin inhibitor blocks the loss of detachment force between Notch receptors and Dl. (A) Detachment forces between the S2-Dl cantilevers and 1% DMSO-pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (B) Detachment force between the S2-Dl cantilevers and 1% DMSO + Psn inhibitor–pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (C) A Western blot showing the amounts of Notch in the presence or absence of the Psn inhibitor. (D) A Western blot showing the amounts of NΔB molecules in the absence or presence of 5× Psn inhibitor.
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fig6: Presenilin inhibitor blocks the loss of detachment force between Notch receptors and Dl. (A) Detachment forces between the S2-Dl cantilevers and 1% DMSO-pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (B) Detachment force between the S2-Dl cantilevers and 1% DMSO + Psn inhibitor–pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (C) A Western blot showing the amounts of Notch in the presence or absence of the Psn inhibitor. (D) A Western blot showing the amounts of NΔB molecules in the absence or presence of 5× Psn inhibitor.

Mentions: To determine if the decrease in detachment force was indeed due to Psn cleavage, we treated the S2 cells expressing various Notch receptors with DFK-167 Psn inhibitor for 30 min, washed the cells, and then measured the detachment force with the S2-Dl cantilever. DFK-167 is dissolved in DMSO. So, we first measured the temporal change in detachment force between the S2-Dl cantilever and the S2 cells expressing the various Notch receptors that were pretreated with the same concentration of DMSO used with the Psn inhibitor. The level of adhesion force at all time points was lower and the rate of decrease to zero was retarded (particularly with S2-N cells), possibly due to the effect of DMSO on the plasma membranes (compare Fig. 6 A with Fig. 5 A). When the S2 cells expressing various Notch receptors were treated with DMSO + Psn inhibitor, the decrease of the detachment force to zero was blocked with N, N1-2155, Nnd3, and Nmf receptors (Fig. 6 B). The detachment forces between the S2-Dl cantilevers and S2-Dl cells or S2 cells treated with the Psn inhibitor did not increase (unpublished data). Also, when Psn inhibitor–treated S2-Dl cells were used on the cantilever and tested against untreated S2-N cells, the loss in detachment force was not blocked (see later, Fig. 8 E). These observations indicated that the block in the decrease in detachment force was due to the Psn inhibitor affecting N rather than some other general cell adhesion molecules. Western blotting showed that the level of the S2-cleaved N fragment in S2-N cells increased after treatments with the Psn inhibitor, even in the absence of Dl; in the presence of Dl, the levels of the S2 fragment further increased (2.6 ± 0.24 at 120 min, P < 0.05, n = 4) in association with reduced levels of the S3 fragment (Fig. 6 C). Although the effect of the Psn inhibitor was apparent even at 10 min of aggregation, data for longer periods are shown to indicate the robustness of the effect.


The adhesion force of Notch with Delta and the rate of Notch signaling.

Ahimou F, Mok LP, Bardot B, Wesley C - J. Cell Biol. (2004)

Presenilin inhibitor blocks the loss of detachment force between Notch receptors and Dl. (A) Detachment forces between the S2-Dl cantilevers and 1% DMSO-pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (B) Detachment force between the S2-Dl cantilevers and 1% DMSO + Psn inhibitor–pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (C) A Western blot showing the amounts of Notch in the presence or absence of the Psn inhibitor. (D) A Western blot showing the amounts of NΔB molecules in the absence or presence of 5× Psn inhibitor.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172611&req=5

fig6: Presenilin inhibitor blocks the loss of detachment force between Notch receptors and Dl. (A) Detachment forces between the S2-Dl cantilevers and 1% DMSO-pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (B) Detachment force between the S2-Dl cantilevers and 1% DMSO + Psn inhibitor–pretreated S2 cells expressing Notch receptors, in 1× PBS+Ca2+. (C) A Western blot showing the amounts of Notch in the presence or absence of the Psn inhibitor. (D) A Western blot showing the amounts of NΔB molecules in the absence or presence of 5× Psn inhibitor.
Mentions: To determine if the decrease in detachment force was indeed due to Psn cleavage, we treated the S2 cells expressing various Notch receptors with DFK-167 Psn inhibitor for 30 min, washed the cells, and then measured the detachment force with the S2-Dl cantilever. DFK-167 is dissolved in DMSO. So, we first measured the temporal change in detachment force between the S2-Dl cantilever and the S2 cells expressing the various Notch receptors that were pretreated with the same concentration of DMSO used with the Psn inhibitor. The level of adhesion force at all time points was lower and the rate of decrease to zero was retarded (particularly with S2-N cells), possibly due to the effect of DMSO on the plasma membranes (compare Fig. 6 A with Fig. 5 A). When the S2 cells expressing various Notch receptors were treated with DMSO + Psn inhibitor, the decrease of the detachment force to zero was blocked with N, N1-2155, Nnd3, and Nmf receptors (Fig. 6 B). The detachment forces between the S2-Dl cantilevers and S2-Dl cells or S2 cells treated with the Psn inhibitor did not increase (unpublished data). Also, when Psn inhibitor–treated S2-Dl cells were used on the cantilever and tested against untreated S2-N cells, the loss in detachment force was not blocked (see later, Fig. 8 E). These observations indicated that the block in the decrease in detachment force was due to the Psn inhibitor affecting N rather than some other general cell adhesion molecules. Western blotting showed that the level of the S2-cleaved N fragment in S2-N cells increased after treatments with the Psn inhibitor, even in the absence of Dl; in the presence of Dl, the levels of the S2 fragment further increased (2.6 ± 0.24 at 120 min, P < 0.05, n = 4) in association with reduced levels of the S3 fragment (Fig. 6 C). Although the effect of the Psn inhibitor was apparent even at 10 min of aggregation, data for longer periods are shown to indicate the robustness of the effect.

Bottom Line: Notch signaling is repeatedly used during animal development to specify cell fates.Reduced turnover or Delta pulling accelerate this loss.These data suggest that strong adhesion between Notch and Delta might serve as a booster for initiating Notch signaling at a high rate.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Molecular Genetics, The University of Vermont, VT 05405, USA.

ABSTRACT
Notch signaling is repeatedly used during animal development to specify cell fates. Using atomic force microscopy on live cells, chemical inhibitors, and conventional analyses, we show that the rate of Notch signaling is linked to the adhesion force between cells expressing Notch receptors and Delta ligand. Both the Notch extracellular and intracellular domains are required for the high adhesion force with Delta. This high adhesion force is lost within minutes, primarily due to the action of Presenilin on Notch. Reduced turnover or Delta pulling accelerate this loss. These data suggest that strong adhesion between Notch and Delta might serve as a booster for initiating Notch signaling at a high rate.

Show MeSH
Related in: MedlinePlus