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Context-Dependent Functional Divergence of the Notch Ligands DLL1 and DLL4 In Vivo.

Preuße K, Tveriakhina L, Schuster-Gossler K, Gaspar C, Rosa AI, Henrique D, Gossler A, Stauber M - PLoS Genet. (2015)

Bottom Line: In the anterior PSM, every cell expresses both Notch receptors and ligands, and DLL1 is the only activator of Notch while DLL4 is not endogenously expressed.Testing several aspects of the complex Notch signalling system in vitro, we found that both ligands have a similar trans-activation potential but that only DLL4 is an efficient cis-inhibitor of Notch signalling, causing a reduced net activation of Notch.These differential cis-inhibitory properties are likely to contribute to the functional divergence of DLL1 and DLL4.

View Article: PubMed Central - PubMed

Affiliation: Institut für Molekularbiologie OE5250, Medizinische Hochschule Hannover, Hannover, Germany.

ABSTRACT
Notch signalling is a fundamental pathway that shapes the developing embryo and sustains adult tissues by direct communication between ligand and receptor molecules on adjacent cells. Among the ligands are two Delta paralogues, DLL1 and DLL4, that are conserved in mammals and share a similar structure and sequence. They activate the Notch receptor partly in overlapping expression domains where they fulfil redundant functions in some processes (e.g. maintenance of the crypt cell progenitor pool). In other processes, however, they appear to act differently (e.g. maintenance of foetal arterial identity) raising the questions of how similar DLL1 and DLL4 really are and which mechanism causes the apparent context-dependent divergence. By analysing mice that conditionally overexpress DLL1 or DLL4 from the same genomic locus (Hprt) and mice that express DLL4 instead of DLL1 from the endogenous Dll1 locus (Dll1Dll4ki), we found functional differences that are tissue-specific: while DLL1 and DLL4 act redundantly during the maintenance of retinal progenitors, their function varies in the presomitic mesoderm (PSM) where somites form in a Notch-dependent process. In the anterior PSM, every cell expresses both Notch receptors and ligands, and DLL1 is the only activator of Notch while DLL4 is not endogenously expressed. Transgenic DLL4 cannot replace DLL1 during somitogenesis and in heterozygous Dll1Dll4ki/+ mice, the Dll1Dll4ki allele causes a dominant segmentation phenotype. Testing several aspects of the complex Notch signalling system in vitro, we found that both ligands have a similar trans-activation potential but that only DLL4 is an efficient cis-inhibitor of Notch signalling, causing a reduced net activation of Notch. These differential cis-inhibitory properties are likely to contribute to the functional divergence of DLL1 and DLL4.

No MeSH data available.


Related in: MedlinePlus

Model of Notch signalling in the PSM triggered by DLL1 and ectopic DLL4.Summary combining our in vivo and in vitro data in three different genetic scenarios (A-C); trans-activation (green arrows) and cis-inhibition (red bars) in cells of the PSM are schematically depicted on the left, representative skeletal preparations to visualise the outcome of somitogenesis are shown on the right; references to Figs. in this paper are given below. (A) In wildtype and Dll1Dll1ki/Dll1ki PSMs, endogenous or transgenic DLL1 (D1) trans-activates Notch (N) signalling and results in a regularly segmented axial skeleton. (B) In our in vitro assays, DLL4 (D4) trans-activates Notch with similar efficiency as DLL1 but has an additional strong cis-inhibitory effect on Notch signalling that partially overrides trans-activation. The reduced net Notch activation in Dll1Dll4ki/Dll4ki and CAG:Dll4;Dll1loxP/loxP;T(s):Cre PSMs is insufficient to support normal segmentation. (C) When both DLL1 and DLL4 are expressed (Dll1Dll4ki/+ PSM), cis-inhibition by DLL4 plays a relatively smaller role, the resulting axial skeletons are mostly regular. However, cis-inhibition by DLL4 reduces the robustness of Notch signalling resulting in minor malformations (arrow indicates a misplaced rib), which are consistently seen in Dll1Dll4ki/+ skeletons.
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pgen.1005328.g006: Model of Notch signalling in the PSM triggered by DLL1 and ectopic DLL4.Summary combining our in vivo and in vitro data in three different genetic scenarios (A-C); trans-activation (green arrows) and cis-inhibition (red bars) in cells of the PSM are schematically depicted on the left, representative skeletal preparations to visualise the outcome of somitogenesis are shown on the right; references to Figs. in this paper are given below. (A) In wildtype and Dll1Dll1ki/Dll1ki PSMs, endogenous or transgenic DLL1 (D1) trans-activates Notch (N) signalling and results in a regularly segmented axial skeleton. (B) In our in vitro assays, DLL4 (D4) trans-activates Notch with similar efficiency as DLL1 but has an additional strong cis-inhibitory effect on Notch signalling that partially overrides trans-activation. The reduced net Notch activation in Dll1Dll4ki/Dll4ki and CAG:Dll4;Dll1loxP/loxP;T(s):Cre PSMs is insufficient to support normal segmentation. (C) When both DLL1 and DLL4 are expressed (Dll1Dll4ki/+ PSM), cis-inhibition by DLL4 plays a relatively smaller role, the resulting axial skeletons are mostly regular. However, cis-inhibition by DLL4 reduces the robustness of Notch signalling resulting in minor malformations (arrow indicates a misplaced rib), which are consistently seen in Dll1Dll4ki/+ skeletons.

Mentions: In summary, our cis-inhibition assays (Fig 5E) reveal a functional difference between DLL1 and DLL4 that was not evident in the trans-activation assays (Fig 5D and 5F): DLL4, but not DLL1, is a potent cis-inhibitor of NOTCH1 and cis-inhibition by DLL4 can significantly reduce Notch activation. Our in vitro results are consistent with our in vivo data: they can explain both why DLL4 appears to be a weaker activator of Notch signalling than DLL1 during somitogenesis in our transgenic mice and why transgenic DLL4 has a dominant effect on segmentation in Dll1Dll4ki/+ mice (see Discussion and Fig 6). We propose that in the PSM, DLL1 is a more efficient net activator of Notch than (ectopic) DLL4 because it does not efficiently cis-inhibit Notch.


Context-Dependent Functional Divergence of the Notch Ligands DLL1 and DLL4 In Vivo.

Preuße K, Tveriakhina L, Schuster-Gossler K, Gaspar C, Rosa AI, Henrique D, Gossler A, Stauber M - PLoS Genet. (2015)

Model of Notch signalling in the PSM triggered by DLL1 and ectopic DLL4.Summary combining our in vivo and in vitro data in three different genetic scenarios (A-C); trans-activation (green arrows) and cis-inhibition (red bars) in cells of the PSM are schematically depicted on the left, representative skeletal preparations to visualise the outcome of somitogenesis are shown on the right; references to Figs. in this paper are given below. (A) In wildtype and Dll1Dll1ki/Dll1ki PSMs, endogenous or transgenic DLL1 (D1) trans-activates Notch (N) signalling and results in a regularly segmented axial skeleton. (B) In our in vitro assays, DLL4 (D4) trans-activates Notch with similar efficiency as DLL1 but has an additional strong cis-inhibitory effect on Notch signalling that partially overrides trans-activation. The reduced net Notch activation in Dll1Dll4ki/Dll4ki and CAG:Dll4;Dll1loxP/loxP;T(s):Cre PSMs is insufficient to support normal segmentation. (C) When both DLL1 and DLL4 are expressed (Dll1Dll4ki/+ PSM), cis-inhibition by DLL4 plays a relatively smaller role, the resulting axial skeletons are mostly regular. However, cis-inhibition by DLL4 reduces the robustness of Notch signalling resulting in minor malformations (arrow indicates a misplaced rib), which are consistently seen in Dll1Dll4ki/+ skeletons.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4482573&req=5

pgen.1005328.g006: Model of Notch signalling in the PSM triggered by DLL1 and ectopic DLL4.Summary combining our in vivo and in vitro data in three different genetic scenarios (A-C); trans-activation (green arrows) and cis-inhibition (red bars) in cells of the PSM are schematically depicted on the left, representative skeletal preparations to visualise the outcome of somitogenesis are shown on the right; references to Figs. in this paper are given below. (A) In wildtype and Dll1Dll1ki/Dll1ki PSMs, endogenous or transgenic DLL1 (D1) trans-activates Notch (N) signalling and results in a regularly segmented axial skeleton. (B) In our in vitro assays, DLL4 (D4) trans-activates Notch with similar efficiency as DLL1 but has an additional strong cis-inhibitory effect on Notch signalling that partially overrides trans-activation. The reduced net Notch activation in Dll1Dll4ki/Dll4ki and CAG:Dll4;Dll1loxP/loxP;T(s):Cre PSMs is insufficient to support normal segmentation. (C) When both DLL1 and DLL4 are expressed (Dll1Dll4ki/+ PSM), cis-inhibition by DLL4 plays a relatively smaller role, the resulting axial skeletons are mostly regular. However, cis-inhibition by DLL4 reduces the robustness of Notch signalling resulting in minor malformations (arrow indicates a misplaced rib), which are consistently seen in Dll1Dll4ki/+ skeletons.
Mentions: In summary, our cis-inhibition assays (Fig 5E) reveal a functional difference between DLL1 and DLL4 that was not evident in the trans-activation assays (Fig 5D and 5F): DLL4, but not DLL1, is a potent cis-inhibitor of NOTCH1 and cis-inhibition by DLL4 can significantly reduce Notch activation. Our in vitro results are consistent with our in vivo data: they can explain both why DLL4 appears to be a weaker activator of Notch signalling than DLL1 during somitogenesis in our transgenic mice and why transgenic DLL4 has a dominant effect on segmentation in Dll1Dll4ki/+ mice (see Discussion and Fig 6). We propose that in the PSM, DLL1 is a more efficient net activator of Notch than (ectopic) DLL4 because it does not efficiently cis-inhibit Notch.

Bottom Line: In the anterior PSM, every cell expresses both Notch receptors and ligands, and DLL1 is the only activator of Notch while DLL4 is not endogenously expressed.Testing several aspects of the complex Notch signalling system in vitro, we found that both ligands have a similar trans-activation potential but that only DLL4 is an efficient cis-inhibitor of Notch signalling, causing a reduced net activation of Notch.These differential cis-inhibitory properties are likely to contribute to the functional divergence of DLL1 and DLL4.

View Article: PubMed Central - PubMed

Affiliation: Institut für Molekularbiologie OE5250, Medizinische Hochschule Hannover, Hannover, Germany.

ABSTRACT
Notch signalling is a fundamental pathway that shapes the developing embryo and sustains adult tissues by direct communication between ligand and receptor molecules on adjacent cells. Among the ligands are two Delta paralogues, DLL1 and DLL4, that are conserved in mammals and share a similar structure and sequence. They activate the Notch receptor partly in overlapping expression domains where they fulfil redundant functions in some processes (e.g. maintenance of the crypt cell progenitor pool). In other processes, however, they appear to act differently (e.g. maintenance of foetal arterial identity) raising the questions of how similar DLL1 and DLL4 really are and which mechanism causes the apparent context-dependent divergence. By analysing mice that conditionally overexpress DLL1 or DLL4 from the same genomic locus (Hprt) and mice that express DLL4 instead of DLL1 from the endogenous Dll1 locus (Dll1Dll4ki), we found functional differences that are tissue-specific: while DLL1 and DLL4 act redundantly during the maintenance of retinal progenitors, their function varies in the presomitic mesoderm (PSM) where somites form in a Notch-dependent process. In the anterior PSM, every cell expresses both Notch receptors and ligands, and DLL1 is the only activator of Notch while DLL4 is not endogenously expressed. Transgenic DLL4 cannot replace DLL1 during somitogenesis and in heterozygous Dll1Dll4ki/+ mice, the Dll1Dll4ki allele causes a dominant segmentation phenotype. Testing several aspects of the complex Notch signalling system in vitro, we found that both ligands have a similar trans-activation potential but that only DLL4 is an efficient cis-inhibitor of Notch signalling, causing a reduced net activation of Notch. These differential cis-inhibitory properties are likely to contribute to the functional divergence of DLL1 and DLL4.

No MeSH data available.


Related in: MedlinePlus