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Repulsive cues combined with physical barriers and cell-cell adhesion determine progenitor cell positioning during organogenesis.

Paksa A, Bandemer J, Hoeckendorf B, Razin N, Tarbashevich K, Minina S, Meyen D, Biundo A, Leidel SA, Peyrieras N, Gov NS, Keller PJ, Raz E - Nat Commun (2016)

Bottom Line: Using primordial germ cells that participate in gonad formation, we present the developmental mechanisms maintaining a motile progenitor cell population at the site where the organ develops.Employing high-resolution live-cell microscopy, we find that repulsive cues coupled with physical barriers confine the cells to the correct bilateral positions.This analysis revealed that cell polarity changes on interaction with the physical barrier and that the establishment of compact clusters involves increased cell-cell interaction time.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell Biology, ZMBE, Von-Esmarch-Street 56, 48149 Muenster, Germany.

ABSTRACT
The precise positioning of organ progenitor cells constitutes an essential, yet poorly understood step during organogenesis. Using primordial germ cells that participate in gonad formation, we present the developmental mechanisms maintaining a motile progenitor cell population at the site where the organ develops. Employing high-resolution live-cell microscopy, we find that repulsive cues coupled with physical barriers confine the cells to the correct bilateral positions. This analysis revealed that cell polarity changes on interaction with the physical barrier and that the establishment of compact clusters involves increased cell-cell interaction time. Using particle-based simulations, we demonstrate the role of reflecting barriers, from which cells turn away on contact, and the importance of proper cell-cell adhesion level for maintaining the tight cell clusters and their correct positioning at the target region. The combination of these developmental and cellular mechanisms prevents organ fusion, controls organ positioning and is thus critical for its proper function.

No MeSH data available.


Related in: MedlinePlus

PGCs avoid regions expressing LPP proteins.(a) Generation of embryos lacking Cxcl12a, whose PGCs are labelled by EGFP and overexpress either LPP proteins or a Control protein in mCherry-labelled half of the embryo. (b) PGCs avoid cellular domains of the embryos, which overexpress LPPs (middle row), as compared to control domains (top row) or those overexpressing phosphatase-inactive versions of LPPs (lower row). (c) A significant reduction in the percentage of PGCs located within the LPPs-overexpressing domain in 10 hpf embryos (one-way analysis of variance; *P≤0.05). Error bars designate minimum to maximum range of the data points. N and n show the number of embryos and PGCs, respectively. See also Supplementary Fig. 8. (d) mCherry-labelled cells overexpressing Cxcl12a with either a Control Protein or LPPs were transplanted into embryos lacking Cxcl12a (medny054) whose PGCs express EGFP. (e) Images and a graph demonstrating the association of PGCs with Cxcl12a-expressing cells in control embryos (upper image, magenta point in graph) and the lack of interaction with cells expressing Cxcl12a and LPPs (starred red cells in lower image, blue points in graph, 77% of embryos showed absolutely no cell association). The statistical significance was evaluated using the Mann–Whitney U-test (****P≤0.0001). Scale bar, 15 μm. See also Supplementary Movie 3 and Supplementary Fig. 9.
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f4: PGCs avoid regions expressing LPP proteins.(a) Generation of embryos lacking Cxcl12a, whose PGCs are labelled by EGFP and overexpress either LPP proteins or a Control protein in mCherry-labelled half of the embryo. (b) PGCs avoid cellular domains of the embryos, which overexpress LPPs (middle row), as compared to control domains (top row) or those overexpressing phosphatase-inactive versions of LPPs (lower row). (c) A significant reduction in the percentage of PGCs located within the LPPs-overexpressing domain in 10 hpf embryos (one-way analysis of variance; *P≤0.05). Error bars designate minimum to maximum range of the data points. N and n show the number of embryos and PGCs, respectively. See also Supplementary Fig. 8. (d) mCherry-labelled cells overexpressing Cxcl12a with either a Control Protein or LPPs were transplanted into embryos lacking Cxcl12a (medny054) whose PGCs express EGFP. (e) Images and a graph demonstrating the association of PGCs with Cxcl12a-expressing cells in control embryos (upper image, magenta point in graph) and the lack of interaction with cells expressing Cxcl12a and LPPs (starred red cells in lower image, blue points in graph, 77% of embryos showed absolutely no cell association). The statistical significance was evaluated using the Mann–Whitney U-test (****P≤0.0001). Scale bar, 15 μm. See also Supplementary Movie 3 and Supplementary Fig. 9.

Mentions: To complement the loss-of-function experiments, we sought to determine whether LPPs expression is sufficient for transforming embryonic structures into less favourable regions. To this end, we generated embryos lacking the guidance cue Cxcl12a, in which certain domains were engineered to overexpress the LPP proteins or a control protein (Fig. 4a and red-labelled regions in Fig. 4b). LPP1-varX1 and LPP3-varX1 that are highly expressed in the tissues dorsal to PGC clusters and exhibit the highest similarity to the Drosophila Wunen proteins were tested in these experiments. Remarkably, we observed a significant bias in PGC localization, as PGCs (green-labelled cells in Fig. 4b) were less likely to reside within domains overexpressing LPPs (Fig. 4b, middle row and Fig. 4c). Similar results were obtained in embryos lacking both guidance cues Cxcl12a and Cxcl12b (Supplementary Fig. 8a). This bias was not detected when the overexpressed protein was irrelevant for the process, nor when the overexpressed LPPs were mutated in their active sites (Fig. 4b, upper and lower rows respectively; Fig. 4c; black boxes in Supplementary Fig. 2 that mark mutation sites24). It is noteworthy that the total number of PGCs in the different experiments remained unchanged (Supplementary Fig. 8b), providing evidence that the observed bias in cell localization resulted from an effect on cell behaviour, rather than enhanced PGC death within domains of LPPs overexpression.


Repulsive cues combined with physical barriers and cell-cell adhesion determine progenitor cell positioning during organogenesis.

Paksa A, Bandemer J, Hoeckendorf B, Razin N, Tarbashevich K, Minina S, Meyen D, Biundo A, Leidel SA, Peyrieras N, Gov NS, Keller PJ, Raz E - Nat Commun (2016)

PGCs avoid regions expressing LPP proteins.(a) Generation of embryos lacking Cxcl12a, whose PGCs are labelled by EGFP and overexpress either LPP proteins or a Control protein in mCherry-labelled half of the embryo. (b) PGCs avoid cellular domains of the embryos, which overexpress LPPs (middle row), as compared to control domains (top row) or those overexpressing phosphatase-inactive versions of LPPs (lower row). (c) A significant reduction in the percentage of PGCs located within the LPPs-overexpressing domain in 10 hpf embryos (one-way analysis of variance; *P≤0.05). Error bars designate minimum to maximum range of the data points. N and n show the number of embryos and PGCs, respectively. See also Supplementary Fig. 8. (d) mCherry-labelled cells overexpressing Cxcl12a with either a Control Protein or LPPs were transplanted into embryos lacking Cxcl12a (medny054) whose PGCs express EGFP. (e) Images and a graph demonstrating the association of PGCs with Cxcl12a-expressing cells in control embryos (upper image, magenta point in graph) and the lack of interaction with cells expressing Cxcl12a and LPPs (starred red cells in lower image, blue points in graph, 77% of embryos showed absolutely no cell association). The statistical significance was evaluated using the Mann–Whitney U-test (****P≤0.0001). Scale bar, 15 μm. See also Supplementary Movie 3 and Supplementary Fig. 9.
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Related In: Results  -  Collection

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f4: PGCs avoid regions expressing LPP proteins.(a) Generation of embryos lacking Cxcl12a, whose PGCs are labelled by EGFP and overexpress either LPP proteins or a Control protein in mCherry-labelled half of the embryo. (b) PGCs avoid cellular domains of the embryos, which overexpress LPPs (middle row), as compared to control domains (top row) or those overexpressing phosphatase-inactive versions of LPPs (lower row). (c) A significant reduction in the percentage of PGCs located within the LPPs-overexpressing domain in 10 hpf embryos (one-way analysis of variance; *P≤0.05). Error bars designate minimum to maximum range of the data points. N and n show the number of embryos and PGCs, respectively. See also Supplementary Fig. 8. (d) mCherry-labelled cells overexpressing Cxcl12a with either a Control Protein or LPPs were transplanted into embryos lacking Cxcl12a (medny054) whose PGCs express EGFP. (e) Images and a graph demonstrating the association of PGCs with Cxcl12a-expressing cells in control embryos (upper image, magenta point in graph) and the lack of interaction with cells expressing Cxcl12a and LPPs (starred red cells in lower image, blue points in graph, 77% of embryos showed absolutely no cell association). The statistical significance was evaluated using the Mann–Whitney U-test (****P≤0.0001). Scale bar, 15 μm. See also Supplementary Movie 3 and Supplementary Fig. 9.
Mentions: To complement the loss-of-function experiments, we sought to determine whether LPPs expression is sufficient for transforming embryonic structures into less favourable regions. To this end, we generated embryos lacking the guidance cue Cxcl12a, in which certain domains were engineered to overexpress the LPP proteins or a control protein (Fig. 4a and red-labelled regions in Fig. 4b). LPP1-varX1 and LPP3-varX1 that are highly expressed in the tissues dorsal to PGC clusters and exhibit the highest similarity to the Drosophila Wunen proteins were tested in these experiments. Remarkably, we observed a significant bias in PGC localization, as PGCs (green-labelled cells in Fig. 4b) were less likely to reside within domains overexpressing LPPs (Fig. 4b, middle row and Fig. 4c). Similar results were obtained in embryos lacking both guidance cues Cxcl12a and Cxcl12b (Supplementary Fig. 8a). This bias was not detected when the overexpressed protein was irrelevant for the process, nor when the overexpressed LPPs were mutated in their active sites (Fig. 4b, upper and lower rows respectively; Fig. 4c; black boxes in Supplementary Fig. 2 that mark mutation sites24). It is noteworthy that the total number of PGCs in the different experiments remained unchanged (Supplementary Fig. 8b), providing evidence that the observed bias in cell localization resulted from an effect on cell behaviour, rather than enhanced PGC death within domains of LPPs overexpression.

Bottom Line: Using primordial germ cells that participate in gonad formation, we present the developmental mechanisms maintaining a motile progenitor cell population at the site where the organ develops.Employing high-resolution live-cell microscopy, we find that repulsive cues coupled with physical barriers confine the cells to the correct bilateral positions.This analysis revealed that cell polarity changes on interaction with the physical barrier and that the establishment of compact clusters involves increased cell-cell interaction time.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell Biology, ZMBE, Von-Esmarch-Street 56, 48149 Muenster, Germany.

ABSTRACT
The precise positioning of organ progenitor cells constitutes an essential, yet poorly understood step during organogenesis. Using primordial germ cells that participate in gonad formation, we present the developmental mechanisms maintaining a motile progenitor cell population at the site where the organ develops. Employing high-resolution live-cell microscopy, we find that repulsive cues coupled with physical barriers confine the cells to the correct bilateral positions. This analysis revealed that cell polarity changes on interaction with the physical barrier and that the establishment of compact clusters involves increased cell-cell interaction time. Using particle-based simulations, we demonstrate the role of reflecting barriers, from which cells turn away on contact, and the importance of proper cell-cell adhesion level for maintaining the tight cell clusters and their correct positioning at the target region. The combination of these developmental and cellular mechanisms prevents organ fusion, controls organ positioning and is thus critical for its proper function.

No MeSH data available.


Related in: MedlinePlus