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NDR Kinases Are Essential for Somitogenesis and Cardiac Looping during Mouse Embryonic Development.

Schmitz-Rohmer D, Probst S, Yang ZZ, Laurent F, Stadler MB, Zuniga A, Zeller R, Hynx D, Hemmings BA, Hergovich A - PLoS ONE (2015)

Bottom Line: In addition, Ndr1/2-double embryos developed a heart defect that manifests itself as pericardial edemas, obstructed heart tubes and arrest of cardiac looping.The resulting cardiac insufficiency is the likely cause of the lethality of Ndr1/2-double embryos around E10.Ndr1/2-double embryos show defects in somitogenesis and cardiac looping, which reveals their essential functions and shows that the NDR kinases are critically required during the early phase of organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.

ABSTRACT
Studies of mammalian tissue culture cells indicate that the conserved and distinct NDR isoforms, NDR1 and NDR2, play essential cell biological roles. However, mice lacking either Ndr1 or Ndr2 alone develop normally. Here, we studied the physiological consequences of inactivating both NDR1 and NDR2 in mice, showing that the lack of both Ndr1/Ndr2 (called Ndr1/2-double mutants) causes embryonic lethality. In support of compensatory roles for NDR1 and NDR2, total protein and activating phosphorylation levels of the remaining NDR isoform were elevated in mice lacking either Ndr1 or Ndr2. Mice retaining one single wild-type Ndr allele were viable and fertile. Ndr1/2-double embryos displayed multiple phenotypes causing a developmental delay from embryonic day E8.5 onwards. While NDR kinases are not required for notochord formation, the somites of Ndr1/2-double embryos were smaller, irregularly shaped and unevenly spaced along the anterior-posterior axis. Genes implicated in somitogenesis were down-regulated and the normally symmetric expression of Lunatic fringe, a component of the Notch pathway, showed a left-right bias in the last forming somite in 50% of all Ndr1/2-double embryos. In addition, Ndr1/2-double embryos developed a heart defect that manifests itself as pericardial edemas, obstructed heart tubes and arrest of cardiac looping. The resulting cardiac insufficiency is the likely cause of the lethality of Ndr1/2-double embryos around E10. Taken together, we show that NDR kinases compensate for each other in vivo in mouse embryos, explaining why mice deficient for either Ndr1 or Ndr2 are viable. Ndr1/2-double embryos show defects in somitogenesis and cardiac looping, which reveals their essential functions and shows that the NDR kinases are critically required during the early phase of organogenesis.

No MeSH data available.


Related in: MedlinePlus

The remaining NDR isoform is upregulated in tissues of mice lacking one of the two Ndr genes.(A) Western blot analysis of NDR1 and NDR2 expression in thymus and colon of wild-type (+/+), Ndr2 heterozygous (+/-) and Ndr2-deficient (-/-) mice. (B) Western Blot analysis of NDR1 and NDR2 proteins in brain, colon, heart, thymus, spleen and lymph nodes (LN) of wild-type (wt), Ndr1-deficient (ko 1) and Ndr2 deficient (ko 2) mice. Hydrophobic motif phosphorylation, which serves as a direct indicator of NDR kinase activity, was examined using anti-phospho-T444/T442 antibodies (detecting NDR1 and NDR2 equally well [65]). The upper band corresponds to T442-P of NDR2, the lower band corresponds to T444-P of NDR1 [65]. HSC-70 served as loading control. Asterisks indicate compensatory HM phosphorylation events.
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pone.0136566.g002: The remaining NDR isoform is upregulated in tissues of mice lacking one of the two Ndr genes.(A) Western blot analysis of NDR1 and NDR2 expression in thymus and colon of wild-type (+/+), Ndr2 heterozygous (+/-) and Ndr2-deficient (-/-) mice. (B) Western Blot analysis of NDR1 and NDR2 proteins in brain, colon, heart, thymus, spleen and lymph nodes (LN) of wild-type (wt), Ndr1-deficient (ko 1) and Ndr2 deficient (ko 2) mice. Hydrophobic motif phosphorylation, which serves as a direct indicator of NDR kinase activity, was examined using anti-phospho-T444/T442 antibodies (detecting NDR1 and NDR2 equally well [65]). The upper band corresponds to T442-P of NDR2, the lower band corresponds to T444-P of NDR1 [65]. HSC-70 served as loading control. Asterisks indicate compensatory HM phosphorylation events.

Mentions: Ndr1 and Ndr2 display partially overlapping expression patterns and in all mouse tissues examined so far at least one of the two NDR isoforms is expressed [2–4, 7, 8]. While NDR1 protein levels are highest in organs of the immune system (thymus, spleen and lymph nodes), NDR2 protein levels peak in the colon and brain [2–4, 7, 8]. Intriguingly, NDR2 protein levels are post-transcriptionally up-regulated upon ablation of Ndr1, suggesting a compensatory link between the two NDR isoforms [7]. More precisely, up-regulation of NDR2 occurs particularly in tissues with high NDR1 expression in the wild-type situation [7]. To address whether the inactivation of Ndr2 is also compensated by up-regulation of NDR1 protein, we analyzed thymus and colon tissue lysates of Ndr2 wild-type, heterozygous and deficient adult littermate mice (Fig 2A). While NDR1 protein levels in the thymus of Ndr2 heterozygous and deficient mice remained unchanged, NDR1 protein levels appeared to be increased in the colon of Ndr2-deficient mice (Fig 2A). This suggested that NDR1 compensates for loss of NDR2 in the colon, a tissue where NDR2 is normally highly expressed.


NDR Kinases Are Essential for Somitogenesis and Cardiac Looping during Mouse Embryonic Development.

Schmitz-Rohmer D, Probst S, Yang ZZ, Laurent F, Stadler MB, Zuniga A, Zeller R, Hynx D, Hemmings BA, Hergovich A - PLoS ONE (2015)

The remaining NDR isoform is upregulated in tissues of mice lacking one of the two Ndr genes.(A) Western blot analysis of NDR1 and NDR2 expression in thymus and colon of wild-type (+/+), Ndr2 heterozygous (+/-) and Ndr2-deficient (-/-) mice. (B) Western Blot analysis of NDR1 and NDR2 proteins in brain, colon, heart, thymus, spleen and lymph nodes (LN) of wild-type (wt), Ndr1-deficient (ko 1) and Ndr2 deficient (ko 2) mice. Hydrophobic motif phosphorylation, which serves as a direct indicator of NDR kinase activity, was examined using anti-phospho-T444/T442 antibodies (detecting NDR1 and NDR2 equally well [65]). The upper band corresponds to T442-P of NDR2, the lower band corresponds to T444-P of NDR1 [65]. HSC-70 served as loading control. Asterisks indicate compensatory HM phosphorylation events.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549247&req=5

pone.0136566.g002: The remaining NDR isoform is upregulated in tissues of mice lacking one of the two Ndr genes.(A) Western blot analysis of NDR1 and NDR2 expression in thymus and colon of wild-type (+/+), Ndr2 heterozygous (+/-) and Ndr2-deficient (-/-) mice. (B) Western Blot analysis of NDR1 and NDR2 proteins in brain, colon, heart, thymus, spleen and lymph nodes (LN) of wild-type (wt), Ndr1-deficient (ko 1) and Ndr2 deficient (ko 2) mice. Hydrophobic motif phosphorylation, which serves as a direct indicator of NDR kinase activity, was examined using anti-phospho-T444/T442 antibodies (detecting NDR1 and NDR2 equally well [65]). The upper band corresponds to T442-P of NDR2, the lower band corresponds to T444-P of NDR1 [65]. HSC-70 served as loading control. Asterisks indicate compensatory HM phosphorylation events.
Mentions: Ndr1 and Ndr2 display partially overlapping expression patterns and in all mouse tissues examined so far at least one of the two NDR isoforms is expressed [2–4, 7, 8]. While NDR1 protein levels are highest in organs of the immune system (thymus, spleen and lymph nodes), NDR2 protein levels peak in the colon and brain [2–4, 7, 8]. Intriguingly, NDR2 protein levels are post-transcriptionally up-regulated upon ablation of Ndr1, suggesting a compensatory link between the two NDR isoforms [7]. More precisely, up-regulation of NDR2 occurs particularly in tissues with high NDR1 expression in the wild-type situation [7]. To address whether the inactivation of Ndr2 is also compensated by up-regulation of NDR1 protein, we analyzed thymus and colon tissue lysates of Ndr2 wild-type, heterozygous and deficient adult littermate mice (Fig 2A). While NDR1 protein levels in the thymus of Ndr2 heterozygous and deficient mice remained unchanged, NDR1 protein levels appeared to be increased in the colon of Ndr2-deficient mice (Fig 2A). This suggested that NDR1 compensates for loss of NDR2 in the colon, a tissue where NDR2 is normally highly expressed.

Bottom Line: In addition, Ndr1/2-double embryos developed a heart defect that manifests itself as pericardial edemas, obstructed heart tubes and arrest of cardiac looping.The resulting cardiac insufficiency is the likely cause of the lethality of Ndr1/2-double embryos around E10.Ndr1/2-double embryos show defects in somitogenesis and cardiac looping, which reveals their essential functions and shows that the NDR kinases are critically required during the early phase of organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.

ABSTRACT
Studies of mammalian tissue culture cells indicate that the conserved and distinct NDR isoforms, NDR1 and NDR2, play essential cell biological roles. However, mice lacking either Ndr1 or Ndr2 alone develop normally. Here, we studied the physiological consequences of inactivating both NDR1 and NDR2 in mice, showing that the lack of both Ndr1/Ndr2 (called Ndr1/2-double mutants) causes embryonic lethality. In support of compensatory roles for NDR1 and NDR2, total protein and activating phosphorylation levels of the remaining NDR isoform were elevated in mice lacking either Ndr1 or Ndr2. Mice retaining one single wild-type Ndr allele were viable and fertile. Ndr1/2-double embryos displayed multiple phenotypes causing a developmental delay from embryonic day E8.5 onwards. While NDR kinases are not required for notochord formation, the somites of Ndr1/2-double embryos were smaller, irregularly shaped and unevenly spaced along the anterior-posterior axis. Genes implicated in somitogenesis were down-regulated and the normally symmetric expression of Lunatic fringe, a component of the Notch pathway, showed a left-right bias in the last forming somite in 50% of all Ndr1/2-double embryos. In addition, Ndr1/2-double embryos developed a heart defect that manifests itself as pericardial edemas, obstructed heart tubes and arrest of cardiac looping. The resulting cardiac insufficiency is the likely cause of the lethality of Ndr1/2-double embryos around E10. Taken together, we show that NDR kinases compensate for each other in vivo in mouse embryos, explaining why mice deficient for either Ndr1 or Ndr2 are viable. Ndr1/2-double embryos show defects in somitogenesis and cardiac looping, which reveals their essential functions and shows that the NDR kinases are critically required during the early phase of organogenesis.

No MeSH data available.


Related in: MedlinePlus