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Insulin-like factor regulates neural induction through an IGF1 receptor-independent mechanism.

Haramoto Y, Takahashi S, Oshima T, Onuma Y, Ito Y, Asashima M - Sci Rep (2015)

Bottom Line: Insulin3 reduced extracellular Wnts and cell surface localised Lrp6.These results suggest that Insulin3 is a novel cell-autonomous inhibitor of Wnt signalling.This study provides the first evidence that an insulin-like factor regulates neural induction through an IGF1R-independent mechanism.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Stem Cell Engineering, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.

ABSTRACT
Insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R) signalling is required for normal embryonic growth and development. Previous reports indicated that the IGF/IGF1R/MAPK pathway contributes to neural induction and the IGF/IGF1R/PI3K/Akt pathway to eye development. Here, we report the isolation of insulin3 encoding a novel insulin-like ligand involved in neural induction. Insulin3 has a similar structure to pro-insulin and mature IGF ligands, but cannot activate the IGF1 receptor. However, similar to IGFs, Insulin3 induced the gene expression of an anterior neural marker, otx2, and enlarged anterior head structures by inhibiting Wnt signalling. Insulin3 are predominantly localised to the endoplasmic reticulum when otx2 is induced by insulin3. Insulin3 reduced extracellular Wnts and cell surface localised Lrp6. These results suggest that Insulin3 is a novel cell-autonomous inhibitor of Wnt signalling. This study provides the first evidence that an insulin-like factor regulates neural induction through an IGF1R-independent mechanism.

No MeSH data available.


Related in: MedlinePlus

Characterization of insulin3 in Xenopus.(a) Stage PCR for insulin3. Stages of samples are indicated (top). (b) Whole-mount in situ hybridization of insulin3 transcripts. Stages of samples are indicated (right). Viewpoints are indicated (top). The line indicates the position of the cross-section. (c) mRNAs were injected into the animal pole of two-cell-stage X. laevis embryos. Embryos were harvested at stage 30. Amounts of mRNA injected per embryos were: insulin1 (2 ng), IGF1 (300 pg) and insulin3 (300 pg). Expanded cement gland was induced by injection of insulin3 and IGF1, but not by injection of insulin1. (d) Animal cap assay. One ng of IGF1 and insulin3 mRNAs were injected into the animal pole of both blastomeres at the two-cell stage in X. laevis embryos. Animal caps were dissected at stage 9 and harvested at stage 11. IGF1 and Insulin3 induce otx2 and inhibit without induction of BMP antagonists. WE: whole embryo. RT-: reverse transcriptase minus reaction. Full-length gels are presented in Supplementary figure S7.
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f1: Characterization of insulin3 in Xenopus.(a) Stage PCR for insulin3. Stages of samples are indicated (top). (b) Whole-mount in situ hybridization of insulin3 transcripts. Stages of samples are indicated (right). Viewpoints are indicated (top). The line indicates the position of the cross-section. (c) mRNAs were injected into the animal pole of two-cell-stage X. laevis embryos. Embryos were harvested at stage 30. Amounts of mRNA injected per embryos were: insulin1 (2 ng), IGF1 (300 pg) and insulin3 (300 pg). Expanded cement gland was induced by injection of insulin3 and IGF1, but not by injection of insulin1. (d) Animal cap assay. One ng of IGF1 and insulin3 mRNAs were injected into the animal pole of both blastomeres at the two-cell stage in X. laevis embryos. Animal caps were dissected at stage 9 and harvested at stage 11. IGF1 and Insulin3 induce otx2 and inhibit without induction of BMP antagonists. WE: whole embryo. RT-: reverse transcriptase minus reaction. Full-length gels are presented in Supplementary figure S7.

Mentions: Insulin3 is expressed from late blastulae to the early tailbud stage (Fig. 1a). At the early gastrulae, Insulin3 was detected in whole presumptive neuroectoderm (Fig. 1b). At the late gastrulae and the early neurulae stage (stage 15), insulin3 expression was detected at the midpoint of the mediolateral axis of the neural plate and slightly at the archenteron roof. The insulin3 expressing cells are located in the subepithelial layer of the neural plate. At the late neurulae stage (stage 20), insulin3 expression appears in the anterior endoderm forward of the prechordal plate (Fig. 1b).


Insulin-like factor regulates neural induction through an IGF1 receptor-independent mechanism.

Haramoto Y, Takahashi S, Oshima T, Onuma Y, Ito Y, Asashima M - Sci Rep (2015)

Characterization of insulin3 in Xenopus.(a) Stage PCR for insulin3. Stages of samples are indicated (top). (b) Whole-mount in situ hybridization of insulin3 transcripts. Stages of samples are indicated (right). Viewpoints are indicated (top). The line indicates the position of the cross-section. (c) mRNAs were injected into the animal pole of two-cell-stage X. laevis embryos. Embryos were harvested at stage 30. Amounts of mRNA injected per embryos were: insulin1 (2 ng), IGF1 (300 pg) and insulin3 (300 pg). Expanded cement gland was induced by injection of insulin3 and IGF1, but not by injection of insulin1. (d) Animal cap assay. One ng of IGF1 and insulin3 mRNAs were injected into the animal pole of both blastomeres at the two-cell stage in X. laevis embryos. Animal caps were dissected at stage 9 and harvested at stage 11. IGF1 and Insulin3 induce otx2 and inhibit without induction of BMP antagonists. WE: whole embryo. RT-: reverse transcriptase minus reaction. Full-length gels are presented in Supplementary figure S7.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4481404&req=5

f1: Characterization of insulin3 in Xenopus.(a) Stage PCR for insulin3. Stages of samples are indicated (top). (b) Whole-mount in situ hybridization of insulin3 transcripts. Stages of samples are indicated (right). Viewpoints are indicated (top). The line indicates the position of the cross-section. (c) mRNAs were injected into the animal pole of two-cell-stage X. laevis embryos. Embryos were harvested at stage 30. Amounts of mRNA injected per embryos were: insulin1 (2 ng), IGF1 (300 pg) and insulin3 (300 pg). Expanded cement gland was induced by injection of insulin3 and IGF1, but not by injection of insulin1. (d) Animal cap assay. One ng of IGF1 and insulin3 mRNAs were injected into the animal pole of both blastomeres at the two-cell stage in X. laevis embryos. Animal caps were dissected at stage 9 and harvested at stage 11. IGF1 and Insulin3 induce otx2 and inhibit without induction of BMP antagonists. WE: whole embryo. RT-: reverse transcriptase minus reaction. Full-length gels are presented in Supplementary figure S7.
Mentions: Insulin3 is expressed from late blastulae to the early tailbud stage (Fig. 1a). At the early gastrulae, Insulin3 was detected in whole presumptive neuroectoderm (Fig. 1b). At the late gastrulae and the early neurulae stage (stage 15), insulin3 expression was detected at the midpoint of the mediolateral axis of the neural plate and slightly at the archenteron roof. The insulin3 expressing cells are located in the subepithelial layer of the neural plate. At the late neurulae stage (stage 20), insulin3 expression appears in the anterior endoderm forward of the prechordal plate (Fig. 1b).

Bottom Line: Insulin3 reduced extracellular Wnts and cell surface localised Lrp6.These results suggest that Insulin3 is a novel cell-autonomous inhibitor of Wnt signalling.This study provides the first evidence that an insulin-like factor regulates neural induction through an IGF1R-independent mechanism.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Stem Cell Engineering, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.

ABSTRACT
Insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R) signalling is required for normal embryonic growth and development. Previous reports indicated that the IGF/IGF1R/MAPK pathway contributes to neural induction and the IGF/IGF1R/PI3K/Akt pathway to eye development. Here, we report the isolation of insulin3 encoding a novel insulin-like ligand involved in neural induction. Insulin3 has a similar structure to pro-insulin and mature IGF ligands, but cannot activate the IGF1 receptor. However, similar to IGFs, Insulin3 induced the gene expression of an anterior neural marker, otx2, and enlarged anterior head structures by inhibiting Wnt signalling. Insulin3 are predominantly localised to the endoplasmic reticulum when otx2 is induced by insulin3. Insulin3 reduced extracellular Wnts and cell surface localised Lrp6. These results suggest that Insulin3 is a novel cell-autonomous inhibitor of Wnt signalling. This study provides the first evidence that an insulin-like factor regulates neural induction through an IGF1R-independent mechanism.

No MeSH data available.


Related in: MedlinePlus