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The Nanos3-3'UTR is required for germ cell specific NANOS3 expression in mouse embryos.

Suzuki H, Saba R, Sada A, Saga Y - PLoS ONE (2010)

Bottom Line: This is fundamental to the continuation of a species.Although Nanos3 is transcribed in both cell lineages, it is efficiently translated only in the germ lineage.Surprisingly, even when under the control of the CAG promoter which induces strong ubiquitous transcription in both germ cells and somatic cells, the addition of the Nanos3-3'UTR sequence to the coding region of exogenous gene was effective in restricting protein expression in germ cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo, Japan.

ABSTRACT

Background: The regulation of gene expression via a 3' untranslated region (UTR) plays essential roles in the discrimination of the germ cell lineage from somatic cells during embryogenesis. This is fundamental to the continuation of a species. Mouse NANOS3 is an essential protein required for the germ cell maintenance and is specifically expressed in these cells. However, the regulatory mechanisms that restrict the expression of this gene in the germ cells is largely unknown at present.

Methodology/principal findings: In our current study, we show that differences in the stability of Nanos3 mRNA between germ cells and somatic cells is brought about in a 3'UTR-dependent manner in mouse embryos. Although Nanos3 is transcribed in both cell lineages, it is efficiently translated only in the germ lineage. We also find that the translational suppression of NANOS3 in somatic cells is caused by a 3'UTR-mediated mRNA destabilizing mechanism. Surprisingly, even when under the control of the CAG promoter which induces strong ubiquitous transcription in both germ cells and somatic cells, the addition of the Nanos3-3'UTR sequence to the coding region of exogenous gene was effective in restricting protein expression in germ cells.

Conclusions/significance: Our current study thus suggests that Nanos3-3'UTR has an essential role in translational control in the mouse embryo.

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Related in: MedlinePlus

Nanos3-3′UTR is sufficient to establish the germ cell-specific expression pattern in the mouse embryo.(A–C) Fluorescence images of CAG-mRFP(Nos3-3′UTR) (A, male; B, female) and CAG-mRFP(BghpA) (C, male) transgenic embryos at E14.5. The upper images are of the abdomens of embryos harboring transgenes (Tg+), whereas the lower panels show corresponding images from embryos lacking a transgene (Tg−). Broken gray lines indicate gonads. (D–E) Quantitative RT-PCR analysis of mRFP in CAG-mRFP(Nos3-3′UTR) (D) or CAG-Lyn-mRFP(BghpA) (E) embryos at E14.5. The data were normalized using G3PDH. M–K, male kidney; M-Li, male limb; M–G, male gonad; F–G, female gonad; N, wild-type embryo. Error bars represent the s.d. Student t-test was used to calculate P values. *, P<0.05.
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pone-0009300-g005: Nanos3-3′UTR is sufficient to establish the germ cell-specific expression pattern in the mouse embryo.(A–C) Fluorescence images of CAG-mRFP(Nos3-3′UTR) (A, male; B, female) and CAG-mRFP(BghpA) (C, male) transgenic embryos at E14.5. The upper images are of the abdomens of embryos harboring transgenes (Tg+), whereas the lower panels show corresponding images from embryos lacking a transgene (Tg−). Broken gray lines indicate gonads. (D–E) Quantitative RT-PCR analysis of mRFP in CAG-mRFP(Nos3-3′UTR) (D) or CAG-Lyn-mRFP(BghpA) (E) embryos at E14.5. The data were normalized using G3PDH. M–K, male kidney; M-Li, male limb; M–G, male gonad; F–G, female gonad; N, wild-type embryo. Error bars represent the s.d. Student t-test was used to calculate P values. *, P<0.05.

Mentions: To further investigate whether the Nanos3-3′UTR affects the transcription or stability of mRNA, we generated two additional transgenic mice. We utilized the CAG promoter, a known strong promoter-enhancer that drives the ubiquitous transcription of mRFP with either Nanos3-3′UTR or BghpA (Fig. 1C–D). Surprisingly, Nanos3-3′UTR proved to be effective in restricting the mRFP expression in the germ cells at E14.5 (Fig. 5A–B), whilst mRFP was always expressed ubiquitously with no pattern observed in the CAG-mRFP(BghpA) embryo (Fig. 5C). The same expression pattern was observed in CAG-lyn-mRFP(BghpA) transgenic embryo that had been previously established in our laboratory [34]. The lyn-mRFP is an mRFP that contains the lyn kinase at its N-terminus, which serves as a membrane localization signal but does not affect neither transcription and translation [34]. Therefore, we considered CAG-lyn-mRFP(BghpA) is compatible with CAG-mRFP(BghpA). By qRT-PCR analyses, we further revealed that the relative amounts of mRFP mRNA in somatic tissues (a kidney and a hind limb) were significantly lower than those in the gonads in CAG-mRFP(Nos3-3′UTR) embryos whereas not significantly altered in CAG-lyn-mRFP(BghpA) (Fig. 5D and E). The results suggest that Nanos3-3′UTR is sufficient to suppress protein expression in somatic cells by destabilizing mRNA and establishing a germ-cell specific expression pattern.


The Nanos3-3'UTR is required for germ cell specific NANOS3 expression in mouse embryos.

Suzuki H, Saba R, Sada A, Saga Y - PLoS ONE (2010)

Nanos3-3′UTR is sufficient to establish the germ cell-specific expression pattern in the mouse embryo.(A–C) Fluorescence images of CAG-mRFP(Nos3-3′UTR) (A, male; B, female) and CAG-mRFP(BghpA) (C, male) transgenic embryos at E14.5. The upper images are of the abdomens of embryos harboring transgenes (Tg+), whereas the lower panels show corresponding images from embryos lacking a transgene (Tg−). Broken gray lines indicate gonads. (D–E) Quantitative RT-PCR analysis of mRFP in CAG-mRFP(Nos3-3′UTR) (D) or CAG-Lyn-mRFP(BghpA) (E) embryos at E14.5. The data were normalized using G3PDH. M–K, male kidney; M-Li, male limb; M–G, male gonad; F–G, female gonad; N, wild-type embryo. Error bars represent the s.d. Student t-test was used to calculate P values. *, P<0.05.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2823788&req=5

pone-0009300-g005: Nanos3-3′UTR is sufficient to establish the germ cell-specific expression pattern in the mouse embryo.(A–C) Fluorescence images of CAG-mRFP(Nos3-3′UTR) (A, male; B, female) and CAG-mRFP(BghpA) (C, male) transgenic embryos at E14.5. The upper images are of the abdomens of embryos harboring transgenes (Tg+), whereas the lower panels show corresponding images from embryos lacking a transgene (Tg−). Broken gray lines indicate gonads. (D–E) Quantitative RT-PCR analysis of mRFP in CAG-mRFP(Nos3-3′UTR) (D) or CAG-Lyn-mRFP(BghpA) (E) embryos at E14.5. The data were normalized using G3PDH. M–K, male kidney; M-Li, male limb; M–G, male gonad; F–G, female gonad; N, wild-type embryo. Error bars represent the s.d. Student t-test was used to calculate P values. *, P<0.05.
Mentions: To further investigate whether the Nanos3-3′UTR affects the transcription or stability of mRNA, we generated two additional transgenic mice. We utilized the CAG promoter, a known strong promoter-enhancer that drives the ubiquitous transcription of mRFP with either Nanos3-3′UTR or BghpA (Fig. 1C–D). Surprisingly, Nanos3-3′UTR proved to be effective in restricting the mRFP expression in the germ cells at E14.5 (Fig. 5A–B), whilst mRFP was always expressed ubiquitously with no pattern observed in the CAG-mRFP(BghpA) embryo (Fig. 5C). The same expression pattern was observed in CAG-lyn-mRFP(BghpA) transgenic embryo that had been previously established in our laboratory [34]. The lyn-mRFP is an mRFP that contains the lyn kinase at its N-terminus, which serves as a membrane localization signal but does not affect neither transcription and translation [34]. Therefore, we considered CAG-lyn-mRFP(BghpA) is compatible with CAG-mRFP(BghpA). By qRT-PCR analyses, we further revealed that the relative amounts of mRFP mRNA in somatic tissues (a kidney and a hind limb) were significantly lower than those in the gonads in CAG-mRFP(Nos3-3′UTR) embryos whereas not significantly altered in CAG-lyn-mRFP(BghpA) (Fig. 5D and E). The results suggest that Nanos3-3′UTR is sufficient to suppress protein expression in somatic cells by destabilizing mRNA and establishing a germ-cell specific expression pattern.

Bottom Line: This is fundamental to the continuation of a species.Although Nanos3 is transcribed in both cell lineages, it is efficiently translated only in the germ lineage.Surprisingly, even when under the control of the CAG promoter which induces strong ubiquitous transcription in both germ cells and somatic cells, the addition of the Nanos3-3'UTR sequence to the coding region of exogenous gene was effective in restricting protein expression in germ cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo, Japan.

ABSTRACT

Background: The regulation of gene expression via a 3' untranslated region (UTR) plays essential roles in the discrimination of the germ cell lineage from somatic cells during embryogenesis. This is fundamental to the continuation of a species. Mouse NANOS3 is an essential protein required for the germ cell maintenance and is specifically expressed in these cells. However, the regulatory mechanisms that restrict the expression of this gene in the germ cells is largely unknown at present.

Methodology/principal findings: In our current study, we show that differences in the stability of Nanos3 mRNA between germ cells and somatic cells is brought about in a 3'UTR-dependent manner in mouse embryos. Although Nanos3 is transcribed in both cell lineages, it is efficiently translated only in the germ lineage. We also find that the translational suppression of NANOS3 in somatic cells is caused by a 3'UTR-mediated mRNA destabilizing mechanism. Surprisingly, even when under the control of the CAG promoter which induces strong ubiquitous transcription in both germ cells and somatic cells, the addition of the Nanos3-3'UTR sequence to the coding region of exogenous gene was effective in restricting protein expression in germ cells.

Conclusions/significance: Our current study thus suggests that Nanos3-3'UTR has an essential role in translational control in the mouse embryo.

Show MeSH
Related in: MedlinePlus