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Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells.

Kunwar PS, Starz-Gaiano M, Bainton RJ, Heberlein U, Lehmann R - PLoS Biol. (2003)

Bottom Line: In tre1 mutant embryos, most germ cells do not exit the PMG.Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells.Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Developmental Genetics Program, New York University School of Medicine, New York, New York, USA.

ABSTRACT
In most organisms, germ cells are formed distant from the somatic part of the gonad and thus have to migrate along and through a variety of tissues to reach the gonad. Transepithelial migration through the posterior midgut (PMG) is the first active step during Drosophila germ cell migration. Here we report the identification of a novel G protein-coupled receptor (GPCR), Tre1, that is essential for this migration step. Maternal tre1 RNA is localized to germ cells, and tre1 is required cell autonomously in germ cells. In tre1 mutant embryos, most germ cells do not exit the PMG. The few germ cells that do leave the midgut early migrate normally to the gonad, suggesting that this gene is specifically required for transepithelial migration and that mutant germ cells are still able to recognize other guidance cues. Additionally, inhibiting small Rho GTPases in germ cells affects transepithelial migration, suggesting that Tre1 signals through Rho1. We propose that Tre1 acts in a manner similar to chemokine receptors required during transepithelial migration of leukocytes, implying an evolutionarily conserved mechanism of transepithelial migration. Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells. Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

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Germ Cell-Specific Rescue of tre1 Phenotype(A) and (C) depict the experimental rationale for the tissue-specific gene expression experiment. EP(X)0496, which drives expression of tre1 RNA, was expressed either in the germline by the germline-specific driver nos-GAL4 (A) or in the soma by somatic blastoderm cell-specific driver o-GAL4 (C). nos-GAL4 (yellow) is maternally localized to the posterior pole (yellow) and drives expression in germ cells (green), starting at stage 7 and persisting through embryogenesis and transiently in posterior somatic tissues at blastoderm stage (data not shown). o-GAL4 (C) (yellow) drives expression in all somatic cells at blastoderm stage except for germ cells (green). (B and D) Embryos at stage 13 (top view) stained with anti-Vasa. Anterior is left. Expression of tre1 only in the germ cells rescued the tre1 phenotype (B). Expression of tre1 in somatic tissues did not rescue the tre1 mutant phenotype (D).
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pbio.0000080-g006: Germ Cell-Specific Rescue of tre1 Phenotype(A) and (C) depict the experimental rationale for the tissue-specific gene expression experiment. EP(X)0496, which drives expression of tre1 RNA, was expressed either in the germline by the germline-specific driver nos-GAL4 (A) or in the soma by somatic blastoderm cell-specific driver o-GAL4 (C). nos-GAL4 (yellow) is maternally localized to the posterior pole (yellow) and drives expression in germ cells (green), starting at stage 7 and persisting through embryogenesis and transiently in posterior somatic tissues at blastoderm stage (data not shown). o-GAL4 (C) (yellow) drives expression in all somatic cells at blastoderm stage except for germ cells (green). (B and D) Embryos at stage 13 (top view) stained with anti-Vasa. Anterior is left. Expression of tre1 only in the germ cells rescued the tre1 phenotype (B). Expression of tre1 in somatic tissues did not rescue the tre1 mutant phenotype (D).

Mentions: In the second approach, we used tissue-specific gene expression to determine where Tre1 function is required. Using the germ cell-specific GAL4 driver nos-GAL4 and the EP line EP0496, we expressed tre1 in the germ cells and tested whether the tre1 mutant phenotype can be rescued (Figure 6A) (Van Doren et al. 1998b). In EP0496, the UAS sites required for GAL4-mediated transcriptional activation are inserted in the tre1 promoter region and drive expression of tre1 RNA under GAL4 control. Since the tre1 gene is located on the X chromosome, only half of the embryos are expected to carry a copy of EP0496 and should thus express the tre1 gene in the germ cells. In this experiment, 50% of the embryos obtained by crossing homozygous ΔEP5 mothers carrying the nos-GAL4 transgene to EP0496 males showed a complete rescue of the transepithelial migration phenotype (Figure 6B). Embryos derived from crossing a UAS-LacZ line to ΔEP5 mothers carrying nos-GAL4 showed only the minor zygotic rescue of the mutant phenotype as described above (data not shown). A difficulty in the interpretation of this experiment lies in the fact that the nos-GAL4 driver also transiently activates somatic expression in the PMG anlage at the blastoderm stage (Van Doren et al. 1998b). Thus, the phenotypic rescue could be due to expression of tre1 in the PMG during the blastoderm stage rather than due to germ cell expression. To rule out this possibility, we wanted to express tre1 in the PMG anlage at the blastoderm stage. Since there are no early GAL4 drivers available that specifically express a reporter in the PMG anlage, we used a somatic driver, o-GAL4, which efficiently drives expression in all somatic tissues, including the PMG, during the blastoderm stage, but does not activate transcription in the germ cells (Figure 6C) (W. Gehring and E. Wieschaus, personal communication). We did not observe any rescue of the tre1 phenotype with this driver (Figure 6D; total number of embryos analyzed, n = 200). These experiments demonstrate that tre1 is required autonomously in germ cells for their migration through the PMG and that transcription of tre1 in early germ cells is sufficient to rescue the migration phenotype.


Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells.

Kunwar PS, Starz-Gaiano M, Bainton RJ, Heberlein U, Lehmann R - PLoS Biol. (2003)

Germ Cell-Specific Rescue of tre1 Phenotype(A) and (C) depict the experimental rationale for the tissue-specific gene expression experiment. EP(X)0496, which drives expression of tre1 RNA, was expressed either in the germline by the germline-specific driver nos-GAL4 (A) or in the soma by somatic blastoderm cell-specific driver o-GAL4 (C). nos-GAL4 (yellow) is maternally localized to the posterior pole (yellow) and drives expression in germ cells (green), starting at stage 7 and persisting through embryogenesis and transiently in posterior somatic tissues at blastoderm stage (data not shown). o-GAL4 (C) (yellow) drives expression in all somatic cells at blastoderm stage except for germ cells (green). (B and D) Embryos at stage 13 (top view) stained with anti-Vasa. Anterior is left. Expression of tre1 only in the germ cells rescued the tre1 phenotype (B). Expression of tre1 in somatic tissues did not rescue the tre1 mutant phenotype (D).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC300690&req=5

pbio.0000080-g006: Germ Cell-Specific Rescue of tre1 Phenotype(A) and (C) depict the experimental rationale for the tissue-specific gene expression experiment. EP(X)0496, which drives expression of tre1 RNA, was expressed either in the germline by the germline-specific driver nos-GAL4 (A) or in the soma by somatic blastoderm cell-specific driver o-GAL4 (C). nos-GAL4 (yellow) is maternally localized to the posterior pole (yellow) and drives expression in germ cells (green), starting at stage 7 and persisting through embryogenesis and transiently in posterior somatic tissues at blastoderm stage (data not shown). o-GAL4 (C) (yellow) drives expression in all somatic cells at blastoderm stage except for germ cells (green). (B and D) Embryos at stage 13 (top view) stained with anti-Vasa. Anterior is left. Expression of tre1 only in the germ cells rescued the tre1 phenotype (B). Expression of tre1 in somatic tissues did not rescue the tre1 mutant phenotype (D).
Mentions: In the second approach, we used tissue-specific gene expression to determine where Tre1 function is required. Using the germ cell-specific GAL4 driver nos-GAL4 and the EP line EP0496, we expressed tre1 in the germ cells and tested whether the tre1 mutant phenotype can be rescued (Figure 6A) (Van Doren et al. 1998b). In EP0496, the UAS sites required for GAL4-mediated transcriptional activation are inserted in the tre1 promoter region and drive expression of tre1 RNA under GAL4 control. Since the tre1 gene is located on the X chromosome, only half of the embryos are expected to carry a copy of EP0496 and should thus express the tre1 gene in the germ cells. In this experiment, 50% of the embryos obtained by crossing homozygous ΔEP5 mothers carrying the nos-GAL4 transgene to EP0496 males showed a complete rescue of the transepithelial migration phenotype (Figure 6B). Embryos derived from crossing a UAS-LacZ line to ΔEP5 mothers carrying nos-GAL4 showed only the minor zygotic rescue of the mutant phenotype as described above (data not shown). A difficulty in the interpretation of this experiment lies in the fact that the nos-GAL4 driver also transiently activates somatic expression in the PMG anlage at the blastoderm stage (Van Doren et al. 1998b). Thus, the phenotypic rescue could be due to expression of tre1 in the PMG during the blastoderm stage rather than due to germ cell expression. To rule out this possibility, we wanted to express tre1 in the PMG anlage at the blastoderm stage. Since there are no early GAL4 drivers available that specifically express a reporter in the PMG anlage, we used a somatic driver, o-GAL4, which efficiently drives expression in all somatic tissues, including the PMG, during the blastoderm stage, but does not activate transcription in the germ cells (Figure 6C) (W. Gehring and E. Wieschaus, personal communication). We did not observe any rescue of the tre1 phenotype with this driver (Figure 6D; total number of embryos analyzed, n = 200). These experiments demonstrate that tre1 is required autonomously in germ cells for their migration through the PMG and that transcription of tre1 in early germ cells is sufficient to rescue the migration phenotype.

Bottom Line: In tre1 mutant embryos, most germ cells do not exit the PMG.Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells.Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Developmental Genetics Program, New York University School of Medicine, New York, New York, USA.

ABSTRACT
In most organisms, germ cells are formed distant from the somatic part of the gonad and thus have to migrate along and through a variety of tissues to reach the gonad. Transepithelial migration through the posterior midgut (PMG) is the first active step during Drosophila germ cell migration. Here we report the identification of a novel G protein-coupled receptor (GPCR), Tre1, that is essential for this migration step. Maternal tre1 RNA is localized to germ cells, and tre1 is required cell autonomously in germ cells. In tre1 mutant embryos, most germ cells do not exit the PMG. The few germ cells that do leave the midgut early migrate normally to the gonad, suggesting that this gene is specifically required for transepithelial migration and that mutant germ cells are still able to recognize other guidance cues. Additionally, inhibiting small Rho GTPases in germ cells affects transepithelial migration, suggesting that Tre1 signals through Rho1. We propose that Tre1 acts in a manner similar to chemokine receptors required during transepithelial migration of leukocytes, implying an evolutionarily conserved mechanism of transepithelial migration. Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells. Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

Show MeSH