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Genome-wide analysis reveals a major role in cell fate maintenance and an unexpected role in endoreduplication for the Drosophila FoxA gene Fork head.

Maruyama R, Grevengoed E, Stempniewicz P, Andrew DJ - PLoS ONE (2011)

Bottom Line: Transcription factors drive organogenesis, from the initiation of cell fate decisions to the maintenance and implementation of these decisions.Thus, unlike the worm FoxA protein PHA-4, Fkh does not function to specify cell fate.Overall, this study demonstrates an important role for Fkh in determining how an organ preserves its identity throughout development and provides an alternative paradigm for how FoxA proteins function in organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Johns Hopkins University, School of Medicine, Baltimore, Maryland, United States of America.

ABSTRACT
Transcription factors drive organogenesis, from the initiation of cell fate decisions to the maintenance and implementation of these decisions. The Drosophila embryonic salivary gland provides an excellent platform for unraveling the underlying transcriptional networks of organ development because Drosophila is relatively unencumbered by significant genetic redundancy. The highly conserved FoxA family transcription factors are essential for various aspects of organogenesis in all animals that have been studied. Here, we explore the role of the single Drosophila FoxA protein Fork head (Fkh) in salivary gland organogenesis using two genome-wide strategies. A large-scale in situ hybridization analysis reveals a major role for Fkh in maintaining the salivary gland fate decision and controlling salivary gland physiological activity, in addition to its previously known roles in morphogenesis and survival. The majority of salivary gland genes (59%) are affected by fkh loss, mainly at later stages of salivary gland development. We show that global expression of Fkh cannot drive ectopic salivary gland formation. Thus, unlike the worm FoxA protein PHA-4, Fkh does not function to specify cell fate. In addition, Fkh only indirectly regulates many salivary gland genes, which is also distinct from the role of PHA-4 in organogenesis. Our microarray analyses reveal unexpected roles for Fkh in blocking terminal differentiation and in endoreduplication in the salivary gland and in other Fkh-expressing embryonic tissues. Overall, this study demonstrates an important role for Fkh in determining how an organ preserves its identity throughout development and provides an alternative paradigm for how FoxA proteins function in organogenesis.

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Six groups of fkh dependent genes.In situ hybridizations of examples from each group of SG genes is shown for WT and fkh mutants. The fkh mutants are homozygous for Df(3L)H99, which blocks the SG cell death associated with fkh loss. Red arrowheads indicate the SGs. Blank arrowheads indicate the salivary duct. (A,B) Lateral views of WT and fkh embryos at stage 11 are shown. (C–H) Left two panels show lateral views of WT and fkh embryos at stage 11. Right two panels show ventral views of WT and fkh embryos at stage 13/14. SGs remain on the ventral surface of fkh mutants. (A) Fkh does not affect expression of most early SG genes, including Noa36. (B) Expression of eight early SG genes was significantly reduced in SGs of fkh mutants, as shown with CG32269. (C) Fourteen early genes affected by loss of fkh showed higher levels of expression in late embryonic SGs. This group included trh and nyo, which are initially expressed throughout the SG and duct primordia but that subsequently become restricted to the duct in WT embryos. (D) Two early expressed SG genes showed decreased expression at early stages and increased expression at late stages, as shown for the Pepck gene. (E) Expression of 27% of continuously expressed SG genes was unaffected by fkh loss. The Hmu gene is an example of this class. Hmu transcripts localize to the apical domains of SG cells. (F) 30% of continuously expressed SG genes showed reduced expression at all stages in fkh mutants, as seen with CG30497. (G) Many continuously expressed genes were unaffected at early stages but showed reduced expression at late stages, including the bHLH transcription factor gene sage. (H) All five late expressed SG genes were affected by loss of fkh, as observed with Mvl.
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pone-0020901-g003: Six groups of fkh dependent genes.In situ hybridizations of examples from each group of SG genes is shown for WT and fkh mutants. The fkh mutants are homozygous for Df(3L)H99, which blocks the SG cell death associated with fkh loss. Red arrowheads indicate the SGs. Blank arrowheads indicate the salivary duct. (A,B) Lateral views of WT and fkh embryos at stage 11 are shown. (C–H) Left two panels show lateral views of WT and fkh embryos at stage 11. Right two panels show ventral views of WT and fkh embryos at stage 13/14. SGs remain on the ventral surface of fkh mutants. (A) Fkh does not affect expression of most early SG genes, including Noa36. (B) Expression of eight early SG genes was significantly reduced in SGs of fkh mutants, as shown with CG32269. (C) Fourteen early genes affected by loss of fkh showed higher levels of expression in late embryonic SGs. This group included trh and nyo, which are initially expressed throughout the SG and duct primordia but that subsequently become restricted to the duct in WT embryos. (D) Two early expressed SG genes showed decreased expression at early stages and increased expression at late stages, as shown for the Pepck gene. (E) Expression of 27% of continuously expressed SG genes was unaffected by fkh loss. The Hmu gene is an example of this class. Hmu transcripts localize to the apical domains of SG cells. (F) 30% of continuously expressed SG genes showed reduced expression at all stages in fkh mutants, as seen with CG30497. (G) Many continuously expressed genes were unaffected at early stages but showed reduced expression at late stages, including the bHLH transcription factor gene sage. (H) All five late expressed SG genes were affected by loss of fkh, as observed with Mvl.

Mentions: Further analysis revealed additional subtlety in how the early and continuously expressed classes of genes were affected by fkh loss. Early expressed genes affected by fkh loss could be classified into three groups, ‘decreased’, ‘upregulated later’ and ‘decreased and upregulated later’ (Figure 2D and 3A–D). Continuously expressed Fkh dependent genes could be classified into two groups, ‘decreased’ and ‘only late expression lost’ (Figure 2D and 3E–G). All late expressed genes were similarly affected by loss of fkh and showed decreased expression (Figure 2D and 3H). The variety of expression changes in fkh mutants suggests that Fkh can both activate and repress gene expression and that SG genes are differentially regulated between early and late stages of development.


Genome-wide analysis reveals a major role in cell fate maintenance and an unexpected role in endoreduplication for the Drosophila FoxA gene Fork head.

Maruyama R, Grevengoed E, Stempniewicz P, Andrew DJ - PLoS ONE (2011)

Six groups of fkh dependent genes.In situ hybridizations of examples from each group of SG genes is shown for WT and fkh mutants. The fkh mutants are homozygous for Df(3L)H99, which blocks the SG cell death associated with fkh loss. Red arrowheads indicate the SGs. Blank arrowheads indicate the salivary duct. (A,B) Lateral views of WT and fkh embryos at stage 11 are shown. (C–H) Left two panels show lateral views of WT and fkh embryos at stage 11. Right two panels show ventral views of WT and fkh embryos at stage 13/14. SGs remain on the ventral surface of fkh mutants. (A) Fkh does not affect expression of most early SG genes, including Noa36. (B) Expression of eight early SG genes was significantly reduced in SGs of fkh mutants, as shown with CG32269. (C) Fourteen early genes affected by loss of fkh showed higher levels of expression in late embryonic SGs. This group included trh and nyo, which are initially expressed throughout the SG and duct primordia but that subsequently become restricted to the duct in WT embryos. (D) Two early expressed SG genes showed decreased expression at early stages and increased expression at late stages, as shown for the Pepck gene. (E) Expression of 27% of continuously expressed SG genes was unaffected by fkh loss. The Hmu gene is an example of this class. Hmu transcripts localize to the apical domains of SG cells. (F) 30% of continuously expressed SG genes showed reduced expression at all stages in fkh mutants, as seen with CG30497. (G) Many continuously expressed genes were unaffected at early stages but showed reduced expression at late stages, including the bHLH transcription factor gene sage. (H) All five late expressed SG genes were affected by loss of fkh, as observed with Mvl.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3116861&req=5

pone-0020901-g003: Six groups of fkh dependent genes.In situ hybridizations of examples from each group of SG genes is shown for WT and fkh mutants. The fkh mutants are homozygous for Df(3L)H99, which blocks the SG cell death associated with fkh loss. Red arrowheads indicate the SGs. Blank arrowheads indicate the salivary duct. (A,B) Lateral views of WT and fkh embryos at stage 11 are shown. (C–H) Left two panels show lateral views of WT and fkh embryos at stage 11. Right two panels show ventral views of WT and fkh embryos at stage 13/14. SGs remain on the ventral surface of fkh mutants. (A) Fkh does not affect expression of most early SG genes, including Noa36. (B) Expression of eight early SG genes was significantly reduced in SGs of fkh mutants, as shown with CG32269. (C) Fourteen early genes affected by loss of fkh showed higher levels of expression in late embryonic SGs. This group included trh and nyo, which are initially expressed throughout the SG and duct primordia but that subsequently become restricted to the duct in WT embryos. (D) Two early expressed SG genes showed decreased expression at early stages and increased expression at late stages, as shown for the Pepck gene. (E) Expression of 27% of continuously expressed SG genes was unaffected by fkh loss. The Hmu gene is an example of this class. Hmu transcripts localize to the apical domains of SG cells. (F) 30% of continuously expressed SG genes showed reduced expression at all stages in fkh mutants, as seen with CG30497. (G) Many continuously expressed genes were unaffected at early stages but showed reduced expression at late stages, including the bHLH transcription factor gene sage. (H) All five late expressed SG genes were affected by loss of fkh, as observed with Mvl.
Mentions: Further analysis revealed additional subtlety in how the early and continuously expressed classes of genes were affected by fkh loss. Early expressed genes affected by fkh loss could be classified into three groups, ‘decreased’, ‘upregulated later’ and ‘decreased and upregulated later’ (Figure 2D and 3A–D). Continuously expressed Fkh dependent genes could be classified into two groups, ‘decreased’ and ‘only late expression lost’ (Figure 2D and 3E–G). All late expressed genes were similarly affected by loss of fkh and showed decreased expression (Figure 2D and 3H). The variety of expression changes in fkh mutants suggests that Fkh can both activate and repress gene expression and that SG genes are differentially regulated between early and late stages of development.

Bottom Line: Transcription factors drive organogenesis, from the initiation of cell fate decisions to the maintenance and implementation of these decisions.Thus, unlike the worm FoxA protein PHA-4, Fkh does not function to specify cell fate.Overall, this study demonstrates an important role for Fkh in determining how an organ preserves its identity throughout development and provides an alternative paradigm for how FoxA proteins function in organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, The Johns Hopkins University, School of Medicine, Baltimore, Maryland, United States of America.

ABSTRACT
Transcription factors drive organogenesis, from the initiation of cell fate decisions to the maintenance and implementation of these decisions. The Drosophila embryonic salivary gland provides an excellent platform for unraveling the underlying transcriptional networks of organ development because Drosophila is relatively unencumbered by significant genetic redundancy. The highly conserved FoxA family transcription factors are essential for various aspects of organogenesis in all animals that have been studied. Here, we explore the role of the single Drosophila FoxA protein Fork head (Fkh) in salivary gland organogenesis using two genome-wide strategies. A large-scale in situ hybridization analysis reveals a major role for Fkh in maintaining the salivary gland fate decision and controlling salivary gland physiological activity, in addition to its previously known roles in morphogenesis and survival. The majority of salivary gland genes (59%) are affected by fkh loss, mainly at later stages of salivary gland development. We show that global expression of Fkh cannot drive ectopic salivary gland formation. Thus, unlike the worm FoxA protein PHA-4, Fkh does not function to specify cell fate. In addition, Fkh only indirectly regulates many salivary gland genes, which is also distinct from the role of PHA-4 in organogenesis. Our microarray analyses reveal unexpected roles for Fkh in blocking terminal differentiation and in endoreduplication in the salivary gland and in other Fkh-expressing embryonic tissues. Overall, this study demonstrates an important role for Fkh in determining how an organ preserves its identity throughout development and provides an alternative paradigm for how FoxA proteins function in organogenesis.

Show MeSH
Related in: MedlinePlus