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The bHLH-PAS transcription factor dysfusion regulates tarsal joint formation in response to Notch activity during drosophila leg development.

Córdoba S, Estella C - PLoS Genet. (2014)

Bottom Line: This novel Dys function depends on its obligated partner Tango to activate the transcription of target genes.We also identified a dedicated dys cis-regulatory module that regulates dys expression in the tarsal presumptive leg joints through direct Su(H) binding.All these data place dys as a key player downstream of Notch, directing distal versus proximal joint morphogenesis.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología Molecular and Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid (UAM), Madrid, Spain.

ABSTRACT
A characteristic of all arthropods is the presence of flexible structures called joints that connect all leg segments. Drosophila legs include two types of joints: the proximal or "true" joints that are motile due to the presence of muscle attachment and the distal joints that lack musculature. These joints are not only morphologically, functionally and evolutionarily different, but also the morphogenetic program that forms them is distinct. Development of both proximal and distal joints requires Notch activity; however, it is still unknown how this pathway can control the development of such homologous although distinct structures. Here we show that the bHLH-PAS transcription factor encoded by the gene dysfusion (dys), is expressed and absolutely required for tarsal joint development while it is dispensable for proximal joints. In the presumptive tarsal joints, Dys regulates the expression of the pro-apoptotic genes reaper and head involution defective and the expression of the RhoGTPases modulators, RhoGEf2 and RhoGap71E, thus directing key morphogenetic events required for tarsal joint development. When ectopically expressed, dys is able to induce some aspects of the morphogenetic program necessary for distal joint development such as fold formation and programmed cell death. This novel Dys function depends on its obligated partner Tango to activate the transcription of target genes. We also identified a dedicated dys cis-regulatory module that regulates dys expression in the tarsal presumptive leg joints through direct Su(H) binding. All these data place dys as a key player downstream of Notch, directing distal versus proximal joint morphogenesis.

No MeSH data available.


Related in: MedlinePlus

dys relationship with the Notch pathway.(A) ptc-Gal4, UAS-GFP; UAS-Notch-RNAi prepupal leg disc. Knockdown of Notch levels in the ptc domain (green and arrow in A′) downregulates dys expression (red and single channel in A′). (B) ptc-Gal4, UAS-GFP; UAS-NotchICD prepupal leg disc. Notch pathway activation in the ptc domain (green and arrow in B′) induced dys expression (red and single channel in B′) in the tarsal segments but not in more proximal ones (asterisk). (C) dpp-Gal4. UAS-GFP; UAS-dys-RNAi prepupal leg disc. Dys knockdown in the dpp domain (green and arrow in C) downregulates bib-Z expression in the tarsal segments (red and single channel in C′). Note that bib expression in the tibial/tarsal (tib) and the tarsal/pretarsal (pre) joints remain unaffected (asterisks). (D) dys2/dys3 mutant prepupal leg disc where bib-Z expression (red and single channel in D′) is downregulated in the tarsal segments while in the tibial/tarsal or the tarsal/pretarsal joints remain unaffected (asterisks). Discs-large (Dlg) is in green. (E) en-Gal4, UAS-GFP; UAS-dys-RNAi prepupal leg discs. Knockdown of Dys levels in the posterior compartment (green and arrow) slightly downregulated E(spl)mβ-CD2 expression in the tarsal segments (red and single channel in E′). (F) dys2/dys3 mutant prepupal leg disc. E(spl)mβ-CD2 (red and single channel in F′) is still active in the presumptive tarsal joints. Dlg is in green. (G) Dl expression pattern (green and arrows) remains unaffected in dys2/dys3 mutant prepupal leg. Dlg is in red and single channel for Dl is in (G′). (H–N) Distal adult legs of the following genotypes: (H) wild type, (I) Notchtsa,(J) ptc-Gal4; UAS-dys; tubGal80ts, (K) Notchtsa; ptc-Gal4, UAS-dys; tubGal80ts, (L) dys2/dys3, (M) ptc-Gal4; UAS-NotchICD; tubGal80ts and (N) ptc-Gal4; UAS-NotchICD; tubGal80ts; dys2/dys3. Normal tarsal joint formation is pointed out with arrowheads while ectopic folds along the PD axis are marked with arrows. Note the absence of tarsal joints in Notchtsa (I) and dys2/dys3 (L) mutant legs, and the ectopic folds after dys (J) or NotchICD (M) misexpression in a wild type background and in a Notchtsa (K) or dys2/dys3 (N) mutant background.
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pgen-1004621-g003: dys relationship with the Notch pathway.(A) ptc-Gal4, UAS-GFP; UAS-Notch-RNAi prepupal leg disc. Knockdown of Notch levels in the ptc domain (green and arrow in A′) downregulates dys expression (red and single channel in A′). (B) ptc-Gal4, UAS-GFP; UAS-NotchICD prepupal leg disc. Notch pathway activation in the ptc domain (green and arrow in B′) induced dys expression (red and single channel in B′) in the tarsal segments but not in more proximal ones (asterisk). (C) dpp-Gal4. UAS-GFP; UAS-dys-RNAi prepupal leg disc. Dys knockdown in the dpp domain (green and arrow in C) downregulates bib-Z expression in the tarsal segments (red and single channel in C′). Note that bib expression in the tibial/tarsal (tib) and the tarsal/pretarsal (pre) joints remain unaffected (asterisks). (D) dys2/dys3 mutant prepupal leg disc where bib-Z expression (red and single channel in D′) is downregulated in the tarsal segments while in the tibial/tarsal or the tarsal/pretarsal joints remain unaffected (asterisks). Discs-large (Dlg) is in green. (E) en-Gal4, UAS-GFP; UAS-dys-RNAi prepupal leg discs. Knockdown of Dys levels in the posterior compartment (green and arrow) slightly downregulated E(spl)mβ-CD2 expression in the tarsal segments (red and single channel in E′). (F) dys2/dys3 mutant prepupal leg disc. E(spl)mβ-CD2 (red and single channel in F′) is still active in the presumptive tarsal joints. Dlg is in green. (G) Dl expression pattern (green and arrows) remains unaffected in dys2/dys3 mutant prepupal leg. Dlg is in red and single channel for Dl is in (G′). (H–N) Distal adult legs of the following genotypes: (H) wild type, (I) Notchtsa,(J) ptc-Gal4; UAS-dys; tubGal80ts, (K) Notchtsa; ptc-Gal4, UAS-dys; tubGal80ts, (L) dys2/dys3, (M) ptc-Gal4; UAS-NotchICD; tubGal80ts and (N) ptc-Gal4; UAS-NotchICD; tubGal80ts; dys2/dys3. Normal tarsal joint formation is pointed out with arrowheads while ectopic folds along the PD axis are marked with arrows. Note the absence of tarsal joints in Notchtsa (I) and dys2/dys3 (L) mutant legs, and the ectopic folds after dys (J) or NotchICD (M) misexpression in a wild type background and in a Notchtsa (K) or dys2/dys3 (N) mutant background.

Mentions: To test whether dys is sufficient to induce joint-like structures in the leg, we ectopically expressed dys in an anterior row of cells along the PD axis of the leg disc using the patched (ptc)-Gal4 line. We restricted dys ectopic expression to mid-third instar stage using the Gal80ts technique (see Material and Methods). dys misexpression induces the formation of cuticle folds along the PD axis of the leg that resembles ectopic joint formation (Figure 2F). These joint-like structures are more evident in the tarsal region, although we also detected ectopic folds in more proximal domains such as the tibia or the femur (Figure S2D). These phenotypes are very similar to the ones described for ectopic Notch pathway activation in the leg [12], [14] (see Figure 3M). Although we can not conclude that ectopic dys is able to induce the complete joint architecture, which would include the ball-and-socket structure, we were able to observe a phenotype that recapitulates some major aspects of joint formation such as indentation of the cuticle and fold formation. Taken together, our results suggest that dys is necessary for tarsal joint formation and sufficient to induce joint-like structures.


The bHLH-PAS transcription factor dysfusion regulates tarsal joint formation in response to Notch activity during drosophila leg development.

Córdoba S, Estella C - PLoS Genet. (2014)

dys relationship with the Notch pathway.(A) ptc-Gal4, UAS-GFP; UAS-Notch-RNAi prepupal leg disc. Knockdown of Notch levels in the ptc domain (green and arrow in A′) downregulates dys expression (red and single channel in A′). (B) ptc-Gal4, UAS-GFP; UAS-NotchICD prepupal leg disc. Notch pathway activation in the ptc domain (green and arrow in B′) induced dys expression (red and single channel in B′) in the tarsal segments but not in more proximal ones (asterisk). (C) dpp-Gal4. UAS-GFP; UAS-dys-RNAi prepupal leg disc. Dys knockdown in the dpp domain (green and arrow in C) downregulates bib-Z expression in the tarsal segments (red and single channel in C′). Note that bib expression in the tibial/tarsal (tib) and the tarsal/pretarsal (pre) joints remain unaffected (asterisks). (D) dys2/dys3 mutant prepupal leg disc where bib-Z expression (red and single channel in D′) is downregulated in the tarsal segments while in the tibial/tarsal or the tarsal/pretarsal joints remain unaffected (asterisks). Discs-large (Dlg) is in green. (E) en-Gal4, UAS-GFP; UAS-dys-RNAi prepupal leg discs. Knockdown of Dys levels in the posterior compartment (green and arrow) slightly downregulated E(spl)mβ-CD2 expression in the tarsal segments (red and single channel in E′). (F) dys2/dys3 mutant prepupal leg disc. E(spl)mβ-CD2 (red and single channel in F′) is still active in the presumptive tarsal joints. Dlg is in green. (G) Dl expression pattern (green and arrows) remains unaffected in dys2/dys3 mutant prepupal leg. Dlg is in red and single channel for Dl is in (G′). (H–N) Distal adult legs of the following genotypes: (H) wild type, (I) Notchtsa,(J) ptc-Gal4; UAS-dys; tubGal80ts, (K) Notchtsa; ptc-Gal4, UAS-dys; tubGal80ts, (L) dys2/dys3, (M) ptc-Gal4; UAS-NotchICD; tubGal80ts and (N) ptc-Gal4; UAS-NotchICD; tubGal80ts; dys2/dys3. Normal tarsal joint formation is pointed out with arrowheads while ectopic folds along the PD axis are marked with arrows. Note the absence of tarsal joints in Notchtsa (I) and dys2/dys3 (L) mutant legs, and the ectopic folds after dys (J) or NotchICD (M) misexpression in a wild type background and in a Notchtsa (K) or dys2/dys3 (N) mutant background.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4199481&req=5

pgen-1004621-g003: dys relationship with the Notch pathway.(A) ptc-Gal4, UAS-GFP; UAS-Notch-RNAi prepupal leg disc. Knockdown of Notch levels in the ptc domain (green and arrow in A′) downregulates dys expression (red and single channel in A′). (B) ptc-Gal4, UAS-GFP; UAS-NotchICD prepupal leg disc. Notch pathway activation in the ptc domain (green and arrow in B′) induced dys expression (red and single channel in B′) in the tarsal segments but not in more proximal ones (asterisk). (C) dpp-Gal4. UAS-GFP; UAS-dys-RNAi prepupal leg disc. Dys knockdown in the dpp domain (green and arrow in C) downregulates bib-Z expression in the tarsal segments (red and single channel in C′). Note that bib expression in the tibial/tarsal (tib) and the tarsal/pretarsal (pre) joints remain unaffected (asterisks). (D) dys2/dys3 mutant prepupal leg disc where bib-Z expression (red and single channel in D′) is downregulated in the tarsal segments while in the tibial/tarsal or the tarsal/pretarsal joints remain unaffected (asterisks). Discs-large (Dlg) is in green. (E) en-Gal4, UAS-GFP; UAS-dys-RNAi prepupal leg discs. Knockdown of Dys levels in the posterior compartment (green and arrow) slightly downregulated E(spl)mβ-CD2 expression in the tarsal segments (red and single channel in E′). (F) dys2/dys3 mutant prepupal leg disc. E(spl)mβ-CD2 (red and single channel in F′) is still active in the presumptive tarsal joints. Dlg is in green. (G) Dl expression pattern (green and arrows) remains unaffected in dys2/dys3 mutant prepupal leg. Dlg is in red and single channel for Dl is in (G′). (H–N) Distal adult legs of the following genotypes: (H) wild type, (I) Notchtsa,(J) ptc-Gal4; UAS-dys; tubGal80ts, (K) Notchtsa; ptc-Gal4, UAS-dys; tubGal80ts, (L) dys2/dys3, (M) ptc-Gal4; UAS-NotchICD; tubGal80ts and (N) ptc-Gal4; UAS-NotchICD; tubGal80ts; dys2/dys3. Normal tarsal joint formation is pointed out with arrowheads while ectopic folds along the PD axis are marked with arrows. Note the absence of tarsal joints in Notchtsa (I) and dys2/dys3 (L) mutant legs, and the ectopic folds after dys (J) or NotchICD (M) misexpression in a wild type background and in a Notchtsa (K) or dys2/dys3 (N) mutant background.
Mentions: To test whether dys is sufficient to induce joint-like structures in the leg, we ectopically expressed dys in an anterior row of cells along the PD axis of the leg disc using the patched (ptc)-Gal4 line. We restricted dys ectopic expression to mid-third instar stage using the Gal80ts technique (see Material and Methods). dys misexpression induces the formation of cuticle folds along the PD axis of the leg that resembles ectopic joint formation (Figure 2F). These joint-like structures are more evident in the tarsal region, although we also detected ectopic folds in more proximal domains such as the tibia or the femur (Figure S2D). These phenotypes are very similar to the ones described for ectopic Notch pathway activation in the leg [12], [14] (see Figure 3M). Although we can not conclude that ectopic dys is able to induce the complete joint architecture, which would include the ball-and-socket structure, we were able to observe a phenotype that recapitulates some major aspects of joint formation such as indentation of the cuticle and fold formation. Taken together, our results suggest that dys is necessary for tarsal joint formation and sufficient to induce joint-like structures.

Bottom Line: This novel Dys function depends on its obligated partner Tango to activate the transcription of target genes.We also identified a dedicated dys cis-regulatory module that regulates dys expression in the tarsal presumptive leg joints through direct Su(H) binding.All these data place dys as a key player downstream of Notch, directing distal versus proximal joint morphogenesis.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología Molecular and Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid (UAM), Madrid, Spain.

ABSTRACT
A characteristic of all arthropods is the presence of flexible structures called joints that connect all leg segments. Drosophila legs include two types of joints: the proximal or "true" joints that are motile due to the presence of muscle attachment and the distal joints that lack musculature. These joints are not only morphologically, functionally and evolutionarily different, but also the morphogenetic program that forms them is distinct. Development of both proximal and distal joints requires Notch activity; however, it is still unknown how this pathway can control the development of such homologous although distinct structures. Here we show that the bHLH-PAS transcription factor encoded by the gene dysfusion (dys), is expressed and absolutely required for tarsal joint development while it is dispensable for proximal joints. In the presumptive tarsal joints, Dys regulates the expression of the pro-apoptotic genes reaper and head involution defective and the expression of the RhoGTPases modulators, RhoGEf2 and RhoGap71E, thus directing key morphogenetic events required for tarsal joint development. When ectopically expressed, dys is able to induce some aspects of the morphogenetic program necessary for distal joint development such as fold formation and programmed cell death. This novel Dys function depends on its obligated partner Tango to activate the transcription of target genes. We also identified a dedicated dys cis-regulatory module that regulates dys expression in the tarsal presumptive leg joints through direct Su(H) binding. All these data place dys as a key player downstream of Notch, directing distal versus proximal joint morphogenesis.

No MeSH data available.


Related in: MedlinePlus