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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.


dys molecular regulation and model for tarsal joint development.(A) Schematic representation of a tarsal joint of a dys640 wild type (left) and dys640Su(H)-1+2 where the two Su(H) binding sites are mutated (right). See text for description. (B) Model for tarsal joint formation. Blue and brown arrowheads mark proximal and distal joints, respectively. In a tarsal joint, Notch ligands Dl and Ser (blue) activate the Notch pathway in the distal adjacent cells. Notch, in turn, activates dys expression, which regulates the high levels of P-Mad observed in the joint. Dys also controls the expression of the pro-apoptotic genes rpr and hid and the RhoGTPases modulators RhoGef2 and RhoGap71E. The sharp boundary of Dpp signaling also regulates rpr expression via JNK pathway activation. The expression domains of E(spl)mβ, dys, rpr, hid, RhoGef2 and RhoGap71E are indicated.
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pgen-1004621-g008: dys molecular regulation and model for tarsal joint development.(A) Schematic representation of a tarsal joint of a dys640 wild type (left) and dys640Su(H)-1+2 where the two Su(H) binding sites are mutated (right). See text for description. (B) Model for tarsal joint formation. Blue and brown arrowheads mark proximal and distal joints, respectively. In a tarsal joint, Notch ligands Dl and Ser (blue) activate the Notch pathway in the distal adjacent cells. Notch, in turn, activates dys expression, which regulates the high levels of P-Mad observed in the joint. Dys also controls the expression of the pro-apoptotic genes rpr and hid and the RhoGTPases modulators RhoGef2 and RhoGap71E. The sharp boundary of Dpp signaling also regulates rpr expression via JNK pathway activation. The expression domains of E(spl)mβ, dys, rpr, hid, RhoGef2 and RhoGap71E are indicated.

Mentions: Our results show that dys expression at the presumptive tarsal segments is controlled by a dedicated CRM 640 bp long that integrates Notch signaling through direct Su(H) binding. This is, to our knowledge, the first characterized Notch direct target described for leg joint development. Interestingly, the mutation of the two identified Su(H) consensus sites, dys640-lacZSu(H)-1+2, resulted in lacZ derepression in the inter-joint domain of the tarsal segments although at lower levels compared to normal signal observed in the presumptive joints in dys640-lacZ control legs. These results are in favor of the “default repression” model in which Su(H) associates with co-repressors in the absence of Notch signaling to repress target gene transcription [39] (Figure 8A). In the event of Notch activation, co-repressors are displaced by NICD, so Su(H) binding could lead to target gene transcription through the recruitment of co-activators. Therefore, dys fulfills the two predictions of the model to occur in the absence of Su(H) binding: (1) target genes will be derepressed and (2) their expression will be reduced in their normal expression domains. These two predictions can be validated in the dys640Su(H)1+2 and dys640B gene reporter constructs, where Su(H) binding is compromised or lost. In both cases we observed a consistent derepression of lacZ expression in the inter-joint domain of the tarsal segments, although with weak levels. We also observed reduced lacZ expression at the presumptive joint domain in the dys CRM with the two Su(H) sites mutated or the dys640B reporter gene compared to the intact dys640. This characteristic is specially evident in the dys640B construct that lacks the two described Su(H) sites and probably others not identified in our analysis.


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 molecular regulation and model for tarsal joint development.(A) Schematic representation of a tarsal joint of a dys640 wild type (left) and dys640Su(H)-1+2 where the two Su(H) binding sites are mutated (right). See text for description. (B) Model for tarsal joint formation. Blue and brown arrowheads mark proximal and distal joints, respectively. In a tarsal joint, Notch ligands Dl and Ser (blue) activate the Notch pathway in the distal adjacent cells. Notch, in turn, activates dys expression, which regulates the high levels of P-Mad observed in the joint. Dys also controls the expression of the pro-apoptotic genes rpr and hid and the RhoGTPases modulators RhoGef2 and RhoGap71E. The sharp boundary of Dpp signaling also regulates rpr expression via JNK pathway activation. The expression domains of E(spl)mβ, dys, rpr, hid, RhoGef2 and RhoGap71E are indicated.
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Related In: Results  -  Collection

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pgen-1004621-g008: dys molecular regulation and model for tarsal joint development.(A) Schematic representation of a tarsal joint of a dys640 wild type (left) and dys640Su(H)-1+2 where the two Su(H) binding sites are mutated (right). See text for description. (B) Model for tarsal joint formation. Blue and brown arrowheads mark proximal and distal joints, respectively. In a tarsal joint, Notch ligands Dl and Ser (blue) activate the Notch pathway in the distal adjacent cells. Notch, in turn, activates dys expression, which regulates the high levels of P-Mad observed in the joint. Dys also controls the expression of the pro-apoptotic genes rpr and hid and the RhoGTPases modulators RhoGef2 and RhoGap71E. The sharp boundary of Dpp signaling also regulates rpr expression via JNK pathway activation. The expression domains of E(spl)mβ, dys, rpr, hid, RhoGef2 and RhoGap71E are indicated.
Mentions: Our results show that dys expression at the presumptive tarsal segments is controlled by a dedicated CRM 640 bp long that integrates Notch signaling through direct Su(H) binding. This is, to our knowledge, the first characterized Notch direct target described for leg joint development. Interestingly, the mutation of the two identified Su(H) consensus sites, dys640-lacZSu(H)-1+2, resulted in lacZ derepression in the inter-joint domain of the tarsal segments although at lower levels compared to normal signal observed in the presumptive joints in dys640-lacZ control legs. These results are in favor of the “default repression” model in which Su(H) associates with co-repressors in the absence of Notch signaling to repress target gene transcription [39] (Figure 8A). In the event of Notch activation, co-repressors are displaced by NICD, so Su(H) binding could lead to target gene transcription through the recruitment of co-activators. Therefore, dys fulfills the two predictions of the model to occur in the absence of Su(H) binding: (1) target genes will be derepressed and (2) their expression will be reduced in their normal expression domains. These two predictions can be validated in the dys640Su(H)1+2 and dys640B gene reporter constructs, where Su(H) binding is compromised or lost. In both cases we observed a consistent derepression of lacZ expression in the inter-joint domain of the tarsal segments, although with weak levels. We also observed reduced lacZ expression at the presumptive joint domain in the dys CRM with the two Su(H) sites mutated or the dys640B reporter gene compared to the intact dys640. This characteristic is specially evident in the dys640B construct that lacks the two described Su(H) sites and probably others not identified in our analysis.

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.