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The COP9 signalosome converts temporal hormone signaling to spatial restriction on neural competence.

Huang YC, Lu YN, Wu JT, Chien CT, Pi H - PLoS Genet. (2014)

Bottom Line: We found that the COP9 signalosome (CSN) suppresses the neural competence of non-innervated bristles at the PWM.Several CSN subunits physically associate with ecdysone receptors to represses br at the transcriptional level.We propose a model in which nuclear hormone receptors cooperate with the deneddylation machinery to temporally shutdown downstream target gene expression, conferring a spatial restriction on neural competence at the PWM.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan; Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan; Insitute of Molecular Biology, Academia Sinica, Taipei, Taiwan.

ABSTRACT
During development, neural competence is conferred and maintained by integrating spatial and temporal regulations. The Drosophila sensory bristles that detect mechanical and chemical stimulations are arranged in stereotypical positions. The anterior wing margin (AWM) is arrayed with neuron-innervated sensory bristles, while posterior wing margin (PWM) bristles are non-innervated. We found that the COP9 signalosome (CSN) suppresses the neural competence of non-innervated bristles at the PWM. In CSN mutants, PWM bristles are transformed into neuron-innervated, which is attributed to sustained expression of the neural-determining factor Senseless (Sens). The CSN suppresses Sens through repression of the ecdysone signaling target gene broad (br) that encodes the BR-Z1 transcription factor to activate sens expression. Strikingly, CSN suppression of BR-Z1 is initiated at the prepupa-to-pupa transition, leading to Sens downregulation, and termination of the neural competence of PWM bristles. The role of ecdysone signaling to repress br after the prepupa-to-pupa transition is distinct from its conventional role in activation, and requires CSN deneddylating activity and multiple cullins, the major substrates of deneddylation. Several CSN subunits physically associate with ecdysone receptors to represses br at the transcriptional level. We propose a model in which nuclear hormone receptors cooperate with the deneddylation machinery to temporally shutdown downstream target gene expression, conferring a spatial restriction on neural competence at the PWM.

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BR-Z1 upregulation is required for Sens upregulation at the PWM of CSN mutants.(A–B′) Upregulation of BR-Z1 (red) in CSN4 (A, A′) and CSN5 clones (B, B′) 20–24 h APF. White arrowheads indicate the anterior-posterior boundary. (C–F) Transient overexpression of BR-Z1 in bristle lineage cells via neur-GAL4 and Tub-GAL80ts. (C–E) Ectopic expression of Sens (red in C), accumulation of Hnt (red in D) and ectopic formation of Elav-positive neurons (red in E) were observed at the PWM 22–24 h APF. Prepupae grown at 18°C were shifted to 37°C for one hour at 8–10 h APF, and then incubated at 29°C until dissection. (F) Bristles with dome-shape sockets (indicated by arrows) were observed at the PWM. Inset: enlarged figure showing single bristle. The experiment was carried out similarly to (C–E) except for incubation at 29°C for 12 hours, and returning back to 18°C until eclosion. (G–H′) Knockdown of CSN2, in MARCM clones (green), induced Sens (red) upregulation at the PWM 20–24 h APF (G, G′), which was abolished in CSN2 RNAi brnpr1 double mutant MARCM clones (green) (H, H′).
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pgen-1004760-g003: BR-Z1 upregulation is required for Sens upregulation at the PWM of CSN mutants.(A–B′) Upregulation of BR-Z1 (red) in CSN4 (A, A′) and CSN5 clones (B, B′) 20–24 h APF. White arrowheads indicate the anterior-posterior boundary. (C–F) Transient overexpression of BR-Z1 in bristle lineage cells via neur-GAL4 and Tub-GAL80ts. (C–E) Ectopic expression of Sens (red in C), accumulation of Hnt (red in D) and ectopic formation of Elav-positive neurons (red in E) were observed at the PWM 22–24 h APF. Prepupae grown at 18°C were shifted to 37°C for one hour at 8–10 h APF, and then incubated at 29°C until dissection. (F) Bristles with dome-shape sockets (indicated by arrows) were observed at the PWM. Inset: enlarged figure showing single bristle. The experiment was carried out similarly to (C–E) except for incubation at 29°C for 12 hours, and returning back to 18°C until eclosion. (G–H′) Knockdown of CSN2, in MARCM clones (green), induced Sens (red) upregulation at the PWM 20–24 h APF (G, G′), which was abolished in CSN2 RNAi brnpr1 double mutant MARCM clones (green) (H, H′).

Mentions: The Zinc-finger transcriptional factor BR-Z1 induces Sens expression in response to ecdysone signaling in chemosensory precursors [5]. Thus, we tested if the CSN also downregulates BR-Z1, leading to the suppression of Sens in PWM cells. As expected, the BR-Z1 levels were strongly elevated in CSN4 and CSN5 wing-disc cells 20–24 h APF (Figure 3A–B′). Elevated BR-Z1 levels were also detected in CSN2-knockdown cells (Figure S3A). Distinct from Sens regulation, CSN suppression of BR-Z1 was ubiquitous in all wing-disc cells, not restricted to the wing margin. Furthermore, forced expression of BR-Z1 in bristle lineage cells by neur-GAL4 elevated Sens and Hnt expression and induced Elav-positive neurons at the PWM (Figure 3C–E). In adult wings, all bristles at the PWM had dome-shape sockets (Figure 3F), suggesting that BR-Z1 promotes the formation of innervated bristles at the PWM. The br locus encodes three additional isoforms BR-Z2, BR-Z3 and BR-Z4, and functional redundancy was observed among these isoforms [46], [47]. We found that the activity to induce innervated bristles is not limited to BR-Z1, as overexpression of BR-Z3 also promoted Sens accumulation and innervated bristle formation at the PWM (Figure S3B, S3C).


The COP9 signalosome converts temporal hormone signaling to spatial restriction on neural competence.

Huang YC, Lu YN, Wu JT, Chien CT, Pi H - PLoS Genet. (2014)

BR-Z1 upregulation is required for Sens upregulation at the PWM of CSN mutants.(A–B′) Upregulation of BR-Z1 (red) in CSN4 (A, A′) and CSN5 clones (B, B′) 20–24 h APF. White arrowheads indicate the anterior-posterior boundary. (C–F) Transient overexpression of BR-Z1 in bristle lineage cells via neur-GAL4 and Tub-GAL80ts. (C–E) Ectopic expression of Sens (red in C), accumulation of Hnt (red in D) and ectopic formation of Elav-positive neurons (red in E) were observed at the PWM 22–24 h APF. Prepupae grown at 18°C were shifted to 37°C for one hour at 8–10 h APF, and then incubated at 29°C until dissection. (F) Bristles with dome-shape sockets (indicated by arrows) were observed at the PWM. Inset: enlarged figure showing single bristle. The experiment was carried out similarly to (C–E) except for incubation at 29°C for 12 hours, and returning back to 18°C until eclosion. (G–H′) Knockdown of CSN2, in MARCM clones (green), induced Sens (red) upregulation at the PWM 20–24 h APF (G, G′), which was abolished in CSN2 RNAi brnpr1 double mutant MARCM clones (green) (H, H′).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4230841&req=5

pgen-1004760-g003: BR-Z1 upregulation is required for Sens upregulation at the PWM of CSN mutants.(A–B′) Upregulation of BR-Z1 (red) in CSN4 (A, A′) and CSN5 clones (B, B′) 20–24 h APF. White arrowheads indicate the anterior-posterior boundary. (C–F) Transient overexpression of BR-Z1 in bristle lineage cells via neur-GAL4 and Tub-GAL80ts. (C–E) Ectopic expression of Sens (red in C), accumulation of Hnt (red in D) and ectopic formation of Elav-positive neurons (red in E) were observed at the PWM 22–24 h APF. Prepupae grown at 18°C were shifted to 37°C for one hour at 8–10 h APF, and then incubated at 29°C until dissection. (F) Bristles with dome-shape sockets (indicated by arrows) were observed at the PWM. Inset: enlarged figure showing single bristle. The experiment was carried out similarly to (C–E) except for incubation at 29°C for 12 hours, and returning back to 18°C until eclosion. (G–H′) Knockdown of CSN2, in MARCM clones (green), induced Sens (red) upregulation at the PWM 20–24 h APF (G, G′), which was abolished in CSN2 RNAi brnpr1 double mutant MARCM clones (green) (H, H′).
Mentions: The Zinc-finger transcriptional factor BR-Z1 induces Sens expression in response to ecdysone signaling in chemosensory precursors [5]. Thus, we tested if the CSN also downregulates BR-Z1, leading to the suppression of Sens in PWM cells. As expected, the BR-Z1 levels were strongly elevated in CSN4 and CSN5 wing-disc cells 20–24 h APF (Figure 3A–B′). Elevated BR-Z1 levels were also detected in CSN2-knockdown cells (Figure S3A). Distinct from Sens regulation, CSN suppression of BR-Z1 was ubiquitous in all wing-disc cells, not restricted to the wing margin. Furthermore, forced expression of BR-Z1 in bristle lineage cells by neur-GAL4 elevated Sens and Hnt expression and induced Elav-positive neurons at the PWM (Figure 3C–E). In adult wings, all bristles at the PWM had dome-shape sockets (Figure 3F), suggesting that BR-Z1 promotes the formation of innervated bristles at the PWM. The br locus encodes three additional isoforms BR-Z2, BR-Z3 and BR-Z4, and functional redundancy was observed among these isoforms [46], [47]. We found that the activity to induce innervated bristles is not limited to BR-Z1, as overexpression of BR-Z3 also promoted Sens accumulation and innervated bristle formation at the PWM (Figure S3B, S3C).

Bottom Line: We found that the COP9 signalosome (CSN) suppresses the neural competence of non-innervated bristles at the PWM.Several CSN subunits physically associate with ecdysone receptors to represses br at the transcriptional level.We propose a model in which nuclear hormone receptors cooperate with the deneddylation machinery to temporally shutdown downstream target gene expression, conferring a spatial restriction on neural competence at the PWM.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan; Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan; Insitute of Molecular Biology, Academia Sinica, Taipei, Taiwan.

ABSTRACT
During development, neural competence is conferred and maintained by integrating spatial and temporal regulations. The Drosophila sensory bristles that detect mechanical and chemical stimulations are arranged in stereotypical positions. The anterior wing margin (AWM) is arrayed with neuron-innervated sensory bristles, while posterior wing margin (PWM) bristles are non-innervated. We found that the COP9 signalosome (CSN) suppresses the neural competence of non-innervated bristles at the PWM. In CSN mutants, PWM bristles are transformed into neuron-innervated, which is attributed to sustained expression of the neural-determining factor Senseless (Sens). The CSN suppresses Sens through repression of the ecdysone signaling target gene broad (br) that encodes the BR-Z1 transcription factor to activate sens expression. Strikingly, CSN suppression of BR-Z1 is initiated at the prepupa-to-pupa transition, leading to Sens downregulation, and termination of the neural competence of PWM bristles. The role of ecdysone signaling to repress br after the prepupa-to-pupa transition is distinct from its conventional role in activation, and requires CSN deneddylating activity and multiple cullins, the major substrates of deneddylation. Several CSN subunits physically associate with ecdysone receptors to represses br at the transcriptional level. We propose a model in which nuclear hormone receptors cooperate with the deneddylation machinery to temporally shutdown downstream target gene expression, conferring a spatial restriction on neural competence at the PWM.

Show MeSH
Related in: MedlinePlus