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Loss of putzig Activity Results in Apoptosis during Wing Imaginal Development in Drosophila.

Zimmermann M, Kugler SJ, Schulz A, Nagel AC - PLoS ONE (2015)

Bottom Line: We present evidence that downregulation of Notch activates Dcp1 caspase and JNK signaling, however, neither induces ectopic wg nor dpp expression.In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction.Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two are required together to maintain cell and tissue homeostasis in Drosophila.

View Article: PubMed Central - PubMed

Affiliation: Institute of Genetics, University of Hohenheim, 70599 Stuttgart, Germany.

ABSTRACT
The Drosophila gene putzig (pzg) encodes a nuclear protein that is an integral component of the Trf2/Dref complex involved in the transcription of proliferation-related genes. Moreover, Pzg is found in a complex together with the nucleosome remodeling factor NURF, where it promotes Notch target gene activation. Here we show that downregulation of pzg activity in the developing wing imaginal discs induces an apoptotic response, accompanied by the induction of the pro-apoptotic gene reaper, repression of Drosophila inhibitor of apoptosis protein accumulation and the activation of the caspases Drice, Caspase3 and Dcp1. As a further consequence 'Apoptosis induced Proliferation' (AiP) and 'Apoptosis induced Apoptosis' (AiA) are triggered. As expected, the activity of the stress kinase Jun N-terminal kinase (JNK), proposed to mediate both processes, is ectopically induced in response to pzg loss. In addition, the expression of the mitogen wingless (wg) but not of decapentaplegic (dpp) is observed. We present evidence that downregulation of Notch activates Dcp1 caspase and JNK signaling, however, neither induces ectopic wg nor dpp expression. In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction. Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two are required together to maintain cell and tissue homeostasis in Drosophila.

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pzg depletion autonomously triggers the apoptotic signaling cascade in wing imaginal discs.RNAi mediated depletion of pzg was induced in the posterior part of the wing disc using en-Gal4. Rpr-lacZ, DIAP1 and caspase activity (in red) was monitored as indicated. (A-A''') A strong activation of the pro-apoptotic gene rpr (arrows) as well as the Drosophila activated caspases Driceact (C-C''', arrow), Caspase 3act (D-D''', arrow) and Dcp-1act (E-E''', arrows) is detected in the posterior half of the disc, whereas DIAP1 protein level is reduced (B-B''', repressive arrow). (A-A''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+; rpr-lacZ/+. (B-E''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+. Pzg protein is shown in blue (anti-Pzg, A-E and A'''- E'''); GFP in green (en-Gal4 GFP) marks the posterior compartment (A-E and A''-E''). Posterior is to the right and dorsal up. The antero-posterior compartment boundary is marked with a dashed line. Scale bars: 100 μm.
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pone.0124652.g002: pzg depletion autonomously triggers the apoptotic signaling cascade in wing imaginal discs.RNAi mediated depletion of pzg was induced in the posterior part of the wing disc using en-Gal4. Rpr-lacZ, DIAP1 and caspase activity (in red) was monitored as indicated. (A-A''') A strong activation of the pro-apoptotic gene rpr (arrows) as well as the Drosophila activated caspases Driceact (C-C''', arrow), Caspase 3act (D-D''', arrow) and Dcp-1act (E-E''', arrows) is detected in the posterior half of the disc, whereas DIAP1 protein level is reduced (B-B''', repressive arrow). (A-A''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+; rpr-lacZ/+. (B-E''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+. Pzg protein is shown in blue (anti-Pzg, A-E and A'''- E'''); GFP in green (en-Gal4 GFP) marks the posterior compartment (A-E and A''-E''). Posterior is to the right and dorsal up. The antero-posterior compartment boundary is marked with a dashed line. Scale bars: 100 μm.

Mentions: Pzg activity was downregulated with the UAS-pzg-RNAi shown before to efficiently reduce Pzg protein levels [40]. Pzg-RNAi induction either in the posterior compartment (using en-Gal4) or in a more central area (using omb-Gal4) of the wing disc triggered the execution of the apoptotic program: Pro-apoptotic gene activity, visualized by the activation of a rpr-lacZ reporter (Fig 2A-A''' and S2A-A'' Fig), a reduced level of the anti-apoptotic protein DIAP1 (Fig 2B-B''' and S2B-B'' Fig) and finally the accumulation of activated initiator and effector caspases Drice, Caspase-3 and Dcp-1 were observed in the pzg mutant area of the wing disc (Fig 2C-E''' and S2C-E'' Fig). To exclude, however, off-target effects, a second independent RNAi-line (VDRC v25542) was included in the analysis of apoptosis induction and gave the same overall results. These data show that an impaired pzg activity leads to an increase in apoptosis.


Loss of putzig Activity Results in Apoptosis during Wing Imaginal Development in Drosophila.

Zimmermann M, Kugler SJ, Schulz A, Nagel AC - PLoS ONE (2015)

pzg depletion autonomously triggers the apoptotic signaling cascade in wing imaginal discs.RNAi mediated depletion of pzg was induced in the posterior part of the wing disc using en-Gal4. Rpr-lacZ, DIAP1 and caspase activity (in red) was monitored as indicated. (A-A''') A strong activation of the pro-apoptotic gene rpr (arrows) as well as the Drosophila activated caspases Driceact (C-C''', arrow), Caspase 3act (D-D''', arrow) and Dcp-1act (E-E''', arrows) is detected in the posterior half of the disc, whereas DIAP1 protein level is reduced (B-B''', repressive arrow). (A-A''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+; rpr-lacZ/+. (B-E''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+. Pzg protein is shown in blue (anti-Pzg, A-E and A'''- E'''); GFP in green (en-Gal4 GFP) marks the posterior compartment (A-E and A''-E''). Posterior is to the right and dorsal up. The antero-posterior compartment boundary is marked with a dashed line. Scale bars: 100 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4403878&req=5

pone.0124652.g002: pzg depletion autonomously triggers the apoptotic signaling cascade in wing imaginal discs.RNAi mediated depletion of pzg was induced in the posterior part of the wing disc using en-Gal4. Rpr-lacZ, DIAP1 and caspase activity (in red) was monitored as indicated. (A-A''') A strong activation of the pro-apoptotic gene rpr (arrows) as well as the Drosophila activated caspases Driceact (C-C''', arrow), Caspase 3act (D-D''', arrow) and Dcp-1act (E-E''', arrows) is detected in the posterior half of the disc, whereas DIAP1 protein level is reduced (B-B''', repressive arrow). (A-A''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+; rpr-lacZ/+. (B-E''') en-Gal4 UAS-GFP UAS-pzg-RNAi/+. Pzg protein is shown in blue (anti-Pzg, A-E and A'''- E'''); GFP in green (en-Gal4 GFP) marks the posterior compartment (A-E and A''-E''). Posterior is to the right and dorsal up. The antero-posterior compartment boundary is marked with a dashed line. Scale bars: 100 μm.
Mentions: Pzg activity was downregulated with the UAS-pzg-RNAi shown before to efficiently reduce Pzg protein levels [40]. Pzg-RNAi induction either in the posterior compartment (using en-Gal4) or in a more central area (using omb-Gal4) of the wing disc triggered the execution of the apoptotic program: Pro-apoptotic gene activity, visualized by the activation of a rpr-lacZ reporter (Fig 2A-A''' and S2A-A'' Fig), a reduced level of the anti-apoptotic protein DIAP1 (Fig 2B-B''' and S2B-B'' Fig) and finally the accumulation of activated initiator and effector caspases Drice, Caspase-3 and Dcp-1 were observed in the pzg mutant area of the wing disc (Fig 2C-E''' and S2C-E'' Fig). To exclude, however, off-target effects, a second independent RNAi-line (VDRC v25542) was included in the analysis of apoptosis induction and gave the same overall results. These data show that an impaired pzg activity leads to an increase in apoptosis.

Bottom Line: We present evidence that downregulation of Notch activates Dcp1 caspase and JNK signaling, however, neither induces ectopic wg nor dpp expression.In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction.Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two are required together to maintain cell and tissue homeostasis in Drosophila.

View Article: PubMed Central - PubMed

Affiliation: Institute of Genetics, University of Hohenheim, 70599 Stuttgart, Germany.

ABSTRACT
The Drosophila gene putzig (pzg) encodes a nuclear protein that is an integral component of the Trf2/Dref complex involved in the transcription of proliferation-related genes. Moreover, Pzg is found in a complex together with the nucleosome remodeling factor NURF, where it promotes Notch target gene activation. Here we show that downregulation of pzg activity in the developing wing imaginal discs induces an apoptotic response, accompanied by the induction of the pro-apoptotic gene reaper, repression of Drosophila inhibitor of apoptosis protein accumulation and the activation of the caspases Drice, Caspase3 and Dcp1. As a further consequence 'Apoptosis induced Proliferation' (AiP) and 'Apoptosis induced Apoptosis' (AiA) are triggered. As expected, the activity of the stress kinase Jun N-terminal kinase (JNK), proposed to mediate both processes, is ectopically induced in response to pzg loss. In addition, the expression of the mitogen wingless (wg) but not of decapentaplegic (dpp) is observed. We present evidence that downregulation of Notch activates Dcp1 caspase and JNK signaling, however, neither induces ectopic wg nor dpp expression. In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction. Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two are required together to maintain cell and tissue homeostasis in Drosophila.

Show MeSH
Related in: MedlinePlus