Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A Diagram showing the different p53 isoforms and their respective proteins. Note that p53-C and p53-A encode the same protein. Red boxes represent exons, while blue, black and green boxes denote the different protein domains. TAD-transactivation domain, DBD-DNA binding domain, and OD-oligomerization domain. The amino acid positions that define the p53 domains are indicated. B Schematic representation of the cell cycle and the different genetic tools used to block it. C , E , G Wing imaginal discs expressing the corresponding transgenes under the sal > GFP driver. Imaginal discs were stained for Dcp1 (red), GFP (green) and DAPI (blue). A dotted green line marks the sal domain. The scale bar is 50 μm. D , F , H Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( C ), ( E ), and ( G ). The data are derived from three independent biological replicates, analyzing more than 13 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 and *** P value < 0.005 and not significant (ns) P value > 0.05 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Binding Assay, Blocking Assay, Expressing, Staining, Derivative Assay

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A A simplified representation of the apoptotic pathway triggered by p53. Created with BioRender.com . B Wing imaginal discs expressing the corresponding transgenes under the sal > GFP driver. Imaginal discs were stained for Dcp1 (red), GFP or Myc as indicated (green) and DAPI (blue). A dotted green line marks the sal domain. Note that the expression of the UAS- miRHG strongly reduces the apoptotic induction by p53-A and p53-E, but not by p53-B. Scale bar is 50 μm. C Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( B ). The data are derived from three independent biological replicates, analyzing more than 14 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA. D Representative images of GFP-marked control clones and clones homozygous for Df(3 L)H99 that simultaneously expressed either p53-A or p53-B in wing imaginal discs stained for Dcp1 (red), GFP (green), and DAPI (blue). A dotted green line marks the clone boundary. The scale bar is 25 μm. E Quantification of Dcp1 staining within the GFP-marked clones from the genotypes presented in ( D ). Error bars indicate SEM. n > 14 disc per genotype. **** P value < 0.0001 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Staining, Derivative Assay, Control, Clone Assay

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A , C Wing imaginal discs expressing the corresponding transgenes under the sal > GFP driver. Where indicated p53-A and p53-B expression was performed in a dronc mutant background. Imaginal discs were stained for Dcp1 (red), GFP (green) and DAPI (blue). A dotted green line marks the sal domain. The scale bar is 50 μm. B , D Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( A ) and ( C ). The data are derived from three independent biological replicates, analyzing more than 13 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA. E Cartoon of the Dronc activity sensor. The modified GFP includes a Dronc-specific cleavage site (TETDG) that when cleaved triggers a conformational change in the protein that leads to the emission of green fluorescence. The different Dronc domains are indicated as CARD, large (L) and small (S) subunits. F Wing imaginal discs expressing the Dronc activity sensor under the sal> driver and the corresponding transgenes. The imaginal discs were stained for GFP (green), Dcp1 (white), Myc (red) and DAPI (blue). A dotted yellow line marks the sal domain delimited by Myc staining. The scale bar is 50 μm. G Quantification of GFP (Dronc activity) in the sal domain of wing imaginal discs from the genotypes presented in ( F ). The data are derived from three independent biological replicates, analyzing more than ten discs per genotype Error bars indicate SEM. **** P value < 0.0001 and not significant (ns) P value > 0.05 by one-way ANOVA. H Wing discs lysates were immunoprecipitated (IP) using anti- HA conjugated to Protein A/G magnetic beads. Input (50% of lysate) and IP eluates were resolved by SDS-PAGE and probed with primary antibody against Myc and HA. I Cartoon illustrating the BiFC principle. No fluorescent N- and C-terminal fragments of the GFP variant Venus are fused to p53-B (VN-p53) and Dronc (Dronc-VC). When both proteins are co-expressed and interact, the Venus fragment is reconstructed and emits green fluorescence. This technique enables direct protein interactions to be visualized in living cells. J Scheme showing the different versions of the p53-B protein generated for the BiFC analysis. Amino acid positions defining p53 domains are indicated. K BiFC analysis by the co-expression of VN-p53-B, Dronc-VC, Dronc-VC and VN-p53-B and or Dronc-VC and VN-p53-B ΔTAD with the sal> driver . A dotted green line marks the sal domain delimited by p53 staining. The scale bar is 50 μm. L Quantification of Venus intensity in the sal domain of wing imaginal discs from the genotypes presented in ( K ), expressed as a ratio between Venus signal and p53 staining intensity levels. The data are derived from three independent biological replicates, analyzing more than 13 discs per genotype. Error bars indicate standard deviation. **** P value < 0.0001 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Mutagenesis, Staining, Derivative Assay, Activity Assay, Modification, Fluorescence, Immunoprecipitation, Magnetic Beads, SDS Page, Variant Assay, Generated, Standard Deviation

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A–D Wing imaginal discs expressing the corresponding UAS transgenes under the sal> driver. A , B show the analysis of the activity of the hid 5′-p53 RE -GFP (green) construct, while in ( C ) and ( D ) show the analysis of the rpr 5′-p53 RE -GFP construct. In all images, the Myc (red) and DAPI (blue) channels are also shown. A dotted red line marks the sal domain. The scale bar is 50 μm. E , F Dcp1 staining (red) in wing imaginal discs expressing the indicated UAS transgenes under sal> driver. GFP or Myc staining (green), as well as DAPI staining is also shown. A green dotted line marks the sal domain. The scale bar is 50 μm. G , H Quantification of hid 5′-p53 RE -GFP or rpr- p53 RE -GFP positive area in the sal domain of wing imaginal discs from the presented genotypes. The data are derived from three independent biological replicates, analyzing more than 13 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA. I Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( E ) and ( F ). The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Activity Assay, Construct, Staining, Derivative Assay

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A , C , E , G Representative images of wing imaginaldiscs ( n > 10) expressing the corresponding transgenes with the nub> driver. When indicated the over-expression of the different versions of p53-A and p53-B was performed in a dronc mutant background. Imaginal discs were stained for GFP (green), DAPI (blue) and Dcp1 (white) in ( A ) and ( E ) or MMP1 (red), pH3 (white), DAPI (blue), Wg (cyan) and Dpp (red) in ( C ) and ( G ). A dotted green line marks the nub domain. The scale bar is 50 μm. B , F Quantification of the tumor overgrowths from the genotypes presented in ( A ) and ( E ) calculated as percentage of the nub domain. The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype Error bars indicate SEM. **** P value < 0.0001 and not significant (ns) P value > 0.05 by one-way ANOVA. D , H Quantification of the tumor overgrowths from the genotypes indicated in ( C ) and ( G ) calculated as percentage of the nub domain. The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Over Expression, Mutagenesis, Staining, Derivative Assay

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A Wing imaginal discs expressing the corresponding UAS transgenes under the sal> driver. When indicated the expression of the hp53 was performed in a dronc mutant background. Imaginal discs were stained for GFP (green), DAPI (blue), Myc (white, only when indicated) and Dcp1 (red). A dotted green line marks the sal domain. Scale bar is 50 μm. B Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( A ). The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 by one-way ANOVA. C Wing imaginal discs expressing UAS- hp53 in different conditions of cell cycle arrest stained for GFP (green), DAPI (blue) and Dcp1 (red). D Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( C ). The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 and ** P value < 0.001 by one-way ANOVA. E Wing imaginal discs expressing the Dronc activity sensor under the sal> driver and the corresponding transgenes. The imaginal discs were stained for GFP (green), Myc (red) and DAPI (blue). A dotted red line marks the sal domain delimited by Myc staining. The scale bar is 50 μm. F Quantification of GFP (Dronc activity) in the sal domain of wing imaginal discs from the genotypes presented in ( E ). Error bars indicate SEM. The data are derived from three independent biological replicates, analyzing more than ten discs per genotype. **** P value < 0.0001 by one-way ANOVA. G Wing imaginal discs expressing UAS- hp53 ΔDBD with the sal > GFP driver in a wildtype, dronc and p53 mutant backgrounds stained for GFP (green), DAPI (blue) and Dcp1 (red). A dotted green line marks the sal domain. The scale bar is 50 μm. H Quantification of Dcp1 staining in the sal domain of wing imaginal discs from the genotypes presented in ( G ). The data are derived from three independent biological replicates, analyzing more than 15 discs per genotype. Error bars indicate SEM. **** P value < 0.0001 and *** P value < 0.005 by one-way ANOVA.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Mutagenesis, Staining, Derivative Assay, Activity Assay

Journal: Cell Death & Disease

Article Title: Transcription-dependent and -independent functions of Drosophila p53 isoforms in the induction of apoptosis and senescence-associated tumorigenesis

doi: 10.1038/s41419-026-08571-x

Figure Lengend Snippet: A Wing imaginal discs expressing hp53 with the nub> driver. When indicated the expression of hp53 was combined with the UAS- miRHG or performed in a dronc mutant background. Imaginal discs were stained for GFP (green), DAPI (blue) and Dcp1 (white). A dotted green line marks the nub domain. The scale bar is 50 μm. B Quantification of the tumor overgrowths from the genotypes indicated in ( A ), calculated as a percentage of the nub domain. The data are derived from three independent biological replicates, analyzing more than ten discs per genotype. Error bars indicate SEM. **** P value < 0.0001 and not significant (ns) P value > 0.05 by one-way ANOVA. C , D Representative wildtype wing imaginal disc and disc expressing hp53 in a dronc mutant background ( n > 10) stained for GFP (green), DAPI (blue), MMP1 (red) and pH3 (white) in ( C ) or GFP (green), DAPI (blue) and EdU (white). A dotted green line marks the nub domain. Scale bar is 50 μm.

Article Snippet: Antibodies used: pH3 (Rabbit, 1:1000, MerckMillipore #06-570), pH3 (mouse, 1:1000, Cell signal technology #9706), Dcp1 (Rabbit, 1:200, Cell signal technology #9578), Myc (mouse, 1:200, DHSB #9E 10), MMP1 (mouse, 1:50, DHSB #5H7B11, #3A6B4, #3B8D), Wg (mouse, 1:50, DHSB #4D4), p53 (mouse, 1:50 DHSB #25F4) and Dpp (rabbit, 1:200) [ ].

Techniques: Expressing, Mutagenesis, Staining, Derivative Assay