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The Drosophila sterile-20 kinase slik controls cell proliferation and apoptosis during imaginal disc development.

Hipfner DR, Cohen SM - PLoS Biol. (2003)

Bottom Line: Tumor-like tissue overgrowth results when apoptosis is prevented.Activation of Raf can compensate for the lack of Slik and support cell survival, but activation of ERK cannot.We suggest that Slik mediates growth and survival cues to promote cell proliferation and control cell survival during Drosophila development.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Cell proliferation and programmed cell death are closely controlled during animal development. Proliferative stimuli generally also induce apoptosis, and anti-apoptotic factors are required to allow net cell proliferation. Genetic studies in Drosophila have led to identification of a number of genes that control both processes, providing new insights into the mechanisms that coordinate cell growth, proliferation, and death during development and that fail to do so in diseases of cell proliferation. We present evidence that the Drosophila Sterile-20 kinase Slik promotes cell proliferation and controls cell survival. At normal levels, Slik provides survival cues that prevent apoptosis. Cells deprived of Slik activity can grow, divide, and differentiate, but have an intrinsic survival defect and undergo apoptosis even under conditions in which they are not competing with normal cells for survival cues. Like some oncogenes, excess Slik activity stimulates cell proliferation, but this is compensated for by increased cell death. Tumor-like tissue overgrowth results when apoptosis is prevented. We present evidence that Slik acts via Raf, but not via the canonical ERK pathway. Activation of Raf can compensate for the lack of Slik and support cell survival, but activation of ERK cannot. We suggest that Slik mediates growth and survival cues to promote cell proliferation and control cell survival during Drosophila development.

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JNK Activity and slik-Dependent Apoptosis(A) Wing disc with a Minute+ slik1 mutant clone. Red shows Slik protein. Green shows puc–lacZ reporter gene expression visualized by anti-βGAL. Increased βGAL staining in the clone indicates puc transcription in response to JNK pathway activation.(B and C) Wing disc with large Minute+ slik1 mutant clones in an otherwise wild-type background. Blue shows activated caspase 3. Green shows actin visualized by phalloidin to show cell outlines. (B) and (C) are different optical sections of the same disc. Genotype: +/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+.(D and E) Wing disc with large Minute+ slik1 mutant clones in a hemipterous mutant background. (D) and (E) are different optical sections of the same disc. Genotype: hepr75/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+. Note the dramatic increase in clone size, relatively normal apical appearance, and reduction of apoptosis in the clones (detected by activated caspase 3 staining) when JNK pathway activity is reduced in the absence of the hep JNKK.
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pbio.0000035-g004: JNK Activity and slik-Dependent Apoptosis(A) Wing disc with a Minute+ slik1 mutant clone. Red shows Slik protein. Green shows puc–lacZ reporter gene expression visualized by anti-βGAL. Increased βGAL staining in the clone indicates puc transcription in response to JNK pathway activation.(B and C) Wing disc with large Minute+ slik1 mutant clones in an otherwise wild-type background. Blue shows activated caspase 3. Green shows actin visualized by phalloidin to show cell outlines. (B) and (C) are different optical sections of the same disc. Genotype: +/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+.(D and E) Wing disc with large Minute+ slik1 mutant clones in a hemipterous mutant background. (D) and (E) are different optical sections of the same disc. Genotype: hepr75/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+. Note the dramatic increase in clone size, relatively normal apical appearance, and reduction of apoptosis in the clones (detected by activated caspase 3 staining) when JNK pathway activity is reduced in the absence of the hep JNKK.

Mentions: The c-Jun N-terminal kinase (JNK) pathway is an effector of apoptotic cell death during imaginal disc development (Adachi-Yamada et al. 1999; Moreno et al. 2002a). JNK pathway activity can be monitored in the discs by transcriptional activation of puckered (puc), a dual-specificity phosphatase that acts in a negative-feedback loop to regulate the JNK pathway (Martin-Blanco et al. 1998). Transcription of a puc–lacZ reporter gene is normally low or absent in the wing disc. However, in response to apoptotic stimuli, puc–lacZ is induced in a JNK-dependent manner (Adachi-Yamada et al. 1999). We observed that puc–lacZ was induced in Minute+ slik1 mutant clones (Figure 4A), indicating apoptosis of slik mutant cells involves recruitment of the JNK pathway to amplify the apoptotic trigger. To confirm this, we compared the level of apoptosis in slik mutant clones induced in flies lacking the hemipterous (hep) gene, which encodes the kinase that activates JNK (JNKK). Removing Hep activity has been shown to suppress JNK-induced apoptosis (Adachi-Yamada et al. 1999). Apoptosis was visualized using an antibody to the activated form of caspase 3. Activated caspase was readily detected throughout the Minute+ slik1 mutant clones in a wild-type background (Figure 4B and 4C). The staining was most prominent in basal optical sections of the disc. Using phalloidin to label cortical actin, we observed that dying Slik-negative cells were extruded out the basal surface of the epithelium. Large Minute+ slik1 mutant clones in a hep- background caused much less distortion of the discs and looked essentially normal in apical optical sections (Figure 4D). Levels of activated caspase were strongly reduced in these clones compared to slik mutant clones in the JNKK+ background, even though many mutant cells were extruded on the basal surface of the epithelium (Figure 4D and 4E). These observations indicate that activation of the JNK pathway contributes to apoptosis in slik mutant clones.


The Drosophila sterile-20 kinase slik controls cell proliferation and apoptosis during imaginal disc development.

Hipfner DR, Cohen SM - PLoS Biol. (2003)

JNK Activity and slik-Dependent Apoptosis(A) Wing disc with a Minute+ slik1 mutant clone. Red shows Slik protein. Green shows puc–lacZ reporter gene expression visualized by anti-βGAL. Increased βGAL staining in the clone indicates puc transcription in response to JNK pathway activation.(B and C) Wing disc with large Minute+ slik1 mutant clones in an otherwise wild-type background. Blue shows activated caspase 3. Green shows actin visualized by phalloidin to show cell outlines. (B) and (C) are different optical sections of the same disc. Genotype: +/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+.(D and E) Wing disc with large Minute+ slik1 mutant clones in a hemipterous mutant background. (D) and (E) are different optical sections of the same disc. Genotype: hepr75/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+. Note the dramatic increase in clone size, relatively normal apical appearance, and reduction of apoptosis in the clones (detected by activated caspase 3 staining) when JNK pathway activity is reduced in the absence of the hep JNKK.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC261876&req=5

pbio.0000035-g004: JNK Activity and slik-Dependent Apoptosis(A) Wing disc with a Minute+ slik1 mutant clone. Red shows Slik protein. Green shows puc–lacZ reporter gene expression visualized by anti-βGAL. Increased βGAL staining in the clone indicates puc transcription in response to JNK pathway activation.(B and C) Wing disc with large Minute+ slik1 mutant clones in an otherwise wild-type background. Blue shows activated caspase 3. Green shows actin visualized by phalloidin to show cell outlines. (B) and (C) are different optical sections of the same disc. Genotype: +/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+.(D and E) Wing disc with large Minute+ slik1 mutant clones in a hemipterous mutant background. (D) and (E) are different optical sections of the same disc. Genotype: hepr75/Y; FRT42D P(πmyc) M(2)531/FRT42D slik1; hsFLP388/+. Note the dramatic increase in clone size, relatively normal apical appearance, and reduction of apoptosis in the clones (detected by activated caspase 3 staining) when JNK pathway activity is reduced in the absence of the hep JNKK.
Mentions: The c-Jun N-terminal kinase (JNK) pathway is an effector of apoptotic cell death during imaginal disc development (Adachi-Yamada et al. 1999; Moreno et al. 2002a). JNK pathway activity can be monitored in the discs by transcriptional activation of puckered (puc), a dual-specificity phosphatase that acts in a negative-feedback loop to regulate the JNK pathway (Martin-Blanco et al. 1998). Transcription of a puc–lacZ reporter gene is normally low or absent in the wing disc. However, in response to apoptotic stimuli, puc–lacZ is induced in a JNK-dependent manner (Adachi-Yamada et al. 1999). We observed that puc–lacZ was induced in Minute+ slik1 mutant clones (Figure 4A), indicating apoptosis of slik mutant cells involves recruitment of the JNK pathway to amplify the apoptotic trigger. To confirm this, we compared the level of apoptosis in slik mutant clones induced in flies lacking the hemipterous (hep) gene, which encodes the kinase that activates JNK (JNKK). Removing Hep activity has been shown to suppress JNK-induced apoptosis (Adachi-Yamada et al. 1999). Apoptosis was visualized using an antibody to the activated form of caspase 3. Activated caspase was readily detected throughout the Minute+ slik1 mutant clones in a wild-type background (Figure 4B and 4C). The staining was most prominent in basal optical sections of the disc. Using phalloidin to label cortical actin, we observed that dying Slik-negative cells were extruded out the basal surface of the epithelium. Large Minute+ slik1 mutant clones in a hep- background caused much less distortion of the discs and looked essentially normal in apical optical sections (Figure 4D). Levels of activated caspase were strongly reduced in these clones compared to slik mutant clones in the JNKK+ background, even though many mutant cells were extruded on the basal surface of the epithelium (Figure 4D and 4E). These observations indicate that activation of the JNK pathway contributes to apoptosis in slik mutant clones.

Bottom Line: Tumor-like tissue overgrowth results when apoptosis is prevented.Activation of Raf can compensate for the lack of Slik and support cell survival, but activation of ERK cannot.We suggest that Slik mediates growth and survival cues to promote cell proliferation and control cell survival during Drosophila development.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, Heidelberg, Germany.

ABSTRACT
Cell proliferation and programmed cell death are closely controlled during animal development. Proliferative stimuli generally also induce apoptosis, and anti-apoptotic factors are required to allow net cell proliferation. Genetic studies in Drosophila have led to identification of a number of genes that control both processes, providing new insights into the mechanisms that coordinate cell growth, proliferation, and death during development and that fail to do so in diseases of cell proliferation. We present evidence that the Drosophila Sterile-20 kinase Slik promotes cell proliferation and controls cell survival. At normal levels, Slik provides survival cues that prevent apoptosis. Cells deprived of Slik activity can grow, divide, and differentiate, but have an intrinsic survival defect and undergo apoptosis even under conditions in which they are not competing with normal cells for survival cues. Like some oncogenes, excess Slik activity stimulates cell proliferation, but this is compensated for by increased cell death. Tumor-like tissue overgrowth results when apoptosis is prevented. We present evidence that Slik acts via Raf, but not via the canonical ERK pathway. Activation of Raf can compensate for the lack of Slik and support cell survival, but activation of ERK cannot. We suggest that Slik mediates growth and survival cues to promote cell proliferation and control cell survival during Drosophila development.

Show MeSH
Related in: MedlinePlus