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Genetic interactions between the Drosophila tumor suppressor gene ept and the stat92E transcription factor.

Gilbert MM, Beam CK, Robinson BS, Moberg KH - PLoS ONE (2009)

Bottom Line: We find that this phenotype is very sensitive to the genetic dose of stat92E, the transcriptional effector of the Jak-Stat signaling pathway, and that this pathway undergoes strong activation in ept mutant cells.Genetic evidence indicates that stat92E contributes to cell cycle deregulation and excess cell size phenotypes that are observed among ept mutant cells.These findings identify ept as a cell-autonomous inhibitor of the Jak-Stat pathway and suggest that excess Jak-Stat signaling makes a significant contribution to proliferative and tissue architectural phenotypes that occur in ept mutant tissues.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Emory University School of Medicine, Atlanta, Georgia, United States of America.

ABSTRACT

Background: Tumor Susceptibility Gene-101 (TSG101) promotes the endocytic degradation of transmembrane proteins and is implicated as a mutational target in cancer, yet the effect of TSG101 loss on cell proliferation in vertebrates is uncertain. By contrast, Drosophila epithelial tissues lacking the TSG101 ortholog erupted (ept) develop as enlarged undifferentiated tumors, indicating that the gene can have anti-growth properties in a simple metazoan. A full understanding of pathways deregulated by loss of Drosophila ept will aid in understanding potential links between mammalian TSG101 and growth control.

Principal findings: We have taken a genetic approach to the identification of pathways required for excess growth of Drosophila eye-antennal imaginal discs lacking ept. We find that this phenotype is very sensitive to the genetic dose of stat92E, the transcriptional effector of the Jak-Stat signaling pathway, and that this pathway undergoes strong activation in ept mutant cells. Genetic evidence indicates that stat92E contributes to cell cycle deregulation and excess cell size phenotypes that are observed among ept mutant cells. In addition, autonomous Stat92E hyper-activation is associated with altered tissue architecture in ept tumors and an effect on expression of the apical polarity determinant crumbs.

Conclusions: These findings identify ept as a cell-autonomous inhibitor of the Jak-Stat pathway and suggest that excess Jak-Stat signaling makes a significant contribution to proliferative and tissue architectural phenotypes that occur in ept mutant tissues.

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stat92E affects tissue architecture in ept tumors.Confocal images of ept2/M(3) (A-A″) and ept2/M(3),stat92E06346/+ (B-B″) eye discs stained for Dlg (red), Crb (blue) and DNA (green). The two main tissue lobes in panel B are separated by the esophagus (e), which remained embedded in the tissue mass during dissection. Areas outlined by dashed boxes in (A) and (B) are magnified in (C-C″) and (D-D″) respectively. Asterisks in (A″) denote internal lumens bounded by Dlg-negative apical membrane of ept2/M(3) mutant cells. Asterisks in B-B″′ denote tissue lobes of ept2/M(3),stat92E06346/+ discs. Note the dominant effect of stat92E06346 on the appearance of Crb aggregates (A′ vs B′) and the lack of the ‘inverted’ tissue phenotype in ept2/M(3),stat92E06346/+ tumors (A″ vs B″). The stat92E06346 allele does not have a dominant effect on the mislocalization of Crb onto the Dlg-positive basolateral membrane domain of ept2 mutant cells (arrowhead in D-D″). (E) A control FRT80B/M(3) disc stained for Crb (blue), Dlg (red), and DNA (green). The peripodial cell layer (PP) and disc proper (DP) are indicated. Panels A″′ and B″′ are tracings of the apical (green) and basal (white) membranes of the tissues in A″ and B″ respectively.
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pone-0007083-g002: stat92E affects tissue architecture in ept tumors.Confocal images of ept2/M(3) (A-A″) and ept2/M(3),stat92E06346/+ (B-B″) eye discs stained for Dlg (red), Crb (blue) and DNA (green). The two main tissue lobes in panel B are separated by the esophagus (e), which remained embedded in the tissue mass during dissection. Areas outlined by dashed boxes in (A) and (B) are magnified in (C-C″) and (D-D″) respectively. Asterisks in (A″) denote internal lumens bounded by Dlg-negative apical membrane of ept2/M(3) mutant cells. Asterisks in B-B″′ denote tissue lobes of ept2/M(3),stat92E06346/+ discs. Note the dominant effect of stat92E06346 on the appearance of Crb aggregates (A′ vs B′) and the lack of the ‘inverted’ tissue phenotype in ept2/M(3),stat92E06346/+ tumors (A″ vs B″). The stat92E06346 allele does not have a dominant effect on the mislocalization of Crb onto the Dlg-positive basolateral membrane domain of ept2 mutant cells (arrowhead in D-D″). (E) A control FRT80B/M(3) disc stained for Crb (blue), Dlg (red), and DNA (green). The peripodial cell layer (PP) and disc proper (DP) are indicated. Panels A″′ and B″′ are tracings of the apical (green) and basal (white) membranes of the tissues in A″ and B″ respectively.

Mentions: Mutations in ept or vps25 have previously been shown to alter epithelial polarity and tissue architecture [4]–[6]. The ability of the stat92E06346 allele to restore a more normal morphology to ept mutant eye-antennal discs led us to examine the epithelial organization of these tissues. Co-staining for the basolateral membrane marker Discs large (Dlg) and the apical membrane determinant Crumbs (Crb) reveals that ept mutant eye-antennal discs are composed of compacted sheets of cells (Fig. 2A-A″) that form multilobular, ‘pouched’ structures with their apical surfaces oriented inward toward an internal lumen (examples denoted by yellow asterisks in Fig. 2A″). These structures are very much different from the normal monolayer organization of the peripodial and disc epithelium seen in control discs (Fig. 2E). Using the location of Dlg protein to trace the apical and basal surfaces of these ept/M(3) tissues highlights this phenotype (Fig. 2A′″). These architectural changes occur in parallel to a build-up of Crb protein in the cytoplasm of ept mutant cells (Fig. 2A′) due to a requirement for ESCRT complexes in vesicular trafficking of Crb [4], [5], [30]. Instead of concentrating at the apical surface of the tissue as in a control disc (Fig. 2E), Crb protein in ept/M(3) tumors is found more dispersed throughout the tissue (Fig. 2A′). A high magnification view of one of the ‘pouched’ structures (boxed in Fig. 2A) shows that ept is also required to prevent the spread of Crb from its normal location on the apical membrane onto the Dlg-positive basolateral domain (Fig. 2C-C″; arrowheads mark overlap of anti-Crb and anti-Dlg signals).


Genetic interactions between the Drosophila tumor suppressor gene ept and the stat92E transcription factor.

Gilbert MM, Beam CK, Robinson BS, Moberg KH - PLoS ONE (2009)

stat92E affects tissue architecture in ept tumors.Confocal images of ept2/M(3) (A-A″) and ept2/M(3),stat92E06346/+ (B-B″) eye discs stained for Dlg (red), Crb (blue) and DNA (green). The two main tissue lobes in panel B are separated by the esophagus (e), which remained embedded in the tissue mass during dissection. Areas outlined by dashed boxes in (A) and (B) are magnified in (C-C″) and (D-D″) respectively. Asterisks in (A″) denote internal lumens bounded by Dlg-negative apical membrane of ept2/M(3) mutant cells. Asterisks in B-B″′ denote tissue lobes of ept2/M(3),stat92E06346/+ discs. Note the dominant effect of stat92E06346 on the appearance of Crb aggregates (A′ vs B′) and the lack of the ‘inverted’ tissue phenotype in ept2/M(3),stat92E06346/+ tumors (A″ vs B″). The stat92E06346 allele does not have a dominant effect on the mislocalization of Crb onto the Dlg-positive basolateral membrane domain of ept2 mutant cells (arrowhead in D-D″). (E) A control FRT80B/M(3) disc stained for Crb (blue), Dlg (red), and DNA (green). The peripodial cell layer (PP) and disc proper (DP) are indicated. Panels A″′ and B″′ are tracings of the apical (green) and basal (white) membranes of the tissues in A″ and B″ respectively.
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Related In: Results  -  Collection

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pone-0007083-g002: stat92E affects tissue architecture in ept tumors.Confocal images of ept2/M(3) (A-A″) and ept2/M(3),stat92E06346/+ (B-B″) eye discs stained for Dlg (red), Crb (blue) and DNA (green). The two main tissue lobes in panel B are separated by the esophagus (e), which remained embedded in the tissue mass during dissection. Areas outlined by dashed boxes in (A) and (B) are magnified in (C-C″) and (D-D″) respectively. Asterisks in (A″) denote internal lumens bounded by Dlg-negative apical membrane of ept2/M(3) mutant cells. Asterisks in B-B″′ denote tissue lobes of ept2/M(3),stat92E06346/+ discs. Note the dominant effect of stat92E06346 on the appearance of Crb aggregates (A′ vs B′) and the lack of the ‘inverted’ tissue phenotype in ept2/M(3),stat92E06346/+ tumors (A″ vs B″). The stat92E06346 allele does not have a dominant effect on the mislocalization of Crb onto the Dlg-positive basolateral membrane domain of ept2 mutant cells (arrowhead in D-D″). (E) A control FRT80B/M(3) disc stained for Crb (blue), Dlg (red), and DNA (green). The peripodial cell layer (PP) and disc proper (DP) are indicated. Panels A″′ and B″′ are tracings of the apical (green) and basal (white) membranes of the tissues in A″ and B″ respectively.
Mentions: Mutations in ept or vps25 have previously been shown to alter epithelial polarity and tissue architecture [4]–[6]. The ability of the stat92E06346 allele to restore a more normal morphology to ept mutant eye-antennal discs led us to examine the epithelial organization of these tissues. Co-staining for the basolateral membrane marker Discs large (Dlg) and the apical membrane determinant Crumbs (Crb) reveals that ept mutant eye-antennal discs are composed of compacted sheets of cells (Fig. 2A-A″) that form multilobular, ‘pouched’ structures with their apical surfaces oriented inward toward an internal lumen (examples denoted by yellow asterisks in Fig. 2A″). These structures are very much different from the normal monolayer organization of the peripodial and disc epithelium seen in control discs (Fig. 2E). Using the location of Dlg protein to trace the apical and basal surfaces of these ept/M(3) tissues highlights this phenotype (Fig. 2A′″). These architectural changes occur in parallel to a build-up of Crb protein in the cytoplasm of ept mutant cells (Fig. 2A′) due to a requirement for ESCRT complexes in vesicular trafficking of Crb [4], [5], [30]. Instead of concentrating at the apical surface of the tissue as in a control disc (Fig. 2E), Crb protein in ept/M(3) tumors is found more dispersed throughout the tissue (Fig. 2A′). A high magnification view of one of the ‘pouched’ structures (boxed in Fig. 2A) shows that ept is also required to prevent the spread of Crb from its normal location on the apical membrane onto the Dlg-positive basolateral domain (Fig. 2C-C″; arrowheads mark overlap of anti-Crb and anti-Dlg signals).

Bottom Line: We find that this phenotype is very sensitive to the genetic dose of stat92E, the transcriptional effector of the Jak-Stat signaling pathway, and that this pathway undergoes strong activation in ept mutant cells.Genetic evidence indicates that stat92E contributes to cell cycle deregulation and excess cell size phenotypes that are observed among ept mutant cells.These findings identify ept as a cell-autonomous inhibitor of the Jak-Stat pathway and suggest that excess Jak-Stat signaling makes a significant contribution to proliferative and tissue architectural phenotypes that occur in ept mutant tissues.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Emory University School of Medicine, Atlanta, Georgia, United States of America.

ABSTRACT

Background: Tumor Susceptibility Gene-101 (TSG101) promotes the endocytic degradation of transmembrane proteins and is implicated as a mutational target in cancer, yet the effect of TSG101 loss on cell proliferation in vertebrates is uncertain. By contrast, Drosophila epithelial tissues lacking the TSG101 ortholog erupted (ept) develop as enlarged undifferentiated tumors, indicating that the gene can have anti-growth properties in a simple metazoan. A full understanding of pathways deregulated by loss of Drosophila ept will aid in understanding potential links between mammalian TSG101 and growth control.

Principal findings: We have taken a genetic approach to the identification of pathways required for excess growth of Drosophila eye-antennal imaginal discs lacking ept. We find that this phenotype is very sensitive to the genetic dose of stat92E, the transcriptional effector of the Jak-Stat signaling pathway, and that this pathway undergoes strong activation in ept mutant cells. Genetic evidence indicates that stat92E contributes to cell cycle deregulation and excess cell size phenotypes that are observed among ept mutant cells. In addition, autonomous Stat92E hyper-activation is associated with altered tissue architecture in ept tumors and an effect on expression of the apical polarity determinant crumbs.

Conclusions: These findings identify ept as a cell-autonomous inhibitor of the Jak-Stat pathway and suggest that excess Jak-Stat signaling makes a significant contribution to proliferative and tissue architectural phenotypes that occur in ept mutant tissues.

Show MeSH
Related in: MedlinePlus