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Scaling the Drosophila Wing: TOR-Dependent Target Gene Access by the Hippo Pathway Transducer Yorkie.

Parker J, Struhl G - PLoS Biol. (2015)

Bottom Line: Here, we show that the TOR pathway regulates Yki by a separate and novel mechanism in the Drosophila wing.Instead of controlling Yki nuclear access, TOR signaling governs Yki action after it reaches the nucleus by allowing it to gain access to its target genes.When TOR activity is inhibited, Yki accumulates in the nucleus but is sequestered from its normal growth-promoting target genes--a phenomenon we term "nuclear seclusion." Hence, we posit that in addition to its well-known role in stimulating cellular metabolism in response to nutrients, TOR also promotes wing growth by liberating Yki from nuclear seclusion, a parallel pathway that we propose contributes to the scaling of wing size with nutrient availability.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics and Development, Columbia University, New York, New York, United States of America; Division of Biology, Imperial College London, London, United Kingdom.

ABSTRACT
Organ growth is controlled by patterning signals that operate locally (e.g., Wingless/Ints [Wnts], Bone Morphogenetic Proteins [BMPs], and Hedgehogs [Hhs]) and scaled by nutrient-dependent signals that act systemically (e.g., Insulin-like peptides [ILPs] transduced by the Target of Rapamycin [TOR] pathway). How cells integrate these distinct inputs to generate organs of the appropriate size and shape is largely unknown. The transcriptional coactivator Yorkie (Yki, a YES-Associated Protein, or YAP) acts downstream of patterning morphogens and other tissue-intrinsic signals to promote organ growth. Yki activity is regulated primarily by the Warts/Hippo (Wts/Hpo) tumour suppressor pathway, which impedes nuclear access of Yki by a cytoplasmic tethering mechanism. Here, we show that the TOR pathway regulates Yki by a separate and novel mechanism in the Drosophila wing. Instead of controlling Yki nuclear access, TOR signaling governs Yki action after it reaches the nucleus by allowing it to gain access to its target genes. When TOR activity is inhibited, Yki accumulates in the nucleus but is sequestered from its normal growth-promoting target genes--a phenomenon we term "nuclear seclusion." Hence, we posit that in addition to its well-known role in stimulating cellular metabolism in response to nutrients, TOR also promotes wing growth by liberating Yki from nuclear seclusion, a parallel pathway that we propose contributes to the scaling of wing size with nutrient availability.

No MeSH data available.


Related in: MedlinePlus

Effects of TOR inhibition on wing growth and Yki-driven cell proliferation.A–C: Adult Drosophila wings. (A) A wing from a well-fed wild type fly. (B) Scaled-down wing produced by raising larvae on nutrient-poor food. (C) Scaled-down wing caused by blocking TOR signaling specifically in wing cells with nubbin.GAL4 UAS.TorTED. (D–G) Wing discs from late third instar larvae-bearing clones of mutant tissue outlined with dashed lines, and marked negatively (“black”) by absence of the GFP marker (green): (D) wild type (control) (E) TorΔP, (F) exe1, (G) exe1TorΔP. Mutant clones were induced at the end of the first instar, 48±2 hr after egg laying and are associated with sibling “twin-spot” clones marked by two copies of the GFP marker (bright green) that serve as an internal control for the growth of w.t. tissue. Numbers denote mean clone size ratio compared to wt, and asterisks denote significances from t tests (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, n. s. = not significant). In (G), the bottom italicised value is a comparison with the TorΔP genotype. Number of clones measured (n) = 32 (wt), 38 (ex), 51 (Tor), 36 (ex Tor). (H–K) Clones of the same genotypes as in (D–G) that coexpress p35 with GFP-NLS (generated using the MARCM technique [36]. Clones are positively labelled by GFP-NLS, and nuclei are counterstained with Hoechst (blue). (n) = 92 (wt+p35), 90 (ex+p35), 97 (Tor+p35), 79 (Ex Tor+p35). Numbers signify mean clone size ratio compared to wt+p35, and bottom italicised value in (K) with Tor+p35.
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pbio.1002274.g001: Effects of TOR inhibition on wing growth and Yki-driven cell proliferation.A–C: Adult Drosophila wings. (A) A wing from a well-fed wild type fly. (B) Scaled-down wing produced by raising larvae on nutrient-poor food. (C) Scaled-down wing caused by blocking TOR signaling specifically in wing cells with nubbin.GAL4 UAS.TorTED. (D–G) Wing discs from late third instar larvae-bearing clones of mutant tissue outlined with dashed lines, and marked negatively (“black”) by absence of the GFP marker (green): (D) wild type (control) (E) TorΔP, (F) exe1, (G) exe1TorΔP. Mutant clones were induced at the end of the first instar, 48±2 hr after egg laying and are associated with sibling “twin-spot” clones marked by two copies of the GFP marker (bright green) that serve as an internal control for the growth of w.t. tissue. Numbers denote mean clone size ratio compared to wt, and asterisks denote significances from t tests (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, n. s. = not significant). In (G), the bottom italicised value is a comparison with the TorΔP genotype. Number of clones measured (n) = 32 (wt), 38 (ex), 51 (Tor), 36 (ex Tor). (H–K) Clones of the same genotypes as in (D–G) that coexpress p35 with GFP-NLS (generated using the MARCM technique [36]. Clones are positively labelled by GFP-NLS, and nuclei are counterstained with Hoechst (blue). (n) = 92 (wt+p35), 90 (ex+p35), 97 (Tor+p35), 79 (Ex Tor+p35). Numbers signify mean clone size ratio compared to wt+p35, and bottom italicised value in (K) with Tor+p35.

Mentions: The Drosophila wing is a classical paradigm of organ growth [2,3]. Here, as in other animals, nutrients influence growth via Target of Rapamycin (TOR) signaling [5,10]. During larval life, this pathway is activated in wing cells by haemolymph signals produced in response to feeding, including Insulin-like peptides (ILPs) that act via the Insulin Receptor (InR)/PI3-Kinase/Akt pathway, as well as sugars and amino acids. These inputs converge to regulate TOR—an intracellular kinase with diverse roles in metabolism [5,10]. Starvation reduces TOR activity and scales wing size (and entire body size) downwards (Fig 1A and 1B), an effect mimicked by genetically inhibiting TOR (Fig 1C). Yet, wing growth is simultaneously governed by intrinsic signaling systems (e.g., Wnt, BMP, and Hh morphogens) that control wing size, shape, and pattern [2,3,6,8]. Many of these organ-intrinsic systems exert their effects at least in part via regulation of the Warts (Wts)/Hippo (Hpo) pathway [11–13]—a network of proteins that inhibit a growth-promoting transcriptional coactivator, Yorkie (Yki; orthologous to vertebrate YES-Associated Protein [YAP]) [14]. Hpo and Wts are kinases that act in sequence, Hpo phosphorylating Wts and Wts phosphorylating Yki to sequester Yki cytoplasmically. Inhibition of either kinase promotes growth by allowing Yki to evade cytosolic sequestration and gain access to the nucleus [15–17]. Nuclear Yki binds transcription factors including Scalloped (Sd, a TEAD [transcriptional enhancer activator domain] protein) [18,19] to up-regulate expression of genes that promote cell growth and proliferation. Morphogens [20,21], the protocadherins Fat and Dachsous [22–25], the Crumbs/Lgl epithelial polarity proteins [26–29], and mechanical strain [30] all modulate either or both Wts and Hpo, exerting effects on wing growth via Yki. But for the wing to scale, Yki activity, or the growth that Yki stimulates, must be contingent on TOR activity. Despite previous attempts to assess the links between Wts/Hpo and InR/TOR signaling [15,31–35], the logic by which cells in growing organs integrate these inputs to achieve organ scaling remains unclear.


Scaling the Drosophila Wing: TOR-Dependent Target Gene Access by the Hippo Pathway Transducer Yorkie.

Parker J, Struhl G - PLoS Biol. (2015)

Effects of TOR inhibition on wing growth and Yki-driven cell proliferation.A–C: Adult Drosophila wings. (A) A wing from a well-fed wild type fly. (B) Scaled-down wing produced by raising larvae on nutrient-poor food. (C) Scaled-down wing caused by blocking TOR signaling specifically in wing cells with nubbin.GAL4 UAS.TorTED. (D–G) Wing discs from late third instar larvae-bearing clones of mutant tissue outlined with dashed lines, and marked negatively (“black”) by absence of the GFP marker (green): (D) wild type (control) (E) TorΔP, (F) exe1, (G) exe1TorΔP. Mutant clones were induced at the end of the first instar, 48±2 hr after egg laying and are associated with sibling “twin-spot” clones marked by two copies of the GFP marker (bright green) that serve as an internal control for the growth of w.t. tissue. Numbers denote mean clone size ratio compared to wt, and asterisks denote significances from t tests (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, n. s. = not significant). In (G), the bottom italicised value is a comparison with the TorΔP genotype. Number of clones measured (n) = 32 (wt), 38 (ex), 51 (Tor), 36 (ex Tor). (H–K) Clones of the same genotypes as in (D–G) that coexpress p35 with GFP-NLS (generated using the MARCM technique [36]. Clones are positively labelled by GFP-NLS, and nuclei are counterstained with Hoechst (blue). (n) = 92 (wt+p35), 90 (ex+p35), 97 (Tor+p35), 79 (Ex Tor+p35). Numbers signify mean clone size ratio compared to wt+p35, and bottom italicised value in (K) with Tor+p35.
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pbio.1002274.g001: Effects of TOR inhibition on wing growth and Yki-driven cell proliferation.A–C: Adult Drosophila wings. (A) A wing from a well-fed wild type fly. (B) Scaled-down wing produced by raising larvae on nutrient-poor food. (C) Scaled-down wing caused by blocking TOR signaling specifically in wing cells with nubbin.GAL4 UAS.TorTED. (D–G) Wing discs from late third instar larvae-bearing clones of mutant tissue outlined with dashed lines, and marked negatively (“black”) by absence of the GFP marker (green): (D) wild type (control) (E) TorΔP, (F) exe1, (G) exe1TorΔP. Mutant clones were induced at the end of the first instar, 48±2 hr after egg laying and are associated with sibling “twin-spot” clones marked by two copies of the GFP marker (bright green) that serve as an internal control for the growth of w.t. tissue. Numbers denote mean clone size ratio compared to wt, and asterisks denote significances from t tests (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, n. s. = not significant). In (G), the bottom italicised value is a comparison with the TorΔP genotype. Number of clones measured (n) = 32 (wt), 38 (ex), 51 (Tor), 36 (ex Tor). (H–K) Clones of the same genotypes as in (D–G) that coexpress p35 with GFP-NLS (generated using the MARCM technique [36]. Clones are positively labelled by GFP-NLS, and nuclei are counterstained with Hoechst (blue). (n) = 92 (wt+p35), 90 (ex+p35), 97 (Tor+p35), 79 (Ex Tor+p35). Numbers signify mean clone size ratio compared to wt+p35, and bottom italicised value in (K) with Tor+p35.
Mentions: The Drosophila wing is a classical paradigm of organ growth [2,3]. Here, as in other animals, nutrients influence growth via Target of Rapamycin (TOR) signaling [5,10]. During larval life, this pathway is activated in wing cells by haemolymph signals produced in response to feeding, including Insulin-like peptides (ILPs) that act via the Insulin Receptor (InR)/PI3-Kinase/Akt pathway, as well as sugars and amino acids. These inputs converge to regulate TOR—an intracellular kinase with diverse roles in metabolism [5,10]. Starvation reduces TOR activity and scales wing size (and entire body size) downwards (Fig 1A and 1B), an effect mimicked by genetically inhibiting TOR (Fig 1C). Yet, wing growth is simultaneously governed by intrinsic signaling systems (e.g., Wnt, BMP, and Hh morphogens) that control wing size, shape, and pattern [2,3,6,8]. Many of these organ-intrinsic systems exert their effects at least in part via regulation of the Warts (Wts)/Hippo (Hpo) pathway [11–13]—a network of proteins that inhibit a growth-promoting transcriptional coactivator, Yorkie (Yki; orthologous to vertebrate YES-Associated Protein [YAP]) [14]. Hpo and Wts are kinases that act in sequence, Hpo phosphorylating Wts and Wts phosphorylating Yki to sequester Yki cytoplasmically. Inhibition of either kinase promotes growth by allowing Yki to evade cytosolic sequestration and gain access to the nucleus [15–17]. Nuclear Yki binds transcription factors including Scalloped (Sd, a TEAD [transcriptional enhancer activator domain] protein) [18,19] to up-regulate expression of genes that promote cell growth and proliferation. Morphogens [20,21], the protocadherins Fat and Dachsous [22–25], the Crumbs/Lgl epithelial polarity proteins [26–29], and mechanical strain [30] all modulate either or both Wts and Hpo, exerting effects on wing growth via Yki. But for the wing to scale, Yki activity, or the growth that Yki stimulates, must be contingent on TOR activity. Despite previous attempts to assess the links between Wts/Hpo and InR/TOR signaling [15,31–35], the logic by which cells in growing organs integrate these inputs to achieve organ scaling remains unclear.

Bottom Line: Here, we show that the TOR pathway regulates Yki by a separate and novel mechanism in the Drosophila wing.Instead of controlling Yki nuclear access, TOR signaling governs Yki action after it reaches the nucleus by allowing it to gain access to its target genes.When TOR activity is inhibited, Yki accumulates in the nucleus but is sequestered from its normal growth-promoting target genes--a phenomenon we term "nuclear seclusion." Hence, we posit that in addition to its well-known role in stimulating cellular metabolism in response to nutrients, TOR also promotes wing growth by liberating Yki from nuclear seclusion, a parallel pathway that we propose contributes to the scaling of wing size with nutrient availability.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics and Development, Columbia University, New York, New York, United States of America; Division of Biology, Imperial College London, London, United Kingdom.

ABSTRACT
Organ growth is controlled by patterning signals that operate locally (e.g., Wingless/Ints [Wnts], Bone Morphogenetic Proteins [BMPs], and Hedgehogs [Hhs]) and scaled by nutrient-dependent signals that act systemically (e.g., Insulin-like peptides [ILPs] transduced by the Target of Rapamycin [TOR] pathway). How cells integrate these distinct inputs to generate organs of the appropriate size and shape is largely unknown. The transcriptional coactivator Yorkie (Yki, a YES-Associated Protein, or YAP) acts downstream of patterning morphogens and other tissue-intrinsic signals to promote organ growth. Yki activity is regulated primarily by the Warts/Hippo (Wts/Hpo) tumour suppressor pathway, which impedes nuclear access of Yki by a cytoplasmic tethering mechanism. Here, we show that the TOR pathway regulates Yki by a separate and novel mechanism in the Drosophila wing. Instead of controlling Yki nuclear access, TOR signaling governs Yki action after it reaches the nucleus by allowing it to gain access to its target genes. When TOR activity is inhibited, Yki accumulates in the nucleus but is sequestered from its normal growth-promoting target genes--a phenomenon we term "nuclear seclusion." Hence, we posit that in addition to its well-known role in stimulating cellular metabolism in response to nutrients, TOR also promotes wing growth by liberating Yki from nuclear seclusion, a parallel pathway that we propose contributes to the scaling of wing size with nutrient availability.

No MeSH data available.


Related in: MedlinePlus