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TIF-IA-dependent regulation of ribosome synthesis in drosophila muscle is required to maintain systemic insulin signaling and larval growth.

Ghosh A, Rideout EJ, Grewal SS - PLoS Genet. (2014)

Bottom Line: When we mimic this decrease in muscle ribosome synthesis using RNAi-mediated knockdown of TIF-IA, we observe delayed larval development and reduced body growth.This reduction in growth is caused by lowered systemic insulin signaling via two endocrine responses: reduced expression of Drosophila insulin-like peptides (dILPs) from the brain and increased expression of Imp-L2-a secreted factor that binds and inhibits dILP activity-from muscle.Finally, we show that activation of TOR specifically in muscle can increase overall body size and this effect requires TIF-IA function.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, and Clark H. Smith Brain Tumour Centre, Southern Alberta Cancer Research Institute, University of Calgary, Health Research Innovation Center, Calgary, Alberta, Canada.

ABSTRACT
The conserved TOR kinase signaling network links nutrient availability to cell, tissue and body growth in animals. One important growth-regulatory target of TOR signaling is ribosome biogenesis. Studies in yeast and mammalian cell culture have described how TOR controls rRNA synthesis-a limiting step in ribosome biogenesis-via the RNA Polymerase I transcription factor TIF-IA. However, the contribution of TOR-dependent ribosome synthesis to tissue and body growth in animals is less clear. Here we show in Drosophila larvae that ribosome synthesis in muscle is required non-autonomously to maintain normal body growth and development. We find that amino acid starvation and TOR inhibition lead to reduced levels of TIF-IA, and decreased rRNA synthesis in larval muscle. When we mimic this decrease in muscle ribosome synthesis using RNAi-mediated knockdown of TIF-IA, we observe delayed larval development and reduced body growth. This reduction in growth is caused by lowered systemic insulin signaling via two endocrine responses: reduced expression of Drosophila insulin-like peptides (dILPs) from the brain and increased expression of Imp-L2-a secreted factor that binds and inhibits dILP activity-from muscle. We also observed that maintaining TIF-IA levels in muscle could partially reverse the starvation-mediated suppression of systemic insulin signaling. Finally, we show that activation of TOR specifically in muscle can increase overall body size and this effect requires TIF-IA function. These data suggest that muscle ribosome synthesis functions as a nutrient-dependent checkpoint for overall body growth: in nutrient rich conditions, TOR is required to maintain levels of TIF-IA and ribosome synthesis to promote high levels of systemic insulin, but under conditions of starvation stress, reduced muscle ribosome synthesis triggers an endocrine response that limits systemic insulin signaling to restrict growth and maintain homeostasis.

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Related in: MedlinePlus

TOR activity in muscle is required and sufficient to promote body growth and TIF-IA inhibition in muscle blocks Rheb induced body growth.(A) Pupal volume of dMef2>+ and dMef2>TORTED pupae, n = 29, n - number of pupae per genotype, (*P = 2.47×10−7, Student's t-test). (B) Pupal volume of dMef2>+ and dMef2>Tsc1,Tsc2 pupae, n>55, n - number of pupae per genotype, (*P = 1.08×10−51, Student's t-test). (C) Pupal volume of dMef2>+ and dMef2>slifAnti pupae, n>80, n - number of pupae per genotype, (*P = 5.14×10−10, Student's t-test). (D) Pupal volume of dMef2>+ and dMef2>Rheb pupae, n>38, n - number of pupae per genotype, (*P = 0.003, Student's t-test). (E-F) Representative figures of larvae and pupae of indicated genotypes, scale bar-500 µm. (G) dMef2>Rheb animals showed increased pupal volume (White bar, *P<0.0001, One-way ANOVA and Tukey's post test) compared to dMef2>+ control. Muscle specific inhibition of TIF-IA (dMef2>TIF-IA IR) reduced pupal volume, with respect to dMef2>+ control (Grey bar, •P = 0.0003, One-way ANOVA and Tukey's post test). TIF-IA knockdown in muscle abrogated the Rheb-induced increase in pupal volume (Blue bar, ⧫P<0.0001, One-way ANOVA and Tukey's post test), n - number of pupae per genotype. All error bars indicate SEM.
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pgen-1004750-g004: TOR activity in muscle is required and sufficient to promote body growth and TIF-IA inhibition in muscle blocks Rheb induced body growth.(A) Pupal volume of dMef2>+ and dMef2>TORTED pupae, n = 29, n - number of pupae per genotype, (*P = 2.47×10−7, Student's t-test). (B) Pupal volume of dMef2>+ and dMef2>Tsc1,Tsc2 pupae, n>55, n - number of pupae per genotype, (*P = 1.08×10−51, Student's t-test). (C) Pupal volume of dMef2>+ and dMef2>slifAnti pupae, n>80, n - number of pupae per genotype, (*P = 5.14×10−10, Student's t-test). (D) Pupal volume of dMef2>+ and dMef2>Rheb pupae, n>38, n - number of pupae per genotype, (*P = 0.003, Student's t-test). (E-F) Representative figures of larvae and pupae of indicated genotypes, scale bar-500 µm. (G) dMef2>Rheb animals showed increased pupal volume (White bar, *P<0.0001, One-way ANOVA and Tukey's post test) compared to dMef2>+ control. Muscle specific inhibition of TIF-IA (dMef2>TIF-IA IR) reduced pupal volume, with respect to dMef2>+ control (Grey bar, •P = 0.0003, One-way ANOVA and Tukey's post test). TIF-IA knockdown in muscle abrogated the Rheb-induced increase in pupal volume (Blue bar, ⧫P<0.0001, One-way ANOVA and Tukey's post test), n - number of pupae per genotype. All error bars indicate SEM.

Mentions: TOR activity in muscle is required for normal larval growth and development [28]. We confirmed this finding by inhibiting TOR in the muscle by two different methods, expression of a dominant negative form of TOR in muscle (dMef2>TORTED) [32] and co-overexpression of UAS-Tsc1 and UAS-Tsc2 - negative regulators of TORC1 - in muscle (dMef2>Tsc1,Tsc2). We measured pupal volume, as an indicator of final body size. Our data showed that in both cases, inhibition of TOR in larval muscle reduced pupal volume (Figure 4A, B). Amino acid availability is an important activator of TOR kinase signaling, and we also found that knockdown of the amino acid transporter slimfast (using a UAS-slifAnti antisense [25]), in the larval muscle led to a significant reduction in pupal volume (Figure 4C). Finally, we also examined whether over-activation of TOR in muscle was sufficient to drive systemic growth. We found that overexpression of Ras homolog enriched in brain (Rheb), an upstream activator specifically of TORC1, in muscle (dMef2>Rheb) was sufficient to increase pupal volume compared to control (dMef2>+) pupae (Figure 4D). Together, these findings suggest that TOR activity in muscle is both necessary and sufficient to control overall systemic growth.


TIF-IA-dependent regulation of ribosome synthesis in drosophila muscle is required to maintain systemic insulin signaling and larval growth.

Ghosh A, Rideout EJ, Grewal SS - PLoS Genet. (2014)

TOR activity in muscle is required and sufficient to promote body growth and TIF-IA inhibition in muscle blocks Rheb induced body growth.(A) Pupal volume of dMef2>+ and dMef2>TORTED pupae, n = 29, n - number of pupae per genotype, (*P = 2.47×10−7, Student's t-test). (B) Pupal volume of dMef2>+ and dMef2>Tsc1,Tsc2 pupae, n>55, n - number of pupae per genotype, (*P = 1.08×10−51, Student's t-test). (C) Pupal volume of dMef2>+ and dMef2>slifAnti pupae, n>80, n - number of pupae per genotype, (*P = 5.14×10−10, Student's t-test). (D) Pupal volume of dMef2>+ and dMef2>Rheb pupae, n>38, n - number of pupae per genotype, (*P = 0.003, Student's t-test). (E-F) Representative figures of larvae and pupae of indicated genotypes, scale bar-500 µm. (G) dMef2>Rheb animals showed increased pupal volume (White bar, *P<0.0001, One-way ANOVA and Tukey's post test) compared to dMef2>+ control. Muscle specific inhibition of TIF-IA (dMef2>TIF-IA IR) reduced pupal volume, with respect to dMef2>+ control (Grey bar, •P = 0.0003, One-way ANOVA and Tukey's post test). TIF-IA knockdown in muscle abrogated the Rheb-induced increase in pupal volume (Blue bar, ⧫P<0.0001, One-way ANOVA and Tukey's post test), n - number of pupae per genotype. All error bars indicate SEM.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4214618&req=5

pgen-1004750-g004: TOR activity in muscle is required and sufficient to promote body growth and TIF-IA inhibition in muscle blocks Rheb induced body growth.(A) Pupal volume of dMef2>+ and dMef2>TORTED pupae, n = 29, n - number of pupae per genotype, (*P = 2.47×10−7, Student's t-test). (B) Pupal volume of dMef2>+ and dMef2>Tsc1,Tsc2 pupae, n>55, n - number of pupae per genotype, (*P = 1.08×10−51, Student's t-test). (C) Pupal volume of dMef2>+ and dMef2>slifAnti pupae, n>80, n - number of pupae per genotype, (*P = 5.14×10−10, Student's t-test). (D) Pupal volume of dMef2>+ and dMef2>Rheb pupae, n>38, n - number of pupae per genotype, (*P = 0.003, Student's t-test). (E-F) Representative figures of larvae and pupae of indicated genotypes, scale bar-500 µm. (G) dMef2>Rheb animals showed increased pupal volume (White bar, *P<0.0001, One-way ANOVA and Tukey's post test) compared to dMef2>+ control. Muscle specific inhibition of TIF-IA (dMef2>TIF-IA IR) reduced pupal volume, with respect to dMef2>+ control (Grey bar, •P = 0.0003, One-way ANOVA and Tukey's post test). TIF-IA knockdown in muscle abrogated the Rheb-induced increase in pupal volume (Blue bar, ⧫P<0.0001, One-way ANOVA and Tukey's post test), n - number of pupae per genotype. All error bars indicate SEM.
Mentions: TOR activity in muscle is required for normal larval growth and development [28]. We confirmed this finding by inhibiting TOR in the muscle by two different methods, expression of a dominant negative form of TOR in muscle (dMef2>TORTED) [32] and co-overexpression of UAS-Tsc1 and UAS-Tsc2 - negative regulators of TORC1 - in muscle (dMef2>Tsc1,Tsc2). We measured pupal volume, as an indicator of final body size. Our data showed that in both cases, inhibition of TOR in larval muscle reduced pupal volume (Figure 4A, B). Amino acid availability is an important activator of TOR kinase signaling, and we also found that knockdown of the amino acid transporter slimfast (using a UAS-slifAnti antisense [25]), in the larval muscle led to a significant reduction in pupal volume (Figure 4C). Finally, we also examined whether over-activation of TOR in muscle was sufficient to drive systemic growth. We found that overexpression of Ras homolog enriched in brain (Rheb), an upstream activator specifically of TORC1, in muscle (dMef2>Rheb) was sufficient to increase pupal volume compared to control (dMef2>+) pupae (Figure 4D). Together, these findings suggest that TOR activity in muscle is both necessary and sufficient to control overall systemic growth.

Bottom Line: When we mimic this decrease in muscle ribosome synthesis using RNAi-mediated knockdown of TIF-IA, we observe delayed larval development and reduced body growth.This reduction in growth is caused by lowered systemic insulin signaling via two endocrine responses: reduced expression of Drosophila insulin-like peptides (dILPs) from the brain and increased expression of Imp-L2-a secreted factor that binds and inhibits dILP activity-from muscle.Finally, we show that activation of TOR specifically in muscle can increase overall body size and this effect requires TIF-IA function.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, and Clark H. Smith Brain Tumour Centre, Southern Alberta Cancer Research Institute, University of Calgary, Health Research Innovation Center, Calgary, Alberta, Canada.

ABSTRACT
The conserved TOR kinase signaling network links nutrient availability to cell, tissue and body growth in animals. One important growth-regulatory target of TOR signaling is ribosome biogenesis. Studies in yeast and mammalian cell culture have described how TOR controls rRNA synthesis-a limiting step in ribosome biogenesis-via the RNA Polymerase I transcription factor TIF-IA. However, the contribution of TOR-dependent ribosome synthesis to tissue and body growth in animals is less clear. Here we show in Drosophila larvae that ribosome synthesis in muscle is required non-autonomously to maintain normal body growth and development. We find that amino acid starvation and TOR inhibition lead to reduced levels of TIF-IA, and decreased rRNA synthesis in larval muscle. When we mimic this decrease in muscle ribosome synthesis using RNAi-mediated knockdown of TIF-IA, we observe delayed larval development and reduced body growth. This reduction in growth is caused by lowered systemic insulin signaling via two endocrine responses: reduced expression of Drosophila insulin-like peptides (dILPs) from the brain and increased expression of Imp-L2-a secreted factor that binds and inhibits dILP activity-from muscle. We also observed that maintaining TIF-IA levels in muscle could partially reverse the starvation-mediated suppression of systemic insulin signaling. Finally, we show that activation of TOR specifically in muscle can increase overall body size and this effect requires TIF-IA function. These data suggest that muscle ribosome synthesis functions as a nutrient-dependent checkpoint for overall body growth: in nutrient rich conditions, TOR is required to maintain levels of TIF-IA and ribosome synthesis to promote high levels of systemic insulin, but under conditions of starvation stress, reduced muscle ribosome synthesis triggers an endocrine response that limits systemic insulin signaling to restrict growth and maintain homeostasis.

Show MeSH
Related in: MedlinePlus