Limits...
p53- and ERK7-dependent ribosome surveillance response regulates Drosophila insulin-like peptide secretion.

Hasygar K, Hietakangas V - PLoS Genet. (2014)

Bottom Line: A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15), which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation.Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation.Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences & Institute of Biotechnology, University of Helsinki, Helsinki, Finland.

ABSTRACT
Insulin-like signalling is a conserved mechanism that coordinates animal growth and metabolism with nutrient status. In Drosophila, insulin-producing median neurosecretory cells (IPCs) regulate larval growth by secreting insulin-like peptides (dILPs) in a diet-dependent manner. Previous studies have shown that nutrition affects dILP secretion through humoral signals derived from the fat body. Here we uncover a novel mechanism that operates cell autonomously in the IPCs to regulate dILP secretion. We observed that impairment of ribosome biogenesis specifically in the IPCs strongly inhibits dILP secretion, which consequently leads to reduced body size and a delay in larval development. This response is dependent on p53, a known surveillance factor for ribosome biogenesis. A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15), which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation. We show that ERK7 is sufficient and essential to inhibit dILP secretion upon impaired ribosome biogenesis, and it acts epistatically to p53. Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation. Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

Show MeSH

Related in: MedlinePlus

Kinome-wide screen identifies novel regulators of IPCs.(A) Overview of the kinome-wide RNAi screen in the IPCs. Each point indicates the mean body weight of RNAi expressing flies normalized to weight of flies from the same vial that do not express RNAi in the IPCs (i.e. relative weight). Dotted lines indicate +/−10% of the median body weight. (B) Kinome-wide RNAi screen identifies 12 kinases whose knockdown in the IPCs by two independent RNAi lines leads to significantly reduced body weight. Error bars represent standard deviation (N≥3, ≥10 flies/group). (C) TORC1 is essential for normal IPC function. Knockdown of Raptor, but not Rictor, in IPCs leads to reduction in body weight. Error bars represent standard deviation (N = 4, ≥10 flies/group). (D) Relative mRNA expression of dilp2, dilp3, and dilp5 upon knockdown of the kinase hits. Knockdown of Raptor, instead of TOR, was used to inhibit TORC1 function. Error bars represent standard deviation (N = 3, 10 brains/group). GAPDH was used as an internal reference. *p<0.05, **p<0.01, ***p<0.001 (Student's t-test).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4230838&req=5

pgen-1004764-g001: Kinome-wide screen identifies novel regulators of IPCs.(A) Overview of the kinome-wide RNAi screen in the IPCs. Each point indicates the mean body weight of RNAi expressing flies normalized to weight of flies from the same vial that do not express RNAi in the IPCs (i.e. relative weight). Dotted lines indicate +/−10% of the median body weight. (B) Kinome-wide RNAi screen identifies 12 kinases whose knockdown in the IPCs by two independent RNAi lines leads to significantly reduced body weight. Error bars represent standard deviation (N≥3, ≥10 flies/group). (C) TORC1 is essential for normal IPC function. Knockdown of Raptor, but not Rictor, in IPCs leads to reduction in body weight. Error bars represent standard deviation (N = 4, ≥10 flies/group). (D) Relative mRNA expression of dilp2, dilp3, and dilp5 upon knockdown of the kinase hits. Knockdown of Raptor, instead of TOR, was used to inhibit TORC1 function. Error bars represent standard deviation (N = 3, 10 brains/group). GAPDH was used as an internal reference. *p<0.05, **p<0.01, ***p<0.001 (Student's t-test).

Mentions: While the role of Drosophila insulin-like peptides (dILPs) in systemic growth control is well established, it remains poorly understood how the secretion of dILPs is regulated in response to changing dietary conditions. To get a better insight into this question, we explored the cell autonomous signalling mechanisms that regulate dILP secretion, by performing a kinome-wide screen with RNAi expressed specifically in the IPCs. Impaired IPC function compromises tissue growth [4] and therefore we used body weight of emerging adults as a screening readout (Figure 1A, Table S1). To normalize variation between vials due to growth conditions, we determined the relative weight wherein the mean weight of the flies expressing both dILP2-Gal4 and RNAi was divided by the mean weight of control flies (not expressing the dILP2-Gal4) from the same vial. In total, 231 kinases were screened. All primary hits displaying >10% difference to the median relative weight were further analyzed by an independent RNAi line. Altogether 12 protein kinases showed significant body weight reduction in two independent RNAi lines (Figure 1B). All identified kinases have mammalian orthologs (Table 1). In addition to these 12 kinases identified here, we have earlier shown that atypical PKC is essential for IPC function [25]. One of the kinase hits was TOR (target of rapamycin). TOR kinase is present in two functionally distinct complexes, TOR complex 1 and 2 (TORC1 and TORC2). To explore which of the TOR complexes have a regulatory role in the IPCs, we depleted Raptor and Rictor, essential components of TORC1 and TORC2, respectively. Knockdown of Raptor in the IPCs significantly reduced total body weight, while depletion of Rictor had no significant impact (Figure 1C), showing that TORC1 regulates growth through the IPCs. However, as RNAi does not completely silence gene expression, we cannot rule out the involvement of TORC2.


p53- and ERK7-dependent ribosome surveillance response regulates Drosophila insulin-like peptide secretion.

Hasygar K, Hietakangas V - PLoS Genet. (2014)

Kinome-wide screen identifies novel regulators of IPCs.(A) Overview of the kinome-wide RNAi screen in the IPCs. Each point indicates the mean body weight of RNAi expressing flies normalized to weight of flies from the same vial that do not express RNAi in the IPCs (i.e. relative weight). Dotted lines indicate +/−10% of the median body weight. (B) Kinome-wide RNAi screen identifies 12 kinases whose knockdown in the IPCs by two independent RNAi lines leads to significantly reduced body weight. Error bars represent standard deviation (N≥3, ≥10 flies/group). (C) TORC1 is essential for normal IPC function. Knockdown of Raptor, but not Rictor, in IPCs leads to reduction in body weight. Error bars represent standard deviation (N = 4, ≥10 flies/group). (D) Relative mRNA expression of dilp2, dilp3, and dilp5 upon knockdown of the kinase hits. Knockdown of Raptor, instead of TOR, was used to inhibit TORC1 function. Error bars represent standard deviation (N = 3, 10 brains/group). GAPDH was used as an internal reference. *p<0.05, **p<0.01, ***p<0.001 (Student's t-test).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230838&req=5

pgen-1004764-g001: Kinome-wide screen identifies novel regulators of IPCs.(A) Overview of the kinome-wide RNAi screen in the IPCs. Each point indicates the mean body weight of RNAi expressing flies normalized to weight of flies from the same vial that do not express RNAi in the IPCs (i.e. relative weight). Dotted lines indicate +/−10% of the median body weight. (B) Kinome-wide RNAi screen identifies 12 kinases whose knockdown in the IPCs by two independent RNAi lines leads to significantly reduced body weight. Error bars represent standard deviation (N≥3, ≥10 flies/group). (C) TORC1 is essential for normal IPC function. Knockdown of Raptor, but not Rictor, in IPCs leads to reduction in body weight. Error bars represent standard deviation (N = 4, ≥10 flies/group). (D) Relative mRNA expression of dilp2, dilp3, and dilp5 upon knockdown of the kinase hits. Knockdown of Raptor, instead of TOR, was used to inhibit TORC1 function. Error bars represent standard deviation (N = 3, 10 brains/group). GAPDH was used as an internal reference. *p<0.05, **p<0.01, ***p<0.001 (Student's t-test).
Mentions: While the role of Drosophila insulin-like peptides (dILPs) in systemic growth control is well established, it remains poorly understood how the secretion of dILPs is regulated in response to changing dietary conditions. To get a better insight into this question, we explored the cell autonomous signalling mechanisms that regulate dILP secretion, by performing a kinome-wide screen with RNAi expressed specifically in the IPCs. Impaired IPC function compromises tissue growth [4] and therefore we used body weight of emerging adults as a screening readout (Figure 1A, Table S1). To normalize variation between vials due to growth conditions, we determined the relative weight wherein the mean weight of the flies expressing both dILP2-Gal4 and RNAi was divided by the mean weight of control flies (not expressing the dILP2-Gal4) from the same vial. In total, 231 kinases were screened. All primary hits displaying >10% difference to the median relative weight were further analyzed by an independent RNAi line. Altogether 12 protein kinases showed significant body weight reduction in two independent RNAi lines (Figure 1B). All identified kinases have mammalian orthologs (Table 1). In addition to these 12 kinases identified here, we have earlier shown that atypical PKC is essential for IPC function [25]. One of the kinase hits was TOR (target of rapamycin). TOR kinase is present in two functionally distinct complexes, TOR complex 1 and 2 (TORC1 and TORC2). To explore which of the TOR complexes have a regulatory role in the IPCs, we depleted Raptor and Rictor, essential components of TORC1 and TORC2, respectively. Knockdown of Raptor in the IPCs significantly reduced total body weight, while depletion of Rictor had no significant impact (Figure 1C), showing that TORC1 regulates growth through the IPCs. However, as RNAi does not completely silence gene expression, we cannot rule out the involvement of TORC2.

Bottom Line: A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15), which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation.Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation.Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosciences & Institute of Biotechnology, University of Helsinki, Helsinki, Finland.

ABSTRACT
Insulin-like signalling is a conserved mechanism that coordinates animal growth and metabolism with nutrient status. In Drosophila, insulin-producing median neurosecretory cells (IPCs) regulate larval growth by secreting insulin-like peptides (dILPs) in a diet-dependent manner. Previous studies have shown that nutrition affects dILP secretion through humoral signals derived from the fat body. Here we uncover a novel mechanism that operates cell autonomously in the IPCs to regulate dILP secretion. We observed that impairment of ribosome biogenesis specifically in the IPCs strongly inhibits dILP secretion, which consequently leads to reduced body size and a delay in larval development. This response is dependent on p53, a known surveillance factor for ribosome biogenesis. A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15), which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation. We show that ERK7 is sufficient and essential to inhibit dILP secretion upon impaired ribosome biogenesis, and it acts epistatically to p53. Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation. Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

Show MeSH
Related in: MedlinePlus