An Integrative Analysis of the InR/PI3K/Akt Network Identifies the Dynamic Response to Insulin Signaling

Abstract

Summary Insulin regulates an essential conserved signaling pathway affecting growth, proliferation and metabolism. To expand our understanding of the insulin pathway, we combine biochemical, genetic and computational approaches to build a comprehensive Drosophila InR/PI3K/Akt network. First, we map the dynamic protein-protein interaction network surrounding the insulin core pathway using bait-prey interactions connecting 566 proteins. Combining RNA interference screening and phospho-specific antibodies, we find that 47% of interacting proteins affect pathway activity, and using quantitative phosphoproteomics, we demonstrate that ~10% of interacting proteins are regulated by insulin stimulation at the level of phosphorylation. Next, we integrate these orthogonal datasets to characterize the structure and dynamics of the insulin network at the level of protein complexes, and validate our method by identifying regulatory roles for the Protein Phosphatase 2A (PP2A) and Reptin-Pontin chromatin-remodeling complexes as negative and positive regulators of ribosome biogenesis, respectively. Altogether, our study represents a comprehensive resource for study of the evolutionary conserved insulin network.

Distinct genomic signals of lifespan and life history evolution in response to postponed reproduction and larval diet inDrosophila

“…) and the gene Psa , known to interact with the major IIS/TOR core component Akt1 (Vinayagam et al. ). Similarly, among the loci that differentiate the LE populations from the other EE populations we identified Tomosyn , a regulator of insulin secretion with a confirmed role in aging in C. elegans (Ch'ng et al.…”

“…discussion in Remolina et al 2012;Fabian et al 2018;Flatt and Partridge 2018), we only found a few loci in this signaling network among our candidates. For instance, among the loci significantly differentiated between the E and P populations, we identified the TOR signaling gene happyhour (Bryk et al 2010;Khan et al 2012) and the gene Psa, known to interact with the major IIS/TOR core component Akt1 (Vinayagam et al 2016). Similarly, among the loci that differentiate the LE populations from the other EE populations we identified Tomosyn, a regulator of insulin secretion with a confirmed role in aging in C. elegans (Ch'ng et al 2008), as well as Pi3K21B and PKCδ, which both play a role in signal transduction from the insulin receptor (Braiman et al 2001;Teleman 2010).…”

Perspective: Potential Impact and Therapeutic Implications of Oncogenic PI3K Activation on Chromosomal Instability

“…Interestingly, this gene set was not enriched in MCF10A cells that had lost PTEN [77]. A study mapping the dynamic protein-protein interaction network within the core insulin signalling pathway in Drosophila also found significant enrichment for proteins involved in 'centrosome duplication' following 10 and 30 minutes of insulin stimulation [78].…”

“…Several other studies support a reciprocal relationship between the PI3K pathway and centrosomal regulation. An RNAi screen in Drosophila thus identified proteins involved in the centrosome cycle as regulators of acute insulin-dependent AKT activation [78]. Microtubule acetylation has also recently been linked to sequestration and inhibition of AKT [127].…”

Reconstruction of global regulatory network from signaling to cellular functions using phosphoproteomic data

“…However, the global regulatory network of insulin action via protein phosphorylation has not thus far been elucidated (Buescher et al, ; Chiappino‐Pepe, Pandey, Ataman, & Hatzimanikatis, ; Hatzimanikatis & Saez‐Rodriguez, ; Hyduke, Lewis, & Palsson, ; Joyce & Palsson, ; Palsson & Zengler, ). Phosphoproteomic studies of insulin signaling have been performed with Drosophila cells (Friedman et al, ; Vinayagam et al, ), 3T3‐L1 mouse adipocytes (Humphrey et al, ), mouse hepatoma cells (Monetti, Nagaraj, Sharma, & Mann, ), mouse brown preadipocytes (Krüger et al, ), rat primary hepatocytes (Zhang, Zhang, & Yu, ), rat Fao hepatoma cells (Yugi et al, ) and mouse hepatoma (Humphrey, Azimifar, & Mann, ). Among these phosphoproteomic studies, the cellular functions of insulin‐dependent protein phosphorylation have been estimated using databases of prior knowledge such as GO (Friedman et al, ; Humphrey, Azimifar, et al, ; Vinayagam et al, ; Zhang et al, ), whereas insulin‐dependent regulatory networks of protein phosphorylation by kinases have been estimated using KSR inference (Humphrey et al, ; Miller et al, ).…”