Ras is a key switch controlling cell behavior. In the GTP-bound form, Ras interacts with numerous effectors in a mutually exclusive manner, where individual Ras-effectors are likely part of larger cellular (sub)complexes. The molecular details of these (sub)complexes and their alteration in specific contexts are not understood. Focusing on KRAS, we performed affinity purification (AP)-mass spectrometry (MS) experiments of exogenously expressed FLAG-KRAS WT and three oncogenic mutants ("genetic contexts") in the human Caco-2 cell line, each exposed to 11 different culture media ("culture contexts") that mimic conditions relevant in the colon and colorectal cancer. We identified four effectors present in complex with KRAS in all genetic and growth contexts ("context-general effectors"). Seven effectors are found in KRAS complexes in only some contexts ("context-specific effectors"). Analyzing all interactors in complex with KRAS per condition, we find that the culture contexts had a larger impact on interaction rewiring than genetic contexts. We investigated how changes in the interactome impact functional outcomes and created a Shiny app for interactive visualization. We validated some of the functional differences in metabolism and proliferation. Finally, we used networks to evaluate how KRAS-effectors are involved in the modulation of functions by random walk analyses of effector-mediated (sub)complexes. Altogether, our work shows the impact of environmental contexts on network rewiring, which provides insights into tissue-specific signaling mechanisms. This may also explain why KRAS oncogenic mutants may be causing cancer only in specific tissues despite KRAS being expressed in most cells and tissues.
© 2023 Ternet et al.

Akt signalling is central to cell survival, metabolism, protein and lipid homeostasis, and is impaired in Parkinson's disease (PD). Akt activation is reduced in the brain in PD, and by many PD-causing genes, including PINK1 This study investigated the mechanisms by which PINK1 regulates Akt signalling. Our results reveal for the first time that PINK1 constitutively activates Akt in a PINK1-kinase dependent manner in the absence of growth factors, and enhances Akt activation in normal growth medium. In PINK1-modified MEFs, agonist-induced Akt signalling failed in the absence of PINK1, due to PINK1 kinase-dependent increases in PI(3,4,5)P3 at both plasma membrane and Golgi being significantly impaired. In the absence of PINK1, PI(3,4,5)P3 levels did not increase in the Golgi, and there was significant Golgi fragmentation, a recognised characteristic of PD neuropathology. PINK1 kinase activity protected the Golgi from fragmentation in an Akt-dependent fashion. This study demonstrates a new role for PINK1 as a primary upstream activator of Akt via PINK1 kinase-dependent regulation of its primary activator PI(3,4,5)P3, providing novel mechanistic information on how loss of PINK1 impairs Akt signalling in PD.This article has an associated First Person interview with the first author of the paper.
© 2019. Published by The Company of Biologists Ltd.

The cancer anorexia cachexia syndrome is a systemic metabolic disorder characterized by the catabolism of stored nutrients in skeletal muscle and adipose tissue that is particularly prevalent in nonsmall cell lung cancer (NSCLC). Loss of skeletal muscle results in functional impairments and increased mortality. The aim of the present study was to characterize the changes in systemic metabolism in a genetically engineered mouse model of NSCLC. We show that a portion of these animals develop loss of skeletal muscle, loss of adipose tissue, and increased inflammatory markers mirroring the human cachexia syndrome. Using noncachexic and fasted animals as controls, we report a unique cachexia metabolite phenotype that includes the loss of peroxisome proliferator-activated receptor-α (PPARα) -dependent ketone production by the liver. In this setting, glucocorticoid levels rise and correlate with skeletal muscle degradation and hepatic markers of gluconeogenesis. Restoring ketone production using the PPARα agonist, fenofibrate, prevents the loss of skeletal muscle mass and body weight. These results demonstrate how targeting hepatic metabolism can prevent muscle wasting in lung cancer, and provide evidence for a therapeutic strategy.

The present study aimed to investigate the effect of and mechanism underlying interleukin (IL)‑17 on human retinal vascular endothelial cell (HREC) capillary tube formation in vitro. The expression of IL‑17 receptor (IL‑17R) in human HRECs was quantified using reverse transcriptase‑polymerase chain reaction (RT‑PCR) and western blot analyses. The roles of IL‑17 in HREC migration and capillary tube formation were detected using a wound scratching assay and three‑dimensional Matrigel assay, respectively, in vitro. HREC proliferation was examined using a cell counting kit‑8 assay with administration of serial doses of IL‑17. The effects of IL‑17 on the expression of vascular endothelial growth factor (VEGF), intercellular cell adhesion molecule (ICAM)‑1, IL‑6 and IL‑8 in HRECs were evaluated using RT‑PCR and western blot analyses. The results revealed that the HRECs expressed IL‑17R, and the number of intact capillary tubes formed by HRECs in the presence of IL‑17 was markedly higher, compared with that in the blank control group. The wound scratching assay showed that the numbers of migrated HRECs stimulated with IL‑17 at concentrations of 100 or 500 ng/ml were significantly higher, compared with the number in the control group. The RT‑PCR and western blot analyses showed that IL‑17 significantly promoted the expression of VEGF, ICAM‑1, IL‑6 and IL‑8 by the HRECs. The proliferation of HRECs in the presence of IL‑17 was also significantly increased. Therefore, IL‑17 increased HREC capillary tube formation through promoting HREC migration, proliferation, and expression levels of VEGF, ICAM‑1, IL‑6 and IL-8.

Signaling initiated by Class Ia phosphatidylinositol-3-kinases (PI3Ks) is essential for cell proliferation and survival. We discovered a novel protein we call PI3K interacting protein 1 (PIK3IP1) that shares homology with the p85 regulatory PI3K subunit. Using a variety of in vitro and cell based assays, we demonstrate that PIK3IP1 directly binds to the p110 catalytic subunit and down modulates PI3K activity. Our studies suggest that PIK3IP1 is a new type of PI3K regulator.