Growing evidence suggests a major role for Src-homology-2-domain-containing phosphatase 2 (SHP2/PTPN11) in MYCN-driven high-risk neuroblastoma, although biologic confirmation and a plausible mechanism for this contribution are lacking. Using a zebrafish model of MYCN-overexpressing neuroblastoma, we demonstrate that mutant ptpn11 expression in the adrenal gland analog of MYCN transgenic fish promotes the proliferation of hyperplastic neuroblasts, accelerates neuroblastomagenesis, and increases tumor penetrance. We identify a similar mechanism in tumors with wild-type ptpn11 and dysregulated Gab2, which encodes a Shp2 activator that is overexpressed in human neuroblastomas. In MYCN transgenic fish, Gab2 overexpression activated the Shp2-Ras-Erk pathway, enhanced neuroblastoma induction, and increased tumor penetrance. We conclude that MYCN cooperates with either GAB2-activated or mutant SHP2 in human neuroblastomagenesis. Our findings further suggest that combined inhibition of MYCN and the SHP2-RAS-ERK pathway could provide effective targeted therapy for high-risk neuroblastoma patients with MYCN amplification and aberrant SHP2 activation.

Lysine succinylation was recently identified as a post-translational modification in cells. However, the molecular mechanism underlying lysine succinylation remains unclear. Here, we show that carnitine palmitoyltransferase 1A (CPT1A) has lysine succinyltransferase (LSTase) activity in vivo and in vitro. Using a stable isotope labeling by amino acid in cell culture (SILAC)-based proteomics approach, we found that 101 proteins were more succinylated in cells expressing wild-type (WT) CPT1A compared with vector control cells. One of the most heavily succinylated proteins in this analysis was enolase 1. We found that CPT1A WT succinylated enolase 1 and reduced enolase enzymatic activity in cells and in vitro. Importantly, mutation of CPT1A Gly710 (G710E) selectively inactivated carnitine palmitoyltransferase (CPTase) activity but not the LSTase activity that decreased enolase activity in cells and promoted cell proliferation under glutamine depletion. These findings suggest that CPT1A acts as an LSTase that can regulate enzymatic activity of a substrate protein and metabolism independent of its classical CPTase activity.

Cancer progresses through distinct stages, and mouse models recapitulating traits of this progression are frequently used to explore genetic, morphological, and pharmacological aspects of tumor development. To complement genomic investigations of this process, we here quantify phosphoproteomic changes in skin cancer development using the SILAC mouse technology coupled to high-resolution mass spectrometry. We distill protein expression signatures from our data that distinguish between skin cancer stages. A distinct phosphoproteome of the two stages of cancer progression is identified that correlates with perturbed cell growth and implicates cell adhesion as a major driver of malignancy. Importantly, integrated analysis of phosphoproteomic data and prediction of kinase activity revealed PAK4-PKC/SRC network to be highly deregulated in SCC but not in papilloma. This detailed molecular picture, both at the proteome and phosphoproteome level, will prove useful for the study of mechanisms of tumor progression.