Ras GTPases mediate a wide variety of cellular processes by converting a multitude of extracellular stimuli into specific biological responses including proliferation, differentiation and survival. In mammalian cells, three ras genes encode four Ras isoforms (H-Ras, K-Ras4A, KRas4B and N-Ras) that are highly homologous but functionally distinct. Differences between the isoforms, including their post-translational modifications and intracellular sorting, mean that Ras has emerged as an important model system of compartmentalised signalling and membrane biology. Ras isoforms in different subcellular locations are proposed to recruit distinct upstream and downstream accessory proteins and activate multiple signalling pathways. Here, we summarise data relating to isoform specific signalling, its role in disease and the mechanisms promoting compartmentalised signalling. Further understanding of this field will reveal the role of Ras signalling in development, cellular homeostasis and cancers and may suggest new therapeutic approaches.

The ubiquitously expressed major Ras isoforms: H-, Kand N-Ras, are highly conserved, yet exhibit different biological outputs. We have compared the relative efficiencies with which epidermal or hepatocyte growth factor activates Ras isoforms and the requirement for specific isoforms in the activation of downstream pathways. We find that the relative coupling efficiencies to each Ras isoform are conserved between stimuli. Furthermore, in both cases, inhibition of receptor endocytosis led to reduced N-and H-Ras activation, but K-Ras was unaffected. Acute knockdown of each isoform with siRNA allows endogenous Ras isoform function and abundance to be probed. This revealed that there is significant variation in the contribution of individual isoforms to total Ras across a panel of cancer cell lines although typically KXNbH. Intriguingly, cancer cell lines where a significant fraction of endogenous Ras is oncogenically mutated showed attenuated activation of canonical Ras effector pathways. We profiled the contribution of each Ras isoform to the total Ras pool allowing interpretation of the effect of isoform-specific knockdown on signalling outcomes. In contrast to previous studies indicating preferential coupling of isoforms to Raf and PtdIns-3-kinase pathways, we find that endogenous Ras isoforms show no specific coupling to these major Ras pathways.

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The ErbB-4 receptors are unique in the EGFR/ErbB family for the ability to associate with WW domain-containing proteins. To identify new ligands of the cytoplasmic tail of ErbB-4, we panned a brain cDNA phage library with ErbB-4 peptides containing sequence motifs corresponding to putative docking sites for class-I WW domains. This approach led to identification of AIP4/Itch, a member of the Nedd4-like family of E3 ubiquitin protein ligases, as a protein that specifically interacts with and ubiquitinates ErbB-4 in vivo. Interaction with the ErbB-4 receptors occurs via the WW domains of AIP4/Itch. Functional analyses demonstrate that AIP4/Itch is recruited to the ErbB-4 receptor to promote its polyubiquitination and degradation, thereby regulating stability of the receptor and access of receptor intracellular domains to the nuclear compartment. These findings expand our understanding of the mechanisms contributing to the integrity of the ErbB signaling network and mechanistically link the cellular ubiquitination pathway of AIP4/Itch to the ErbB-4 receptor.

Differential subcellular compartmentalization of the three main Ras isoforms (H‐Ras, N‐Ras and K‐Ras) is believed to underlie their biological differences. Modulatable interactions between cellular membranes and Ras C‐terminal hypervariable region motifs determine differences in trafficking and the relative proportions of each isoform in cell‐surface signalling nanoclusters and intracellular endoplasmic reticulum/Golgi, endosomal and mitochondrial compartments. Ras regulators, effectors and scaffolds are also differentially distributed, potentially enabling preferential coupling to specific signalling pathways in each subcellular location. Here we summarize the mechanisms underlying compartment‐specific Ras signalling and the outputs generated.