A multiprotein signaling complex sustains AKT and mTOR/S6K activity necessary for the survival of cancer cells undergoing stress.

Cancer cells encounter stresses during tumor progression and metastatic spread, however, how they survive these challenges is not fully understood. We now identify a mechanism for cancer cell survival through the discovery of a multiprotein signaling complex that includes the GTPase Cdc42, the Cdc42 GEF/effector protein Dock7, AKT, mTOR and the mTORC1 regulatory partners TSC1, TSC2, and Rheb. This pro-survival signaling complex sustains the activated state of AKT by preventing its dephosphorylation at Ser473 during serum starvation, resulting in a low but critical activation of a Raptor-independent mTOR/S6K activity. We demonstrate that the Dock7 DHR1 domain, previously of unknown function, is responsible for preserving AKT phosphorylation through an interaction requiring its C2-like motif. Collectively, these findings help address long-standing questions of how Cdc42 signals mTOR activation by elucidating the unique functions of its signaling partner Dock7 as an AKT regulator necessary for resistance to anoikis and apoptosis in cancer cells.

Inhibiting Glutamine Metabolism Blocks Coronavirus Replication in Mammalian Cells.

Developing therapeutic strategies against COVID-19 has gained widespread interest given the likelihood that new viral variants will continue to emerge. Here we describe one potential therapeutic strategy which involves targeting members of the glutaminase family of mitochondrial metabolic enzymes (GLS and GLS2), which catalyze the first step in glutamine metabolism, the hydrolysis of glutamine to glutamate. We show three examples where GLS expression increases during coronavirus infection of host cells, and another in which GLS2 is upregulated. The viruses hijack the metabolic machinery responsible for glutamine metabolism to generate the building blocks for biosynthetic processes and satisfy the bioenergetic requirements demanded by the 'glutamine addiction' of virus-infected host cells. We demonstrate how genetic silencing of glutaminase enzymes reduces coronavirus infection and that newer members of two classes of small molecule allosteric inhibitors targeting these enzymes, designated as SU1, a pan-GLS/GLS2 inhibitor, and UP4, which is specific for GLS, block viral replication in mammalian epithelial cells. Overall, these findings highlight the importance of glutamine metabolism for coronavirus replication in human cells and show that glutaminase inhibitors can block coronavirus infection and thereby may represent a novel class of anti-viral drug candidates. Teaser: Inhibitors targeting glutaminase enzymes block coronavirus replication and may represent a new class of anti-viral drugs.

Identification of mTORC2 as a necessary component of HRG/ErbB2-dependent cellular transformation.

UNLABELLED: Overexpression of the receptor tyrosine kinase HER2/ErbB2 (ERBB2) has been linked to a poor prognosis for patients with breast cancer; thus, its activity is a central target for cancer therapy. Likewise, overexpression of heregulin (HRG/NRG1), a growth factor responsible for ErbB2 activation, has also been shown to be a driver of breast cancer progression. Although ErbB2 inhibitors offer a major advancement in the treatment of ErbB2-dependent breast cancers, patients are highly susceptible to developing clinical resistance to these drugs. Therefore, a detailed understanding of the molecular mechanism that underlies HRG/ErbB2-induced tumorigenesis is essential for the development of effective therapeutic strategies for this subset of patients with breast cancer. Here, it was demonstrated that HRG promoted anchorage-independent breast cancer cell growth more potently than EGF, and that the HRG-dependent activation of phosphoinositide 3-kinase and mTORC1 are necessary events for cell transformation. Functional evaluation of two distinct mTOR (MTOR) inhibitors, rapamycin and INK-128, on HRG-dependent signaling activities, uncovered a necessary role for mTORC2 in the regulation of the AKT/TSC2/mTORC1 axis by affecting the phosphorylation of AKT at the PDK1(PDPK1)-dependent site (T308) as well as at the mTORC2-dependent site (S473). The elimination of Rictor (RICTOR), a critical component of mTORC2, is detrimental to both the activation of mTORC1 and HRG-mediated cellular transformation. Similar results were obtained in multiple breast cancer model systems, highlighting an important role for mTORC2 in HRG/ErbB2-dependent breast cancer. IMPLICATIONS: These findings suggest the potential benefits of targeting mTORC2 in HRG/ErbB2-induced breast cancer.

Activation of the Ran GTPase is subject to growth factor regulation and can give rise to cellular transformation.

Although the small GTPase Ran is best known for its roles in nucleocytoplasmic transport, mitotic spindle assembly, and nuclear envelope formation, recent studies have demonstrated the overexpression of Ran in multiple tumor types and that its expression is correlated with a poor patient prognosis, providing evidence for the importance of this GTPase in cell growth regulation. Here we show that Ran is subject to growth factor regulation by demonstrating that it is activated in a serum-dependent manner in human breast cancer cells and, in particular, in response to heregulin, a growth factor that activates the Neu/ErbB2 tyrosine kinase. The heregulin-dependent activation of Ran requires mTOR (mammalian target of rapamycin) and stimulates the capped RNA binding capability of the cap-binding complex in the nucleus, thus influencing gene expression at the level of mRNA processing. We further demonstrate that the excessive activation of Ran has important consequences for cell growth by showing that a novel, activated Ran mutant is sufficient to transform NIH-3T3 cells in an mTOR- and epidermal growth factor receptor-dependent manner and that Ran-transformed cells form tumors in mice.

The molecular basis for the regulation of the cap-binding complex by the importins.

The binding of capped RNAs to the cap-binding complex (CBC) in the nucleus, and their dissociation from the CBC in the cytosol, represent essential steps in RNA processing. Here we show how the nucleocytoplasmic transport proteins importin-alpha and importin-beta have key roles in regulating these events. As a first step toward understanding the molecular basis for this regulation, we determined a 2.2-A resolution X-ray structure for a CBC-importin-alpha complex that provides a detailed picture for how importin-alpha binds to the CBP80 subunit of the CBC. Through a combination of biochemical studies, X-ray crystallographic information and small-angle scattering experiments, we then determined how importin-beta binds to the CBC through its CBP20 subunit. Together, these studies enable us to propose a model describing how importin-beta stimulates the dissociation of capped RNA from the CBC in the cytosol following its nuclear export.