In humans, the S100 protein family is composed of 21 members that exhibit a high degree of structural similarity, but are not functionally interchangeable. This family of proteins modulates cellular responses by functioning both as intracellular Ca2+ sensors and as extracellular factors. Dysregulated expression of multiple members of the S100 family is a common feature of human cancers, with each type of cancer showing a unique S100 protein profile or signature. Emerging in vivo evidence indicates that the biology of most S100 proteins is complex and multifactorial, and that these proteins actively contribute to tumorigenic processes such as cell proliferation, metastasis, angiogenesis and immune evasion. Drug discovery efforts have identified leads for inhibiting several S100 family members, and two of the identified inhibitors have progressed to clinical trials in patients with cancer. This Review highlights new findings regarding the role of S100 family members in cancer diagnosis and treatment, the contribution of S100 signalling to tumour biology, and the discovery and development of S100 inhibitors for treating cancer.

Cellular lamellipodia bind to the matrix and probe its rigidity through forces generated by rearward F-actin transport. Cells respond to matrix rigidity by moving toward more rigid matrices using an unknown mechanism. In spreading and migrating cells we find local periodic contractions of lamellipodia that depend on matrix rigidity, fibronectin binding and myosin light chain kinase (MLCK). These contractions leave periodic rows of matrix bound beta3-integrin and paxillin while generating waves of rearward moving actin bound alpha-actinin and MLCK. The period between contractions corresponds to the time for F-actin to move across the lamellipodia. Shortening lamellipodial width by activating cofilin decreased this period proportionally. Increasing lamellipodial width by Rac signaling activation increased this period. We propose that an actin bound, contraction-activated signaling complex is transported locally from the tip to the base of the lamellipodium, activating the next contraction/extension cycle.

Viral infections continue to represent major challenges to public health, and an enhanced mechanistic understanding of the processes that contribute to viral life cycles is necessary for the development of new therapeutic strategies . Viperin, a member of the radical S-adenosyl-L-methionine (SAM) superfamily of enzymes, is an interferon-inducible protein implicated in the inhibition of replication of a broad range of RNA and DNA viruses, including dengue virus, West Nile virus, hepatitis C virus, influenza A virus, rabies virus and HIV. Viperin has been suggested to elicit these broad antiviral activities through interactions with a large number of functionally unrelated host and viral proteins. Here we demonstrate that viperin catalyses the conversion of cytidine triphosphate (CTP) to 3'-deoxy-3',4'-didehydro-CTP (ddhCTP), a previously undescribed biologically relevant molecule, via a SAM-dependent radical mechanism. We show that mammalian cells expressing viperin and macrophages stimulated with IFNα produce substantial quantities of ddhCTP. We also establish that ddhCTP acts as a chain terminator for the RNA-dependent RNA polymerases from multiple members of the Flavivirus genus, and show that ddhCTP directly inhibits replication of Zika virus in vivo. These findings suggest a partially unifying mechanism for the broad antiviral effects of viperin that is based on the intrinsic enzymatic properties of the protein and involves the generation of a naturally occurring replication-chain terminator encoded by mammalian genomes.

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SummaryWe have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (Mena INV ) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated Mena INV increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by Mena INV is dependent on a macrophage-tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of Mena INV and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells. Journal of Cell Science metastatic progression. Mena is a member of the Ena/VASP family of proteins and binds actin to regulate the geometry and assembly of filament networks through: (1) an anti-capping protein activity (Bear et al., 2002;Barzik et al., 2005; Hansen and Mullins, 2010) that involves binding to profilin and both G-and F-actin; (2) Mena tetramerization, and (3) reduction in the density of actin-related proteins 2 and 3 (Arp2/3)-mediated branching (Gertler et al., 1996;Barzik et al., 2005;Ferron et al., 2007;Pasic et al., 2008;Bear and Gertler, 2009; Hansen and Mullins, 2010). Alternative splicing for the Mena gene has been reported: a 19 amino acid residue insertion just after the EVH1 domain generates the Mena invasion isoform (Mena INV , formerly Mena +++ ) (Gertler et al., 1996;Philippar et al., 2008), whereas a 21 residue insertion in the EVH2 domain generates the Mena11a isoform (Di Modugno et al., 2007). A comparison of the invasive and migratory tumor cells collected in vivo, with primary tumor cells isolated from mouse, rat and human cell-line-derived mammary tumors, revealed that Mena INV expression is upregulated and Mena11a is downregulated selectively in the invasive and migrating carcinoma cell population (Goswami et al., 2009). The differential regulation of Mena isoforms across species suggests that these two isoforms have important roles in invasion and metastasis.In previous studies, we showed that expression of Mena INV in a xenograft mouse mammary tumor promotes increased formation of spontaneous lung metastases from orthotopic tumors and alters the sensitivity of tumor cells to epidermal growth factor (EGF) . We undertook the current study to identify the step(s) in the metastatic cascade that are affected by Mena INV expression and investigate the effect of expression of...

Synergistic activation by heterotrimeric guanine nucleotide–binding protein (G protein)-coupled receptors (GPCRs) and receptor tyrosine kinases distinguishes p110β from other class IA phosphoinositide 3-kinases (PI3Ks). Activation of p110β is specifically implicated in various physiological and pathophysiological processes, such as the growth of tumors deficient in phosphatase and tensin homolog deleted from chromosome 10 (PTEN). To determine the specific contribution of GPCR signaling to p110β-dependent functions, we identified the site in p110β that binds to the Gβγ subunit of G proteins. Mutation of this site eliminated Gβγ-dependent activation of PI3Kβ (a dimer of p110β and the p85 regulatory subunit) in vitro and in cells, without affecting basal activity or phosphotyrosine peptide–mediated activation. Disrupting the p110β-Gβγ interaction by mutation or with a cell-permeable peptide inhibitor blocked the transforming capacity of PI3Kβ in fibroblasts, and reduced proliferation, chemotaxis, and invasiveness of PTEN-null tumor cells in culture. Our data suggest that specifically targeting GPCR signaling to PI3Kβ could provide a therapeutic approach for tumors that depend on p110β for growth and metastasis.

Phosphoinositide 3-kinase gamma (PI3Kγ) has profound roles downstream of G-protein-coupled receptors in inflammation, cardiac function, and tumor progression. To gain insight into how the enzyme's activity is shaped by association with its p101 adaptor subunit, lipid membranes, and Gβγ heterodimers, we mapped these regulatory interactions using hydrogen-deuterium exchange mass spectrometry. We identify residues in both the p110γ and p101 subunits that contribute critical interactions with Gβγ heterodimers, leading to PI3Kγ activation. Mutating Gβγ-interaction sites of either p110γ or p101 ablates G-protein-coupled receptor-mediated signaling to p110γ/p101 in cells and severely affects chemotaxis and cell transformation induced by PI3Kγ overexpression. Hydrogen-deuterium exchange mass spectrometry shows that association with the p101 regulatory subunit causes substantial protection of the RBD-C2 linker as well as the helical domain of p110γ. Lipid interaction massively exposes that same helical site, which is then stabilized by Gβγ. Membrane-elicited conformational change of the helical domain could help prepare the enzyme for Gβγ binding. Our studies and others identify the helical domain of the class I PI3Ks as a hub for diverse regulatory interactions that include the p101, p87 (also known as p84), and p85 adaptor subunits; Rab5 and Gβγ heterodimers; and the β-adrenergic receptor kinase.