Erk/Src Phosphorylation of Cortactin Acts as a Switch On-Switch Off Mechanism That Controls Its Ability To Activate N-WASP

Ding

Proc. Natl. Acad. Sci. U.S.A.

et al. 2007

Cell movement requires morphological polarization characterized by formation of a leading pseudopodium (PD) at the front and a trailing rear at the back. However, little is known about how protein networks are spatially integrated to regulate this process at the system level. Here, we apply global proteome profiling in combination with newly developed quantitative phosphoproteomics approaches for comparative analysis of the cell body (CB) and PD proteome of chemotactic cells. The spatial relationship of 3,509 proteins and 228 distinct sites of phosphorylation were mapped revealing networks of signaling proteins that partition to the PD and/or the CB compartments. The major network represented in the PD includes integrin signaling, actin regulatory, and axon guidance proteins, whereas the CB consists of DNA/RNA metabolism, cell cycle regulation, and structural maintenance. Our findings provide insight into the spatial organization of signaling networks that control cell movement and provide a comprehensive system-wide profile of proteins and phosphorylation sites that control cell polarization.bioinformatics ͉ chemotaxis ͉ phosphoproteomics ͉ proteomics ͉ cell migration D irected cell migration or chemotaxis in response to a chemokine gradient is involved in development, immune function, angiogenesis, as well as pathological processes associated with inflammation and cancer cell metastasis (1). Cell locomotion is a highly polarized process characterized by protrusion of a leading pseudopodium (PD) (lamellipodium) at the front and establishment of a trailing rear compartment or tail region at the back (1, 2). This process is regulated through organization of the actin-myosin cytoskeleton in response to signal transduction processes that operate downstream of chemokine and integrin adhesion receptors. These receptors serve as antennae to sense changes in chemokine and extracellular matrix gradients, which provide navigational cues to direct cell movement. However, the molecular signaling mechanisms that control cell polarity and chemotaxis are not fully understood.Emerging evidence indicates that cell behavior is regulated by the complex organization of multiple signaling networks in time and space. The specific propagation of biological information in a complex biological system is achieved through the spatiotemporal assembly of multiprotein scaffolds, protein activation cascades, protein turnover, and specific posttranslational modifications of proteins, including the phosphorylation and dephosphorylation of specific amino acid residues. Although significant progress has been made in identification of specific signaling proteins and regulatory pathways that mediate cell migration, little is known about how these signals are integrated into spatial networks to achieve morphological polarity and directional movement at a system level.To understand the complexity of signal organization in migrating cells, our laboratory developed a system that facilitates the differential purification of the PD and cell body (CB) co...

Section: Functional Comparison Of Phosphoproteins In the Cb And Pdsupporting

confidence: 76%

Section: Characterization Of the Ras/erk-signaling Pathway In The Pd Bymentioning

confidence: 99%

Profiling signaling polarity in chemotactic cells

Ding

Proc. Natl. Acad. Sci. U.S.A.

et al. 2007

“…In contrast, two extracellular signal-regulated kinase (ERK) sites located at Ser-405 and Ser-418 sites, and ERK phosphorylation enhances cortactin binding and activation of N-WASP (Huang et al, 1998;Campbell et al, 1999;Weed and Parsons, 2001;Martinez-Quiles et al, 2004). Recent studies using mass spectrometry have identified 17 new sites of phosphorylation (12 serine, 4 threonine and 1 tyrosine), suggesting that cortactin is heavily phosphorylated in vivo (Martin et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Deacetylation of cortactin by SIRT1 promotes cell migration

et al. 2008

Cortactin binds F-actin and promotes cell migration. We showed earlier that cortactin is acetylated. Here, we identify SIRT1 (a class III histone deacetylase) as a cortactin deacetylase and p300 as a cortactin acetylase. We show that SIRT1 deacetylates cortactin in vivo and in vitro and that the SIRT1 inhibitor EX-527 increases amounts of acetylated cortactin in ovarian cancer cells. We also show that p300 acetylates cortactin in vivo and that cells lacking or depleted of p300 express lessacetylated cortactin than do control cells. Deletion analysis mapped the SIRT1-binding domain of cortactin to its repeat region, which also binds F-actin. Mouse embryo fibroblasts (MEFs) lacking sir2a (the mouse homolog of SIRT1) migrated more slowly than did wildtype cells. The expression of SIRT1 in sir2a-null cells restored migratory capacity, as did expression of a deacetylation-mimetic mutant of cortactin. SIRT1 and cortactin were more abundant in breast tumor tissue than in their normal counterparts, whereas SIRT1 expression inversely correlates with the ratio of acetylation cortactin versus total cortactin. These data suggest that deacetylation of cortactin is associated with high levels of SIRT1 and tumorigenesis. Finally, breast and ovarian cancer cell lines expressing an acetylation mimetic mutant of cortactin are less motile than that of control cells, whereas cells expressing the deacetylation mimetic mutant of cortactin migrate faster than that of control cells in Transwell migration assays. In summary, our results suggest that cortactin is a novel substrate for SIRT1 and p300 and, for the first time, a possible role for SIRT1 in cell motility through deacetylation of cortactin.

Journal of Biological Chemistry

“…WAVE and cortactin colocalize with Arp2/3 complex in lamellipodia, and RNAi knockdown experiments indicate that both NPFs contribute to nucleation of actin filaments in these structures (26,58,59). WAVE contains one V region and as a dimer was found to have slightly higher activity than dimeric N-WASP-VCA but less activity than dimeric WASP VCA (Fig.…”

Section: The Distinct Modes Of Activation Of Arp2/3 Complex By Cortacmentioning

confidence: 99%

“…Therefore, it is unclear whether potent synergy with cortactin is a general feature of WASP family proteins and if so, whether the mechanism is conserved. Cortactin colocalizes with WASP family proteins such as N-WASP, WASP, and WAVE (Wiskott-Aldrich syndrome protein family verproline-homologous protein) in multiple branched actin networks, including in podosomes (25), at the leading edge of lamellipodia (26), and at sites of endocytosis (27). Determining how branched actin is assembled in each of these structures will require an understanding of how each WASP family protein coordinately regulates Arp2/3 complex with cortactin.…”

mentioning

confidence: 99%

Interactions with Actin Monomers, Actin Filaments, and Arp2/3 Complex Define the Roles of WASP Family Proteins and Cortactin in Coordinately Regulating Branched Actin Networks

Helgeson

Prendergast

Wagner

et al. 2014

Background: How cortactin and WASP proteins coordinately regulate branched actin assembly is poorly understood. Results: The Arp2/3 complex-and actin-interacting regions of cortactin and WASP proteins are functionally distinct and tailored to their regulatory roles. Conclusion: Mechanistic distinctions between activators are important in allowing coordinate regulation of branched actin networks. Significance: Understanding mechanistic distinctions between activators is required to understand cellular structures like lamellipodia.