Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation

Herrington, Kari A.; Trinh, Andrew; Dang, Carolyn T.; O’Shaughnessy, Ellen C.; Hahn, Klaus M.; Gratton, Enrico; Digman, Michelle A.; Sütterlin, Christine

doi:10.1091/mbc.e16-09-0665

Cited by 17 publications

(15 citation statements)

References 56 publications

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“…Thus, we favour the hypothesis that ribbon integrity may be necessary to build a rigid structural platform for the anchoring of asymmetric MTs and for the targeted delivery of cargoes and signalling proteins to PM subdomains (Sutterlin and Colanzi, 2010). In agreement with this suggestion, common features of golgin depletion are the reduced efficacy of polarised delivery of a cargo reporter to Review the leading edge (Yadav et al, 2009) and the lack of PM-associated cdc42 activation (Herrington et al, 2017).…”

Section: Key Regulators Of Golgi Structuresupporting

confidence: 77%

Mitotic inheritance of the Golgi complex and its role in cell division

Ayala

Colanzi

2017

Biology of the Cell

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The Golgi apparatus plays essential roles in the processing and sorting of proteins and lipids, but it can also act as a signalling hub and a microtubule-nucleation centre. The Golgi complex (GC) of mammalian cells is composed of stacks connected by tubular bridges to form a continuous membranous system. In spite of this structural complexity, the GC is highly dynamic, and this feature becomes particularly evident during mitosis, when the GC undergoes a multi-step disassembly process that allows its correct partitioning and inheritance by daughter cells. Strikingly, different steps of Golgi disassembly control mitotic entry and progression, indicating that cells actively monitor Golgi integrity during cell division. Here, we summarise the basic mechanisms and the molecular players that are involved in Golgi disassembly, focussing in particular on recent studies that have revealed the fundamental signalling pathways that connect Golgi inheritance to mitotic entry and progression.

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Section: Key Regulators Of Golgi Structuresupporting

confidence: 77%

Mitotic inheritance of the Golgi complex and its role in cell division

Ayala

Colanzi

2017

Biology of the Cell

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“…Likewise, in mammalian cells, Cdc42 has been detected on Arf6-positive intracellular vesicles, which are trafficked en route to the leading edge of migration cells [55]. Notably, a connected Golgi ribbon was necessary for Cdc42 activity at the PM, further confirming the postulated role of the Golgi-localized Cdc42 in replenishing PM Cdc42 pools [62].…”

Section: Rho Gtpases Acting Along the Secretory Pathwaymentioning

confidence: 73%

“…Importantly, live-cell imaging using biosensors has provided spatial information on the activity of Cdc42 at the Golgi complex [33,61]. The development of a new generation of genetically encoded fluorescence resonance energy transfer (FRET) biosensors with high selectivity, sensitivity, and dynamic range has further pushed the field forward [62][63][64].…”

Section: Rho Gtpases Acting Along the Secretory Pathwaymentioning

confidence: 99%

“…Up to date, four Cdc42-specific GEFs, Tuba (DNMBP), faciogenital dysplasia 1 protein (FGD1), intersectin-1 (ITSN1), and Dbs-130 (MCF2L) have been reported to localize to the Golgi. Endogenous Tuba was initially observed at the Golgi membranes in rat brain cryosections and in HeLa cells using indirect immunofluorescence and co-staining with GM130 [66,67] and has been reported to regulate Cdc42 activity in this compartment [62]. By performing fluorescence lifetime-resolved imaging microscopy (FLIM)-FRET measurements in living cells, Sütterlin and co-workers showed that the activity of Golgi-localized Cdc42 was regulated by ARHGAP21 and Tuba but not FGD1 [62].…”

Section: Rho Gtpases Acting Along the Secretory Pathwaymentioning

confidence: 99%

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Spatiotemporal Control of Intracellular Membrane Trafficking by Rho GTPases

Noll

Haußer

2019

Cells

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As membrane-associated master regulators of cytoskeletal remodeling, Rho GTPases coordinate a wide range of biological processes such as cell adhesion, motility, and polarity. In the last years, Rho GTPases have also been recognized to control intracellular membrane sorting and trafficking steps directly; however, how Rho GTPase signaling is regulated at endomembranes is still poorly understood. In this review, we will specifically address the local Rho GTPase pools coordinating intracellular membrane trafficking with a focus on the endo-and exocytic pathways. We will further highlight the spatiotemporal molecular regulation of Rho signaling at endomembrane sites through Rho regulatory proteins, the GEFs and GAPs. Finally, we will discuss the contribution of dysregulated Rho signaling emanating from endomembranes to the development and progression of cancer.an inactive GDP-bound and an active GTP-bound state. Rho GDP/GTP cycling is tightly regulated by guanine nucleotide exchange factors (GEFs) that promote the formation of the active GTP-bound form through exchanging GDP for GTP. On the contrary, GTPase-activating proteins (GAPs) catalyze the intrinsic GTPase activity and promote the formation of inactive GDP-bound Rho. This inactive state can be subsequently recognized by guanine nucleotide dissociation inhibitors (GDIs), which sequester Rho GTPases in the cytosol [5,6] (Figure 1). Whereas classical Rho GTPases are regulated by GDP/GTP cycling, atypical Rho GTPases are regulated by other mechanisms that occur in particular at the transcriptional and post-translational level [7]. In the case of atypical Rho GTPases, GTP is usually constitutively bound, either because these Rho GTPases possess high intrinsic nucleotide exchange activity or have substitutions in their GTPase domain that prevent GTP hydrolysis.

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“…For studies of GEF-GTPase circuitry, we focused on GEFs that were stimulated by EGF, to examine protrusion/retraction cycles during EGF-induced chemokinesis. Biosensors for Asef, Vav2, and improved versions of our previously published biosensors for Rac1 3 and Cdc42 32,33 (Supplemental methods and Extended Data Figure 3a) were stably expressed in MDA-MB-231 cells and studied in EGF-containing medium. Biosensors were kept below expression levels that perturbed motility behaviors (Extended Data Figure 4).…”

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confidence: 99%

Connectivity analysis of GEF/GTPase networks in living cells

Marston

Vilela

Ren

et al. 2019

Preprint

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contributionsThe cytoskeleton is regulated by dynamic, multi-layered signaling networks that interconnect Rho family small GTPases with exquisite spatiotemporal precision 1 .Understanding the organization of these networks is challenging, as protein activation and interaction occur transiently and with precise subcellular localization. We and others have used fluorescent biosensors in living cells to map the activation patterns of the Rho family small GTPases relative to the changes in cell edge dynamics that they produce 2-4 . These GTPases are controlled by the localized activity of numerous Rho guanine nucleotide exchange factors (RhoGEFs) with overlapping GTPase specificity 5,6 . Here we extend this analysis to determine functional relationships between GEFs and GTPases in controlling cell edge movements. First, biosensors for GEF activation are produced, and their activity correlated with edge dynamics. We then shift the wavelengths of GTPase biosensors to image and correlate the activation of GEFs and GTPases in the same cell. Using partial correlation analysis, we can parse out from such multiplexed data the contribution of each GEF -GTPase interaction to edge dynamics, i.e. we identify when and where specific GEF activation events regulate specific downstream GTPases to affect motility. We describe biosensors for eight Dbl family RhoGEFs based on a broadly applicable new design strategy (Asef, Tiam1, Vav isoforms, Tim, LARG, and β -Pix), and red shifted biosensors for RhoA, Rac1 and Cdc42. In the context of motility, functional interactions were identified for Asef regulation of Cdc42 and Rac1. This approach exemplifies a powerful means to elucidate the real-time connectivity of signal transduction networks.In previous studies of Rho GTPases in motility we could determine the relative timing and location of Rac1, Cdc42, RhoA, and RhoC activation by first using fluorescent biosensors to

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Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation

Abstract: A Cdc42 biosensor and the phasor approach to FLIM-FRET are used to determine Cdc42 activity at specific subcellular locations, revealing new regulatory principles and functions of this small GTPase.

Cited by 17 publications

References 56 publications

Mitotic inheritance of the Golgi complex and its role in cell division

Mitotic inheritance of the Golgi complex and its role in cell division

Spatiotemporal Control of Intracellular Membrane Trafficking by Rho GTPases

Connectivity analysis of GEF/GTPase networks in living cells

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