Actuation of Single Downstream Nodes in Growth Factor Network Mediates Neutrophil Guidance

Pal, Dhiman Sankar; Banerjee, Tatsat; Lin, Yiyan; Borleis, Jane; Devreotes, Peter N.

doi:10.2139/ssrn.4295347

Cited by 2 publications

(27 citation statements)

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“…Our study sheds light on the complexity of Ras function in cells. Previous reports have shown that expression of oncogenic Ras leads to excessive protrusions in multiple different cell types 4,20,61 . The extraneous protrusions impair cell movement.…”

Section: Discussionmentioning

confidence: 97%

“…We extended our investigation to differentiated HL-60 neutrophils to assess the conservation of RasGAP function on cytoskeletal remodeling. As previously reported, we developed a recruitable RASAL3 by fusing it with CRY2-mCherry, allowing for recruitment to the membrane anchor, CIBN-CAAX (Figure 2B) 4 . Recruiting RASAL3 to the F-actin-rich front, marked by LifeAct, caused mature protrusions to immediately disappear.…”

Section: Rasgaps Inhibit Ras and Pi3k Activities To Shut Off Cellular...mentioning

confidence: 99%

“…Similarly, tgRFPt-SspB R73Q ORF was introduced into pDM335 to generate tgRFPt-SspB R73Q-Ctrl construct. The construct for the membrane anchor, N150-Venus-iLID/pDM358, was made previously 4 . This construct was used to subclone PHcrac-YFP, LimE∆coil-YFP, or RBD-YFP ORF to generate dual expressing N150-Venus-iLID/PHcrac-YFP, N150-Venus-iLID/LimE-YFP, or N150-Venus-iLID/RBD-YFP construct, respectively.…”

Section: Plasmid Constructionmentioning

confidence: 99%

“…Next day, 50 μg/mL hygromycin and/or 20 μg/mL G418 sulfate was added, and cells were selected over a period of 3-4 weeks 4,88 .…”

Section: Cell Culturementioning

confidence: 99%

“…While extensive research has been dedicated to understanding the role of Ras mutations in promoting cancer growth, the impact of these mutations on cell migration and metastasis has received relatively less attention. Nevertheless, studies conducted in the model systems, Dictyostelium amoebae and human neutrophils, suggest that local Ras activity plays a direct and immediate role in cell polarity and motility [4][5][6][7][8][9] .…”

Section: Introductionmentioning

confidence: 99%

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Ras-mediated homeostatic control of front-back signaling dictates cell polarity

Lin,

Pal,

Banerjee

et al. 2023

Preprint

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Studies in the model systems, Dictyostelium amoebae and HL-60 neutrophils, have shown that local Ras activity directly regulates cell motility or polarity. Localized Ras activation on the membrane is spatiotemporally regulated by its activators, RasGEFs, and inhibitors, RasGAPs, which might be expected to create a stable front and back, respectively, in migrating cells. Focusing on C2GAPB in amoebae and RASAL3 in neutrophils, we investigated how Ras activity along the cortex controls polarity. Since existing gene knockout and overexpression studies can be circumvented, we chose optogenetic approaches to assess the immediate, local effects of these Ras regulators on the cell cortex. In both cellular systems, optically targeting the respective RasGAPs to the cell front extinguished existing protrusions and changed the direction of migration, as might be expected. However, when the expression of C2GAPB was induced globally, amoebae polarized within hours. Furthermore, within minutes of globally recruiting either C2GAPB in amoebae or RASAL3 in neutrophils, each cell type polarized and moved more rapidly. Targeting the RasGAPs to the cell backs exaggerated these effects on migration and polarity. Overall, in both cell types, RasGAP-mediated polarization was brought about by increased actomyosin contractility at the back and sustained, localized F-actin polymerization at the front. These experimental results were accurately captured by computational simulations in which Ras levels control front and back feedback loops. The discovery that context-dependent Ras activity on the cell cortex has counterintuitive, unanticipated effects on cell polarity can have important implications for future drug-design strategies targeting oncogenic Ras.

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Section: Discussionmentioning

confidence: 97%

Section: Rasgaps Inhibit Ras and Pi3k Activities To Shut Off Cellular...mentioning

confidence: 99%

Section: Plasmid Constructionmentioning

confidence: 99%

“…Next day, 50 μg/mL hygromycin and/or 20 μg/mL G418 sulfate was added, and cells were selected over a period of 3-4 weeks 4,88 .…”

Section: Cell Culturementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Ras-mediated homeostatic control of front-back signaling dictates cell polarity

Lin,

Pal,

Banerjee

et al. 2023

Preprint

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Complementary Cytoskeletal Feedback Loops Control Signal Transduction Excitability and Cell Polarity

Kuhn,

Banerjee,

Haye

et al. 2024

Preprint

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To move through complex environments, cells must constantly integrate chemical and mechanical cues. Signaling networks, such as those comprising Ras and PI3K, transmit chemical cues to the cytoskeleton, but the cytoskeleton must also relay mechanical information back to those signaling systems. Using novel synthetic tools to acutely control specific elements of the cytoskeleton in Dictyostelium and neutrophils, we delineate feedback mechanisms that alter the signaling network and promote front- or back-states of the cell membrane and cortex. First, increasing branched actin assembly increases Ras/PI3K activation while reducing polymeric actin levels overall decreases activation. Second, reducing myosin II assembly immediately increases Ras/PI3K activation and sensitivity to chemotactic stimuli. Third, inhibiting branched actin alone increases cortical actin assembly and strongly blocks Ras/PI3K activation. This effect is mitigated by reducing filamentous actin levels and in cells lacking myosin II. Finally, increasing actin crosslinking with a controllable activator of cytoskeletal regulator RacE leads to a large decrease in Ras activation both globally and locally. Curiously, RacE activation can trigger cell spreading and protrusion with no detectable activation of branched actin nucleators. Taken together with legacy data that Ras/PI3K promotes branched actin assembly and myosin II disassembly, our results define front- and back-promoting positive feedback loops. We propose that these loops play a crucial role in establishing cell polarity and mediating signal integration by controlling the excitable state of the signal transduction networks in respective regions of the membrane and cortex. This interplay enables cells to navigate intricate topologies like tissues containing other cells, the extracellular matrix, and fluids.

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Actuation of Single Downstream Nodes in Growth Factor Network Mediates Neutrophil Guidance

Cited by 2 publications

References 0 publications

Ras-mediated homeostatic control of front-back signaling dictates cell polarity

Ras-mediated homeostatic control of front-back signaling dictates cell polarity

Complementary Cytoskeletal Feedback Loops Control Signal Transduction Excitability and Cell Polarity

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