Polarized focal adhesion kinase activity within a focal adhesion during cell migration

Li, Xiaoquan; Combs, Joseph Dale; Salaita, Khalid; Shu, Xiaokun

doi:10.1038/s41589-023-01353-y

Cited by 21 publications

(11 citation statements)

References 68 publications

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“…The proportion of mobile versus immobile nanocluster regions was measured and each of their half-life was calculated. This was measured for different focal adhesion proteins which serve distinct functions-integrin transmembrane receptors bridge the ECM to the cell interior, that upon activation activates FAK [13][14][15][16]83], a key force-dependent kinase required to activate downstream signalling molecules. For instance paxillin which is a LIM domain protein that upon force-dependent phosphorylation is translocated to the nucleus to bring about regulation of specific gene expression [18,19].…”

Section: Discussionmentioning

confidence: 99%

“…Taken together, our results showed that signalling active phosphorylated FAK immobilises in nanoclusters that serve as signalling hubs. FAK is a force dependent kinase, which is phosphorylated and activated on stiff substrates [14,16,70]. Hence, if the stable nanoclusters of FAK are signalling hubs necessary for signalling, absence of force would eliminate these nanoclusters.…”

Section: Nanokymo-flap/frap Reveals Differential Dynamics Of Core Foc...mentioning

confidence: 99%

“…One key signalling event that is the most well studied hallmark of signalling in biology, in response to external signals, is phosphorylation of kinases, e.g. force dependent phosphorylation of focal adhesion kinase (FAK) at focal adhesions on rigid substrates [13][14][15][16], or DAPK1 at podosomes on soft substrates [17], which in turn phosphorylate downstream signalling molecules and activates them. This signalling coordinates downstream cellular functions including cytoskeletal reorganisation, gene expression (regulated by translocation of activated LIM domain protein); paxillin translocation to the nucleus (activated by phosphorylation at the focal adhesions) [18,19], or DAPK1-induced anoikis on soft substrates [17], among others.…”

Section: Introductionmentioning

confidence: 99%

“…This signalling coordinates downstream cellular functions including cytoskeletal reorganisation, gene expression (regulated by translocation of activated LIM domain protein); paxillin translocation to the nucleus (activated by phosphorylation at the focal adhesions) [18,19], or DAPK1-induced anoikis on soft substrates [17], among others. The dynamics of adhesions is closely regulated by force dependent activation of kinases such as FAK, and deactivation of signal by force dependent phosphatase-PTPN12, (also known as PTP-Pest) [13][14][15][16][20][21][22][23]. The absence of PTPN12 function leads to the formation of large adhesions on compliant substrates lacking force [24].…”

Section: Introductionmentioning

confidence: 99%

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Nano-clusters of ligand-activated integrins organize immobile, signalling active, nano-clusters of phosphorylated FAK required for mechanosignaling in focal adhesions

Jain,

Minhaj,

Kanchanawong

et al. 2024

Preprint

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Transmembrane signalling receptors, such as integrins, organise as nanoclusters that are thought to provide several advantages including, increasing avidity, sensitivity (increasing the signal-to-noise ratio) and robustness (signalling above a threshold rather than activation by a single receptor) of the signal compared to signalling by single receptors. Compared to large micron-sized clusters, nanoclusters offer the advantage of rapid turnover for the disassembly of the signal. However, if nanoclusters function as signalling hubs remains poorly understood. Here, we employ fluorescence nanoscopy combined with photoactivation and photobleaching at sub-diffraction limited resolution of ∼100nm length scale within a focal adhesion to examine the dynamics of diverse focal adhesion proteins. We show that (i) subregions of focal adhesions are enriched in immobile population of integrin β3 organised as nanoclusters, which (ii) in turn serve to organise nanoclusters of associated key adhesome proteins-vinculin, focal adhesion kinase (FAK) and paxillin, demonstrating that signalling proceeds by formation of nanoclusters rather than through individual proteins. (iii) Distinct focal adhesion protein nanoclusters exhibit distinct dynamics dependent on function. (iv) long-lived nanoclusters function as signalling hubs-wherein phosphorylated FAK and paxillin formed stable nanoclusters in close proximity to immobile integrin nanoclusters which are disassembled in response to inactivation signal by phosphatase PTPN12 (v) signalling takes place in response to an external signal such as force or geometric arrangement of the nanoclusters and when the signal is removed, these nanoclusters disassemble. Taken together, these results demonstrate that signalling downstream of transmembrane receptors is organised as hubs of signalling proteins (FAK, paxillin, vinculin) seeded by nanoclusters of the transmembrane receptor (integrin).

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Section: Discussionmentioning

confidence: 99%

Section: Nanokymo-flap/frap Reveals Differential Dynamics Of Core Foc...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Nano-clusters of ligand-activated integrins organize immobile, signalling active, nano-clusters of phosphorylated FAK required for mechanosignaling in focal adhesions

Jain,

Minhaj,

Kanchanawong

et al. 2024

Preprint

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show abstract

“…These optogenetics-driven biosensors exhibited superior performance when compared to their non-optimized counterparts. Moreover, to further improve the resolution of biosensors, we used CRY2/CIBN system to optically regulate the phase separation dynamics of Homo-Oligomeric Tag3 (HOTag3) and HOTag6 [15][16][17] . This innovative approach led to a substantial improvement in the detection resolution of the phosphatidylinositol-4-phosphate (PI4P) biosensor.…”

mentioning

confidence: 99%

Optogenetic strategies for optimizing the performance of biosensors of membrane phospholipids in live cells

Yao,

Lou,

Jin

et al. 2023

Preprint

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High-performance biosensors are crucial for elucidating the spatiotemporal regulatory roles and dynamics of membrane lipids, but there is a lack of improvement strategies for biosensors with low sensitivity and low-content substrates detection. Here we developed universal optogenetic strategies to improve a set of membrane biosensors by trapping them into specific region and further reducing the background signal, or by optically-controlled phase separation for membrane lipids detection and tracking. These improved biosensors were superior to typical tools and light simulation would enhance their detection performance and resolution, which might contribute to the design and optimization of other biosensors.

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Optogenetic Strategies for Optimizing the Performance of Phospholipids Biosensors

Yao,

Lou,

Jin

et al. 2024

Advanced Science

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High‐performance biosensors play a crucial role in elucidating the intricate spatiotemporal regulatory roles and dynamics of membrane phospholipids. However, enhancing the sensitivity and imaging performance remains a significant challenge. Here, optogenetic‐based strategies are presented to optimize phospholipid biosensors. These strategies involves presequestering unbound biosensors in the cell nucleus and regulating their cytosolic levels with blue light to minimize background signal interference in phospholipid detection, particularly under conditions of high expression levels of biosensor. Furthermore, optically controlled phase separation and the SunTag system are employed to generate punctate probes for substrate detection, thereby amplifying biosensor signals and enhancing visualization of the detection process. These improved phospholipid biosensors hold great potential for enhancing the understanding of the spatiotemporal dynamics and regulatory roles of membrane lipids in live cells and the methodological insights in this study might be valuable for developing other high‐performance biosensors.

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Polarized focal adhesion kinase activity within a focal adhesion during cell migration

Cited by 21 publications

References 68 publications

Nano-clusters of ligand-activated integrins organize immobile, signalling active, nano-clusters of phosphorylated FAK required for mechanosignaling in focal adhesions

Nano-clusters of ligand-activated integrins organize immobile, signalling active, nano-clusters of phosphorylated FAK required for mechanosignaling in focal adhesions

Optogenetic strategies for optimizing the performance of biosensors of membrane phospholipids in live cells

Optogenetic Strategies for Optimizing the Performance of Phospholipids Biosensors

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