Intracellular Force Research Articles

Detection of microtubules in vivo using antibody-immobilized nanoneedles

We present here an alternative, force-based measurement method for the detection of intracellular cytoskeletal proteins in the live cell. High aspect ratio nanoneedles of 200 nm in diameter were functionalized with anti-tubulin antibodies and inserted, using an atomic force microscope (AFM), into live NIH3T3 cells, without affecting cell viability. Force curves were recorded during insertion and evacuation of nanoneedles from the cells, and used to analyse intracellular interactions of the nanoneedles with the microtubule cytoskeleton during evacuation from the cell. Disruption of microtubules led to a correlated time-dependent decrease in the measured intracellular binding forces, pointing to the high-sensitivity and high-specificity of this detection method. This analytical technique allows for real-time evaluation of the microtubule network in the live cell, without the need to use potentially harmful molecular markers as do conventional detection methods, and may prove beneficial in the diagnosis and investigation of cytoskeleton-associated diseases.

Acoustic tweezing cytometry for live-cell subcellular modulation of intracellular cytoskeleton contractility

Mechanical forces are critical to modulate cell spreading, contractility, gene expression, and even stem cell differentiation. Yet, existing tools that can apply controllable subcellular forces to a large number of single cells simultaneously are still limited. Here we report a novel ultrasound tweezing cytometry utilizing ultrasound pulses to actuate functionalized lipid microbubbles covalently attached to single live cells to exert mechanical forces in the pN - nN range. Ultrasonic excitation of microbubbles could elicit a rapid and sustained reactive intracellular cytoskeleton contractile force increase in different adherent mechanosensitive cells. Further, ultrasound-mediated intracellular cytoskeleton contractility enhancement was dose-dependent and required an intact actin cytoskeleton as well as RhoA/ROCK signaling. Our results demonstrated the great potential of ultrasound tweezing cytometry technique using functionalized microbubbles as an actuatable, biocompatible, and multifunctional agent for biomechanical stimulations of cells.

Apical Oscillations in Amnioserosa Cells: Basolateral Coupling and Mechanical Autonomy

Holographic laser microsurgery is used to isolate single amnioserosa cells in vivo during early dorsal closure. During this stage of Drosophila embryogenesis, amnioserosa cells undergo oscillations in apical surface area. The postisolation behavior of individual cells depends on their preisolation phase in these contraction/expansion cycles: cells that were contracting tend to collapse quickly after isolation; cells that were expanding do not immediately collapse, but instead pause or even continue to expand for ∼40 s. In either case, the postisolation apical collapse can be prevented by prior anesthetization of the embryos with CO2. These results suggest that although the amnioserosa is under tension, its cells are subjected to only small elastic strains. Furthermore, their postisolation apical collapse is not a passive elastic relaxation, and both the contraction and expansion phases of their oscillations are driven by intracellular forces. All of the above require significant changes to existing computational models.

The Role of Mechanical Force and ROS in Integrin-Dependent Signals

Cells are exposed to several types of integrin stimuli, which generate responses generally referred to as “integrin signals”, but the specific responses to different integrin stimuli are poorly defined. In this study, signals induced by integrin ligation during cell attachment, mechanical force from intracellular contraction, or cell stretching by external force were compared. The elevated phosphorylation levels of several proteins during the early phase of cell attachment and spreading of fibroblast cell lines were not affected by inhibition of ROCK and myosin II activity, i.e. the reactions occurred independently of intracellular contractile force acting on the adhesion sites. The contraction-independent phosphorylation sites included ERK1/2 T202/Y204, AKT S473, p130CAS Y410, and cofilin S3. In contrast to cell attachment, cyclic stretching of the adherent cells induced a robust phosphorylation only of ERK1/2 and the phosphorylation levels of the other investigated proteins were not or only moderately affected by stretching. No major differences between signaling via α5β1 or αvβ3 integrins were detected. The importance of mitochondrial ROS for the integrin-induced signaling pathways was investigated using rotenone, a specific inhibitor of complex I in the respiratory chain. While rotenone only moderately reduced ATP levels and hardly affected the signals induced by cyclic cell stretching, it abolished the activation of AKT and reduced the actin polymerization rate in response to attachment in both cell lines. In contrast, scavenging of extracellular ROS with catalase or the vitamin C analog Asc-2P did not significantly influence the attachment-derived signaling, but caused a selective and pronounced enhancement of ERK1/2 phosphorylation in response to stretching. In conclusion, the results showed that “integrin signals” are composed of separate sets of reactions triggered by different types of integrin stimulation. Mitochondrial ROS and extracellular ROS had specific and distinct effects on the integrin signals induced by cell attachment and mechanical stretching.

High extensibility of stress fibers revealed by in vitro micromanipulation with fluorescence imaging

Stress fibers (SFs), subcellular bundles of actin and myosin filaments, are physically connected at their ends to cell adhesions. The intracellular force transmitted via SFs plays an essential role in cell adhesion regulation and downstream signaling. However, biophysical properties intrinsic to individual SFs remain poorly understood partly because SFs are surrounded by other cytoplasmic components that restrict the deformation of the embedded materials. To characterize their inherent properties independent of other structural components, we isolated SFs from vascular smooth muscle cells and mechanically stretched them by in vitro manipulation while visualizing strain with fluorescent quantum dots attached along their length. SFs were elongated along their entire length, with the length being approximately 4-fold of the stress-free length. This surprisingly high extensibility was beyond that explained by the tandem connection of actin filaments and myosin II bipolar filaments present in SFs, thus suggesting the involvement of other structural components in their passive biophysical properties.

Different Function of α5β1 and αVβ3 Integrins: Elucidating their Interactions and Spatial Requirements for Adhesion and Migration

The coordination of specific functions of α5β1 and αvβ3 integrins is crucial for cell spreading and migration. For the spatio-temporal transduction of signals, cells interacting with their environment regulate the localization of α5β1 and αvβ3 integrins at adhesive contacts. The understanding of integrin function has progressed because of new techniques and analytical tools; however, the analysis of integrin specific contribution and crosstalk during adhesion remains a challenge.To determine αvβ3 and α5β1 integrin specific functions we used nanoarrays of gold particles presenting immobilized integrin-selective peptidomimetics. For measuring forces generated upon adhesion-mediated by these integrins, we produced pillar arrays decorated with gold particles presenting the peptidomimetics. Finally, to characterize integrin crosstalk, we achieved the controlled sumultaneous presentation of αvβ3 and α5β1 integrin peptidomimetics by using binary nanoarrays of alternating gold and iron oxide particles.We show that integrin binding to the peptidomimetics is highly selective and cells spread on both ligands. However, spreading is faster and cell area and intracellular forces are increased on the α5β1 mimetic. Analysis of integrin clusters indicates that αvβ3 clusters are more pronounced on αvβ3 ligand and are present in cells adhering to α5β1 ligand. Quantitative analysis of adhesion plaques shows that focal adhesion size is increased in cells adhering to αvβ3 ligand. Integrin α5β1clusters are present in cells adhering to α5β1 ligand and colocalize with αvβ3 clusters. There are numerous fibrillar structures in cells adhering to α5β1 ligand, while clusters are mostly localized at the cell margins in cells adhering to αvβ3 ligand.These findings suggest that the activation by ligand binding of αvβ3 integrin doesn't play an active role in promoting intial adhesion and spreading, but it is essential for the formation of stable focal adhesions.

Intracellular Microrheology in the Presence of Myosin-Generated Forces in Living Cells

The mechanics of cells are governed by a network of cytoskeletal filaments and molecular motor forming a dynamic mechanical entity. A recent experimental study by Mizuno showed local shear modulus of a synthesized cytoskeletal network could increase in the presence of myosin-generated internal stresses. It was speculated whether similar behaviors could also take place in cells. To examine this, we used an experimental method similar to Mizuno's that combined active and passive microrheology to measure myosin-generated fluctuating force and the local intracellular stiffness. Our data obtained from measurements in 15 HeLa cells showed a linear relationship between the magnitude of the fluctuating forces and the time-averaged shear modulus. Moreover, when myosin activities were inhibited by ML7, we saw a dramatic decrease in the intracellular forces, but surprisingly no significant changes were observed in the time-averaged intracellular stiffness by ML7. While we did not see a direct correlation between the mean intracellular shear modulus and the motor-generated fluctuating force, we did observe an increase in the fluctuation of the shear modulus with increasing motor forces.While it is not clear why results from the intracellular study were so different from the actin construct, some differences between the two systems may be attributed to the difference. First, the intracellular fluctuating forces were found to be weaker than that in the actin construct, thus perhaps not strong enough to modulate the cell stiffness. Secondly, the steady-state internal tension, which could dictate mechanical proprieties of a network, may be different between the two systems. Unfortunately, the experimental method we used could only measure fluctuating forces but not the steady-state tension in the system. Further studies that measure both fluctuating and steady-state forces at the same time may be required to address this problem.

Three-Dimensional Modeling of Active Migration of Living Cells in a Microchannel

Leukocytes and other living cells can deform and migrate actively toward the source of chemoattractant. This directed movement of cells, known as chemotaxis, plays a crucial role in both innate and adaptive immune response of the body, and cancer metastasis. The cell's active deformation and migration are driven by intracellular forces generated by polymerization/depolymerization of cytoskeletal filaments. To numerically study these processes, we have implemented the Dembo's reactive interpenetrating flow formalism and polymerization force model into our three-dimensional volume-of-fluid (VOF)-based computational algorithm for viscoelastic cell deformation. The resulting computational model takes into account passive mechanical properties of the cell, extracellular diffusion of chemoattractant molecules, intracellular diffusion of signaling molecules, intracellular active force generation, and physiological shear flow conditions. We have applied this algorithm to study chemotaxis of nonadherent leukocytes in a rectangular microchannel in the presence or absence of shear flow. The numerical data have been tested against the corresponding experimental data.

Regulation of Nuclear Shape and Function with Cell Elongation

Many cellular events require dynamic changes in the shape and structural organization of the cell morphology and presumably affect gene expression by changing the nuclear shape. In this work, we investigate the effect of cell shape on the nucleus and the mechanism by which intracellular forces are transmitted to the nucleus. We analyzed the intracellular reorganization of individual endothelial cells plated on micropatterned substrates, imposing cells to spread on various aspect ratios. Specific drugs were used to alter each component of the cytoskeleton and we observed the spatial reorganization of the actin network, microtubules, intermediate filaments and focal adhesions, as well as the nuclear shape. Our data demonstrate the key role of the actin cytoskeleton in the adaptation of nuclear shape with cell elongation. Indeed, we show that the nucleus is subjected to pincer forces generated within the cytoskeleton via actin stress fibers. These intracellular forces drastically affect the nuclear shape and decrease the nuclear volume by 40-50% before attaining a state that is highly resistant to further deformation. Based on the quantification of cell traction forces by traction force microscopy, we propose a mechanical model that accounts for our observation and quantitatively predicts the nuclear shape. Our work also demonstrates that nucleus adaptation to cell elongation leads to a modification of nuclear functions. Indeed, DNA staining reveals an increase in chromatin condensation in highly deformed and compressed nucleus. We show that nuclear deformation in response of cellular elongation results in a strong decrease of cell ability to enter S phase and thus to proliferate. In conclusion, our results demonstrate that the shape of the cell is transposed by the actin cytoskeleton to the nucleus and suggest that it can alter the accessibility of genes to the transcription machinery.

Polymorphism and bistability in adherent cells

The optimal shapes attained by contractile cells on adhesive substrates are determined by the interplay between intracellular forces and adhesion with the extracellular matrix. We model the cell as a contractile film bounded by an elastic cortex and connected to the substrate via elastic links. When the adhesion sites are continuously distributed, optimal cell shape is constrained by the adhesion geometry, with a spread area sensitively dependent on the substrate stiffness and contractile tension. For discrete adhesion sites, equilibrium cell shape is convex at weak contractility, while developing local concavities at intermediate values of contractility. Increasing contractility beyond a critical value, controlled by mechanical and geometrical properties of adhesion, cell boundary undergoes a discontinuous transition to a star-shaped configuration with cusps and protrusions, accompanied by a region of bistability and hysteresis.

Force-Induced In Vitro Cancer Metastasis

Cancer deaths are primarily caused by metastases, not by the parent tumor. During metastasis, malignant cells detach from the parent tumor, and spread through the circulatory system to invade new tissues. The physical-chemical mechanisms and parameters within the cellular microenvironment that initiate the onset of metastasis, however, are not understood. Such a gap in the understanding and prediction of the onset of metastasis is of particular concern in colon cancer, since at the time of diagnosis, most colon cancer patients already have advanced disease. In this talk, we will present our recent discoveries that human colon carcinoma (HCT-8, HCT116 and SW480) cells can be induced to undergo a phenotypic, genetic and proteomic transition similar to the early stage of metastasis simply by being grown on a substrate with appropriate mechanical stiffness. This force-induced transition is observed as a change from a flattened, epithelial to a rounded, detached morphology, which we have termed an “E-to-R transition”. The post-transition cancer cells express a remarkable number of in vivo biophysical and biochemical metastasis hallmarks, such as loss of cell-cell and cell-substrate adhesion, reduction of cell-substrate traction force and cell-cell tugging force, gain of anchorage-independence capability, decrease of cell elasticity, and increase of cell migratory behaviors, etc. Furthermore, the post-transition cancer cells are verified to be significantly more invasive than the original cancer cells that were never exposed to soft substrates. Our novel finding suggests that the onset of metastasis may, in part, be linked to the intracellular forces and the mechanical, biophysical microenvironment of the parent tumor.

Spatial and Temporal Sensing Limits of Microtubule Polarization in Neuronal Growth Cones by Intracellular Gradients and Forces

Neuronal growth cones are the most sensitive among eukaryotic cells in responding to directional chemical cues. Although a dynamic microtubule cytoskeleton has been shown to be essential for growth-cone turning, the precise nature of coupling of the spatial cue with microtubule polarization is less understood. Here we present a computational model of microtubule polarization in a turning neuronal growth cone. We explore the limits of directional cues in modifying the spatial polarization of microtubules by testing the role of microtubule dynamics, gradients of regulators, and retrograde forces along filopodia. We analyze the steady state and transition behavior of microtubules on being presented with a directional stimulus. Our model makes novel, to our knowledge, predictions about the minimal angular spread of the chemical signal at the growth cone and the fastest polarization times. A regulatory reaction-diffusion network based on the cyclic phosphorylation-dephosphorylation of a regulator predicts that the receptor-signal magnitude can generate the maximal polarization of microtubules and not feedback loops or amplifications in the network. Using both the phenomenological and network models, we have demonstrated some of the physical limits within which the microtubule polarization system works in turning the neuron.

α-Tubulin acetylation from the inside out

Posttranslational modification of proteins, including phosphorylation, ubiquitylation, and acetylation, is an important means of signaling and functional specialization in the cell. The acetylation of lysine residues is best understood in histone proteins, in which histone acetyltransferases modify histone tail sequences to regulate chromatin structure. However, lysine acetylation has recently become more appreciated as a widely distributed phenomenon regulating diverse proteins and cellular systems (1). One such system is the microtubule (MT) cytoskeleton. MTs are highly dynamic tubular polymers assembled from protofilaments of α/β-tubulin dimers, and are essential for intracellular transport, architectural organization, and force production in eukaryotic cells (2). Acetylation of the conserved lysine 40 (K40) residue of α-tubulin, first reported more than 25 y ago (3), is one of several conserved posttranslational modifications found in the tubulin protein (2, 3). Although MTs generally function as highly dynamic polymers, α-tubulin K40 acetylation is associated with unusually stable MTs. These act as tracks for organelle and macromolecule transport in neurons, and form the structural scaffolds, called axonemes, at the core of beating cilia and flagella (4, 5). α-Tubulin K40 acetylation is catalyzed by a conserved α-tubulin acetyltransferase (α-TAT), but neither the detailed mechanism of tubulin acetylation nor its downstream functional consequences are well understood (6, 7). Now, in parallel studies presented in PNAS, Friedmann et al. (8) and Taschner et al. (9) describe the 3D structure of human α-TAT and the chemical basis for α-tubulin K40 acetylation. The two studies show how α-TAT selectively acetylates α-tubulin K40 and explore α-TAT’s catalytic mechanism through detailed enzymatic analysis (8, 9).

The Integrin-Ligand Interaction Regulates Adhesion and Migration through a Molecular Clutch

Adhesive and migratory behavior can be cell type, integrin, and substrate dependent. We have compared integrin and substrate differences using three integrin receptors: α5β1, α6β1, and αLβ2 expressed in a common cell type, CHO.B2 cells, which lack integrin α subunits, as well as in different cell types that express one or more of these integrins. We find that CHO.B2 cells expressing either α6β1 or αLβ2 integrins migrate and protrude faster and are more directionally persistent on laminin or ICAM-1, respectively, than CHO.B2 cells expressing α5β1 on fibronectin. Despite rapid adhesion maturation and the presence of large adhesions in both the α6β1- and αLβ2-expressing cells, they display robust tyrosine phosphorylation. In addition, whereas myosin II regulates adhesion maturation and turnover, protrusion rates, and polarity in cells migrating on fibronectin, surprisingly, it does not have comparable effects in cells expressing α6β1 or αLβ2. This apparent difference in the integration of myosin II activity, adhesion, and migration arises from alterations in the ligand–integrin–actin linkage (molecular clutch). The elongated adhesions in the protrusions of the α6β1-expressing cells on laminin or the αLβ2-expressing cells on ICAM-1 display a novel, rapid retrograde flux of integrin; this was largely absent in the large adhesions in protrusions of α5β1-expressing cells on fibronectin. Furthermore, the force these adhesions exert on the substrate in protrusive regions is reduced compared to similar regions in α5-expressing cells, and the adhesion strength is reduced. This suggests that intracellular forces are not efficiently transferred from actomyosin to the substratum due to altered adhesion strength, that is, avidity, affinity, or the ligand-integrin-actin interaction. Finally, we show that the migration of fast migrating leukocytes on fibronectin or ICAM-1 is also largely independent of myosin II; however, their adhesions are small and do not show retrograde fluxing suggesting other intrinsic factors determine their migration differences.

Development of a novel liquid crystal based cell traction force transducer system

Keratinocyte traction forces play a crucial role in wound healing. The aim of this study was to develop a novel cell traction force (CTF) transducer system based on cholesteryl ester liquid crystals (LC). Keratinocytes cultured on LC induced linear and isolated deformation lines in the LC surface. As suggested by the fluorescence staining, the deformation lines appeared to correlate with the forces generated by the contraction of circumferential actin filaments which were transmitted to the LC surface via the focal adhesions. Due to the linear viscoelastic behavior of the LC, Hooke's equation was used to quantify the CTFs by associating Young's modulus of LC to the cell induced stresses and biaxial strain in forming the LC deformation. Young's modulus of the LC was profiled by using spherical indentation and determined at approximately 87.1±17.2kPa. A new technique involving cytochalasin-B treatment was used to disrupt the intracellular force generating actin fibers, and consequently the biaxial strain in the LC induced by the cells was determined. Due to the improved sensitivity and spatial resolution (∼1μm) of the LC based CTF transducer, a wide range of CTFs was determined (10–120nN). These were found to be linearly proportional to the length of the deformations. The linear relationship of CTF-deformations was then applied in a bespoke CTF mapping software to estimate CTFs and to map CTF fields. The generated CTF map highlighted distinct distributions and different magnitude of CTFs were revealed for polarized and non-polarized keratinocytes.

Roles of cell confluency and fluid shear in 3-dimensional intracellular forces in endothelial cells

We use a novel 3D inter-/intracellular force microscopy technique based on 3D traction force microscopy to measure the cell-cell junctional and intracellular tensions in subconfluent and confluent vascular endothelial cell (EC) monolayers under static and shear flow conditions. We found that z-direction cell-cell junctional tensions are higher in confluent EC monolayers than those in subconfluent ECs, which cannot be revealed in the previous 2D methods. Under static conditions, subconfluent cells are under spatially non-uniform tensions, whereas cells in confluent monolayers are under uniform tensions. The shear modulations of EC cytoskeletal remodeling, extracellular matrix (ECM) adhesions, and cell-cell junctions lead to significant changes in intracellular tensions. When a confluent monolayer is subjected to flow shear stresses with a high forward component comparable to that seen in the straight part of the arterial system, the intracellular and junction tensions preferentially increase along the flow direction over time, which may be related to the relocation of adherens junction proteins. The increases in intracellular tensions are shown to be a result of chemo-mechanical responses of the ECs under flow shear rather than a direct result of mechanical loading. In contrast, the intracellular tensions do not show a preferential orientation under oscillatory flow with a very low mean shear. These differences in the directionality and magnitude of intracellular tensions may modulate translation and transcription of ECs under different flow patterns, thus affecting their susceptibility for atherogenesis.

Force communication in multicellular tissues addressed by laser nanosurgery

Cell contractility is a prominent mechanism driving multicellular tissue development and remodeling. Forces originated by the actomyosin cytoskeleton not only act within the cell body but can also propagate many layers away from the contraction source and grant tissues the ability to organize collectively and to achieve robust remodeling through development. Tissue tension is being thoroughly investigated in model organisms and increasing evidence is revealing the major role played by the communication, dynamics and propagation of cell-to-cell physical forces in multicellular remodeling. Recently, pulsed-laser-based surgery has fostered in vivo experimental studies to investigate intracellular and supracellular forces in action. The technique offers a unique method to perturb mechanical equilibrium in a subpopulation of cells or in a single cell, while the overall tissue remains intact. In particular, improved ablation precision with short laser pulses and the combination of this technique with biophysical models now allow an in-depth understanding of the role of cellular mechanics in tissue morphogenesis. We first characterize laser ablation modes available to perform intracellular, cellular, or multi-cellular ablation via the example of the model monolayer tissue of the amnioserosa of Drosophila by relating subnanosecond laser pulse energy to ablation efficiency and the probability of cavitation bubble formation. We then review recent laser nanosurgery experiments that have been performed in cultured cells and that tackle actomyosin mechanics and provide molecular insights into force-sensing mechanisms. We finally review studies showing the central role of laser ablation in revealing the nature and orientation of forces involved in intracellular contractility and force mechanosensing in tissue development, e.g., axis elongation, branching morphogenesis, or tissue invagination. We discuss the perspectives offered by the technique in force-based cell-cell communication and mechanosensing pathways.

The influence of FAs complexity and architectures of cytoskeleton on morphology during cell initial spreading

The cytoskeleton controls intracellular force equilibrium and force transmission from attachment substrate to focal adhesions (FAs). During epithelial‐mesenchymal transition (EMT), essential process for numerous developmental processes, cell morphology has dramatically change by alter force balance between extracellular matrix (ECM) and cytoskeletons. The force and tension pattern change in cytoskeleton in relation to different morphology is still unknown. Our objective is to simulate the dynamics of intracellular force distribution and total stored energy during cell spreading by using the cytoskeleton tensegrity model with the pre‐tensed actin filaments and the pre‐compressed microtubules. The Madin‐Darby Canine Kidney (MDCK) cell, bovine aortic endothelium cell (BAEC), and myoblastoma cell (C2C12) and NIH 3T3 fibroblast (NIH 3T3) represents each step of EMT. The coordination of FA was used for stimulation. Our results indicated that most of all MDCK and C2C12 is circle shape as compared to BAEC and NIH 3T3 which is polygon. FAs in C2C12 but not BAEC and MDCK fuse together during cell spreading. The total energy in stress fiber and microtubules will increase and decrease with spreading area respectively. Cells have different FA distribution and stress fiber arrangement present different cell morphology due to energy distribution. This study was supported by NSC in Taiwan (NSC 98‐2627‐B‐006‐010).

Asymmetric Mode of Ca2+-S100A4 Interaction with Nonmuscle Myosin IIA Generates Nanomolar Affinity Required for Filament Remodeling

SummaryFilament assembly of nonmuscle myosin IIA (NMIIA) is selectively regulated by the small Ca2+-binding protein, S100A4, which causes enhanced cell migration and metastasis in certain cancers. Our NMR structure shows that an S100A4 dimer binds to a single myosin heavy chain in an asymmetrical configuration. NMIIA in the complex forms a continuous helix that stretches across the surface of S100A4 and engages the Ca2+-dependent binding sites of each subunit in the dimer. Synergy between these sites leads to a very tight association (KD ∼1 nM) that is unique in the S100 family. Single-residue mutations that remove this synergy weaken binding and ameliorate the effects of S100A4 on NMIIA filament assembly and cell spreading in A431 human epithelial carcinoma cells. We propose a model for NMIIA filament disassembly by S100A4 in which initial binding to the unstructured NMIIA tail initiates unzipping of the coiled coil and disruption of filament packing.

Numerically bridging lamellipodial and filopodial activity during cell spreading reveals a potentially novel trigger of focal adhesion maturation

We present a novel approach to modeling cell spreading, and use it to reveal a potentially central mechanism regulating focal adhesion maturation in various cell phenotypes. Actin bundles that span neighboring focal complexes at the lamellipodium-lamellum interface were assumed to be loaded by intracellular forces in proportion to bundle length. We hypothesized that the length of an actin bundle (with the corresponding accumulated force at its adhesions) may thus regulate adhesion maturation to ensure cell mechanical stability and morphological integrity. We developed a model to test this hypothesis, implementing a "top-down" approach to simplify certain cellular processes while explicitly incorporating complexity of other key subcellular mechanisms. Filopodial and lamellipodial activities were treated as modular processes with functional spatiotemporal interactions coordinated by rules regarding focal adhesion turnover and actin bundle dynamics. This theoretical framework was able to robustly predict temporal evolution of cell area and cytoskeletal organization as reported from a wide range of cell spreading experiments using micropatterned substrates. We conclude that a geometric/temporal modeling framework can capture the key functional aspects of the rapid spreading phase and resultant cytoskeletal complexity. Hence the model is used to reveal mechanistic insight into basic cell behavior essential for spreading. It demonstrates that actin bundles spanning nascent focal adhesions such that they are aligned to the leading edge may accumulate centripetal endogenous forces along their length, and could thus trigger focal adhesion maturation in a force-length dependent fashion. We suggest that this mechanism could be a central "integrating" factor that effectively coordinates force-mediated adhesion maturation at the lamellipodium-lamellum interface.