Biology subjects

Hockenberry, M. A.

Publications and source records attributed to Hockenberry, M. A..

5 recordsLinked to original sources

bioRxiv2026

Nuclear SUN2 coordinates endothelial cell-matrix interactions to regulate blood vessel homeostasis and barrier function

Vascular endothelial cells respond to environmental forces to remodel vessels during development and to achieve homeostasis, and mis-regulated responses lead to vascular dysfunction and disease. The nucleus participates in force transduction to cell-matrix junctions via the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex that resides in the nuclear envelope, but how these forces are regulated and relayed is incompletely understood. We found that the LINC complex protein SUN2 is required for proper endothelial cell-matrix interactions that occur far from the nucleus and affect angiogenic expansion, vascular responses to flow, and barrier integrity. Endothelial cells lacking SUN2 had inappropriate flow responses and reduced expression of flow-mediated transcription factors in vitro and in vivo. Expression of several matrix and adhesion genes was reduced in SUN2-depleted cells, leading to defective extracellular matrix, dysmorphic focal adhesions resistant to dynamic turnover, and disturbed cell-matrix force distribution. Mechanistically, nuclear SUN2 affected dynamic regulation of the microtubule cytoskeleton that correlated with matrix metalloprotease-dependent barrier dysfunction. These findings indicate that nuclear SUN2 establishes and maintains blood vessel homeostasis by controlling microtubule-mediated effects on focal adhesion turnover and extracellular matrix properties, with implications for cardiovascular aging and diseases such as Marfan syndrome that affect vessel wall integrity.

cell biology↗

bioRxiv2026

Quantitative analysis of fibroblast migration reveals migratory states characterized by force generation, cell shape and motion

Cell migration depends on coordinating cell shape changes with force generation, yet how these processes are integrated remains unclear. Here, we combine live-cell imaging with traction force microscopy and computational analysis to quantify cell morphology, motility and force generation in migrating fibroblasts. We find that traction force magnitudes display a multimodal distribution, suggesting discrete migratory regimes. Using a Hidden Markov Model, we identify distinct force states that exhibit differences in shape and motion metrics, and show that individual cells transition between force states over time. To test the role of cytoskeletal organization in establishing the identified states, we analyzed cells lacking Arpc2, which disrupts branched actin assembly. Despite reduced forces and altered morphology, these cells also exhibit three migratory states. State transitions occur more frequently in cells lacking Arpc2 and unlike normal cells their protrusion geometry is force dependent. Together, our findings show that cell migration is organized into discrete mechanical states that couple morphology, motility and force generation. SUMMARY STATEMENTFibroblast motility involves distinct migratory states. These states exist independent of branched actin. However, state transition frequencies, traction force magnitudes and protrusion geometry are branched actin dependent.

cell biology↗

bioRxiv2026

Actin-membrane interface stress regulates Arp2/3-branched actin density during lamellipodial protrusion

Motile cells can sense and exert forces on the extracellular environment through dynamic actin networks. Increased stress against the polymerizing barbed ends of branched actin networks has been shown to lead to an increase in the density of these networks through a force feedback mechanism, though this phenomenon has not been explored through the examination of real-time responses of endogenous branched actin in cells. Here, we utilize mouse CRISPR knock-in fibroblasts with labeled Arp2/3 complex to identify cellular and extracellular conditions that regulate branched actin density and enrichment at the leading edge of lamellipodial protrusions. A common theme shared among all branched actin density-increasing conditions is higher levels of predicted interface stress between the plasma membrane and the barbed ends of the lamellipodial actin network. Among these conditions, we find that Arp2/3 is specifically required for robust spreading and protrusion in response to increased extracellular viscosity. Interestingly, time-lapse traction force microscopy of Arp2/3-dependent viscosity responses show significantly reduced changes in strain energy applied to the substrate when compared to spreading and motility utilizing cell-matrix adhesion. In addition, we find that increased extracellular viscosity can bypass the need for extracellular matrix proteins to support lamellipodial protrusion driven by optogenetic Rac activation. Our studies provide strong support for in vitro models of branched actin force feedback responses and further characterize an essential role for branched actin in mediating dramatic cell shape changes in response to increased extracellular viscosity.

cell biology↗

bioRxiv2024

Measurement of cellular traction forces during confined migration

To migrate efficiently through tissues, cells must transit through small constrictions within the extracellular matrix. However, in vivo environments are geometrically, mechanically, and chemically complex, and it has been difficult to understand how each of these parameters contribute to the propulsive strategy utilized by cells in these diverse settings. To address this, we employed a sacrificial micromolding approach to generate polymer substrates with tunable stiffness, controlled adhesivity, and user-defined microscale geometries. We combined this together with live-cell imaging and three-dimensional traction force microscopy (TFM) to quantify the forces that cells use to transit through constricting channels. Surprisingly, we observe that cells migrating through compliant constrictions take longer to transit and experience greater nuclear deformation than those migrating through more rigid constrictions. TFM reveals that this deformation is generated by inwardly directed contractile forces that decrease the size of the opening and pull the walls closed around the nucleus. These findings show that nuclear deformation during confined migration can be accomplished by internal cytoskeletal machinery rather than by reactive forces from the substrate, and our approach provides a mechanism to test between different models for how cells translocate their nucleus through narrow constrictions. The methods, analysis, and results presented here will be useful to understand how cells choose between propulsive strategies in different physical environments. Significance StatementCell migration is critical for both physiological events like wound healing and pathological events like metastasis. Understanding how cells move through complex environments will assist efforts to enhance or inhibit such processes. We developed a method to quantify the forces that cells use to move through multidimensional environments, including through narrow constrictions like those in tissues. Surprisingly, we find that cells transiting through soft constrictions take longer and deform more than those transiting through rigid constrictions, and we connect this finding to inwardly directed contractile forces generated by migrating cells. Together, this work reveals a key role for substrate rigidity to regulate cell transit through confining geometries and provides a quantitative platform to investigate similar processes in other settings.

biophysics↗

bioRxiv2022

Zyxin and non-muscle myosin are required for single fibroblast durotaxis, but Rho-kinase activity and the Arp2/3 complex are dispensable

Durotaxis, migration of cells directed by stiffness gradient, is critical in development and disease. To study the molecular determinants of single cell durotaxis, we developed an all-in-one photopolymerized hydrogel system containing areas of stiffness gradients with different slopes, along with uniform stiffness (soft and stiff) regions. We find that fibroblasts rely on non-muscle myosin II (NMII) activity and the LIM-domain protein zyxin for durotaxis on both steep and shallow stiffness gradients. Importantly, unlike haptotaxis, the Arp2/3 complex is dispensable for durotaxis on both stiffness gradients. Lack of Arp2/3 results in a filopodia-based durotactic migration that is equally efficient as that of lamellipodia-based durotactic migration. Finally, we reveal an essential role for the actin-bundler fascin in the formation and asymmetric distribution of filopodia during filopodia-based durotaxis in shallow, but not steep, stiffness gradient. Together, our all-in-one hydrogel system can serve as a platform to identify, discriminate, and characterize stiffness gradient specific molecular mechanisms that cells employ to efficiently durotax.

cell biology↗