Changes In Cytoskeletal Structure Research Articles

The exact phenomenon and early signaling events of the endothelial cytoskeleton response to ultrasound

Low-intensity ultrasound can be applied for medical imaging and disease treatment in clinical and experimental studies. However, the biological effects of ultrasound on blood vessels, especially endothelial cells (ECs) are still unclear. In this study, the laws of endothelial cytoskeleton changes under ultrasound induction are investigated. ECs are exposed to low-intensity ultrasound, and the cytoskeletal morphology is analyzed by a filamentous (F)-actin staining technique. We further analyze the characteristics of cytoskeleton rupture using indirect immunofluorescence techniques and cytoskeleton electron microscopy. Finally, the biological effects induced by ultrasound at the tissue level are investigated in an ex vivo blood-vessel model. Significant changes in cytoskeletal structure are detected when induced by ultrasound, including cytoskeletal rupture, blebbing and apoptosis. Moreover, a temporal threshold of ECs injury under different ultrasonic intensities is established. This study illustrates a pattern of significant changes in the cytoskeletal structure of ECs induced by ultrasound. The finding serves as a guide for selecting a safe threshold for clinical ultrasound applications.

Crystal structure of a guanine nucleotide exchange factor encoded by the scrub typhus pathogen Orientia tsutsugamushi

Rho family GTPases regulate an array of cellular processes and are often modulated by pathogens to promote infection. Here, we identify a cryptic guanine nucleotide exchange factor (GEF) domain in the OtDUB protein encoded by the pathogenic bacterium Orientia tsutsugamushi A proteomics-based OtDUB interaction screen identified numerous potential host interactors, including the Rho GTPases Rac1 and Cdc42. We discovered a domain in OtDUB with Rac1/Cdc42 GEF activity (OtDUBGEF), with higher activity toward Rac1 in vitro. While this GEF bears no obvious sequence similarity to known GEFs, crystal structures of OtDUBGEF alone (3.0 Å) and complexed with Rac1 (1.7 Å) reveal striking convergent evolution, with a unique topology, on a V-shaped bacterial GEF fold shared with other bacterial GEF domains. Structure-guided mutational analyses identified residues critical for activity and a mechanism for nucleotide displacement. Ectopic expression of OtDUB activates Rac1 preferentially in cells, and expression of the OtDUBGEF alone alters cell morphology. Cumulatively, this work reveals a bacterial GEF within the multifunctional OtDUB that co-opts host Rac1 signaling to induce changes in cytoskeletal structure.

Heavy Metal Exposure Leads to Rapid Changes in Cellular Biophysical Properties.

Biophysical properties of cells, such as cell mechanics, cell shape, and cell migration, are emerging hallmarks for characterizing various cell functions. Conversely, disruptions to these biophysical properties may be used as reliable indicators of disruptions to cell homeostasis, such as in the case of chemical-induced toxicity. In this study, we demonstrate that treatment of lead(II) nitrate and cadmium nitrate leads to dosage-dependent changes in a collection of biophysical properties, including cellular traction forces, focal adhesions, mechanical stiffness, cell shape, migration speed, permeability, and wound-healing efficacy in mammalian cells. As those changes appear within a few hours after the treatment with a trace amount of lead/cadmium, our results highlight the promise of using biophysical properties to screen environmental chemicals to identify potential toxicants and establish dose response curves. Our systematic and quantitative characterization of the rapid changes in cytoskeletal structure and cell functions upon heavy metal treatment may inspire new research on the mechanisms of toxicity.

MicroRNAs Contribute to Breast Cancer Invasiveness.

Cancer statistics in 2018 highlight an 8.6 million incidence in female cancers, and 4.2 million cancer deaths globally. Moreover, breast cancer is the most frequent malignancy in females and twenty percent of these develop metastasis. This provides only a small chance for successful therapy, and identification of new molecular markers for the diagnosis and prognostic prediction of metastatic disease and development of innovative therapeutic molecules are therefore urgently required. Differentially expressed microRNAs (miRNAs) in cancers cause multiple changes in the expression of the tumorigenesis-promoting genes which have mostly been investigated in breast cancers. Herein, we summarize recent data on breast cancer-specific miRNA expression profiles and their participation in regulating invasive processes, in association with changes in cytoskeletal structure, cell-cell adhesion junctions, cancer cell-extracellular matrix interactions, tumor microenvironments, epithelial-to-mesenchymal transitions and cancer cell stem abilities. We then focused on the epigenetic regulation of individual miRNAs and their modified interactions with other regulatory genes, and reviewed the function of miRNA isoforms and exosome-mediated miRNA transfer in cancer invasiveness. Although research into miRNA’s function in cancer is still ongoing, results herein contribute to improved metastatic cancer management.

The regulator of G-protein signalling Thn1 links pheromone response to volatile production in Schizophyllum commune.

The regulator of G-protein signalling, Thn1, is involved in sexual development through pheromone signalling in the mushroom forming basidiomycete Schizophyllum commune affecting hyphal morphology and mating interactions. Thn1 plays a key role in coordinating sesquiterpene production, pheromone response and sexual development. The gene thn1 is transcriptionally regulated in response to mating with a role in clamp cell development and hydrophobin gene transcription. Further, it negatively regulates cAMP signalling and secondary metabolism. Disruption of thn1 affects dikaryotization by reducing clamp fusion and development with predominant non-fused pseudoclamps. Enhanced protein kinase A (PKA) activities in Δthn1 strains indicate that Thn1 regulates pheromone signalling by de-activating G-protein α subunits, which control cAMP-dependent PKA. The repressed formation of aerial hyphae could be linked to a reduced metabolic activity and to a transcriptional down-regulation of hyd6 and sc3 hydrophobin genes. Thn1 was also shown to be necessary for the biosynthesis of sesquiterpenes and an altered spectrum of sesquiterpenes in Δthn1 is linked to transcriptional up-regulation of biosynthesis genes. Proteome analysis indicated changes in cytoskeletal structure affecting actin localization, linking the major regulator Thn1 to growth and development of S. commune. The results support a role for Thn1 in G-protein signalling connecting development and secondary metabolism.

Mechanical load increase-induced changes in cytoskeletal structure and cellular barrier function in human cerebral endothelial cells.

Globally, approximately a billion patients are estimated to suffer from neurological disorders. Although there are many therapeutic candidates for the central nervous system, treatment of brain disorders is restricted by the blood-brain barrier (BBB), which is a highly selective membrane that protects the brain from exogenous substances. This study was undertaken to develop a novel strategy to overcome the BBB and improve the efficiency of drug delivery to the brain by mechanical load increase using hypergravity. Human cerebral microvascular endothelial cells were exposed three times to 20 min hypergravity (10g), with a 20-min rest period between each exposure. The applied hypergravity reversibly decreased the cellular metabolic activity and increased the permeation rate of fluorescein sodium salt, fluorescein isothiocyanate-labeled dextran (FD-4), and fluorescein-labeled jacalin. Following the exposure to hypergravity, we also observed structural changes of the cytoskeleton and tight junctions, and an alteration in the expression levels of related genes. These results indicate that increased mechanical load due to the applied hypergravity affects the cytoskeletal arrangement and tight junctions, thereby weakening the cell barrier function and enhancing the permeability of the paracellular pathway. Thus, the mechanical load increase by hypergravity has the potential of being used as a novel strategy to overcome the BBB for brain drug delivery.

A bifurcated signaling cascade of NIMA-related kinases controls distinct kinesins in anaphase.

In mitosis, cells undergo a precisely orchestrated series of spatiotemporal changes in cytoskeletal structure to divide their genetic material. These changes are coordinated by a sophisticated network of protein-protein interactions and posttranslational modifications. In this study, we report a bifurcation in a signaling cascade of the NIMA-related kinases (Neks) Nek6, Nek7, and Nek9 that is required for the localization and function of two kinesins essential for cytokinesis, Mklp2 and Kif14. We demonstrate that a Nek9, Nek6, and Mklp2 signaling module controls the timely localization and bundling activity of Mklp2 at the anaphase central spindle. We further show that a separate Nek9, Nek7, and Kif14 signaling module is required for the recruitment of the Rho-interacting kinase citron to the anaphase midzone. Our findings uncover an anaphase-specific function for these effector kinesins that is controlled by specific Nek kinase signaling modules to properly coordinate cytokinesis.

Proteomic and Immunohistological Characterization of Cytoskeletal Alterations during the Progression of Human Cardiomyopathy

Background: Intermediate filaments (IFs) and microtubules (MTs) play diverse structural and functional roles in the cardiomyocyte, including newly defined regulatory roles in cardiac mechanics and mechanosignaling. Defects in cytoskeleton may contribute to cardiomyopathies through a combination of structural and signaling mechanisms. Previously, we demonstrated that detyrosination of α-tubulin increases MT-IF interaction to regulate the stiffness of cardiomyocytes and impede contractility. Levels of detyrosination are inversely correlated with left ventricular ejection fraction in patients with heart disease. Aim: To broadly characterize cytoskeletal alterations in diverse human cardiac disease. To this end we performed immunohistological and proteomic analysis of left ventricle tissue samples from patients with heart disease of varying origin and severity. Results: Our analysis reveals alterations in a range of cytoskeletal proteins, including different tubulin isotypes, microtubule associated proteins (MAPs), IFs, and sarcomeric proteins. Among these, IFs (such as desmin and synemin) and MAP4 are strikingly and progressively upregulated in hypertrophy and hypertrophic cardiomyopathy, as well as dilated and ischemic cardiomyopathy. This suggests a broadly conserved upregulation and stabilization of IFs and MTs during the progression of cardiomyopathy. Protein expression changes were confirmed by western blot, and quantitative immunofluorescence was used to characterize the density and distribution of cytoskeletal networks in diseased myocardium. Through these studies we aim to provide the most complete characterization to date of cytoskeletal changes associated with human heart disease, in an effort to better understand how pathological changes in cytoskeletal structure correlate with declining function.

Distinct localizations and roles of non‐muscle myosin II during proplatelet formation and platelet release

At the end of maturation, megakaryocytes (MKs) form long cytoplasmic extensions called proplatelets (PPT). Enormous changes in cytoskeletal structures cause PPT to extend further, to re-localize organelles such as mitochondria and to fragment, leading to platelet release. Two non-muscle myosin IIs (NMIIs) are expressed in MKs; however, only NMII-A (MYH9), but not NMII-B (MYH10), is expressed in mature MKs and is implicated in PPT formation. To provide invivo evidence on the specific role of NMII-A and IIB in MK PPT formation. We studied two transgenic mouse models in which non-muscle myosin heavy chain (NMHC) II-A was genetically replaced either by II-B or by a chimeric NMHCII that combined the head domain of II-A with the rod and tail domains of II-B. This work demonstrates that the kinetic properties of NM-IIA, depending on the N-terminal domain, render NMII-A the better NMII candidate to control PPT formation. Furthermore, the carboxyl-terminal domain determines myosin II localization in the constriction region of PPT and is responsible for the specific role of NMII in platelet release.

The Dynamic Interplay between Cleavage Furrow Proteins in Cellular Mechanoresponsiveness

Cell shape changes associated with processes like cytokinesis and motility proceed on several second time-scales. However, they are derived from molecular events occurring much faster, including protein-protein interactions, filament assembly, and force generation. How these fast dynamics define cellular outcomes remain unknown. While accumulation of cytoskeletal elements is often portrayed as being driven by signaling pathways, mechanical stresses also direct proteins. A myosin II-based mechanosensory system controls cellular contractility and shape during cytokinesis and under applied stress. In Dictyostelium, this system tunes myosin II accumulation under mechanical stress by feedback through the actin network, particularly through the crosslinker cortexillin I. Cortexillin-binding IQGAP proteins are major regulators of this system. We examined the dynamic interplay between these key cytoskeletal proteins using fluorescence recovery after photobleaching and fluorescence correlation spectroscopy, defining the short time-scale dynamics of these players during cytokinesis and under mechanical stress. Actin and its polar cortex-enriched crosslinkers showed sub-second recovery, while equatorially enriched proteins including cortexillin I, IQGAP2, and myosin II recovered in 1-5 seconds. Mobility of these equatorial proteins was greatly reduced at the furrow, compared to their interphase dynamics. This mobility shift did not arise from a single biochemical event, but rather from global inhibition of protein dynamics by mechanical stress-associated changes in cytoskeletal structure. We further expanded our genetic and biochemical understanding of this mechanosensory system using a proteomics approach to identify relevant protein-protein interactions. We identified that, in addition to binding to each other, both cortexillin I and IQGAP2 also interact with myosin II under conditions that prevent myosin II-F-actin binding. This validates the high crosstalk occurring between various mechanosensitive elements. Mechanical tuning of contractile protein dynamics provides robustness to the cytoskeletal framework responsible for regulating cell shape and contributes to the fidelity of cytokinesis.

Mechanical regulation of human vocal fold stellate cells.

It is generally accepted that tensile and compressive strains have direct effects on cell morphology and structure, including changes in cytoskeletal structure and organization. Cytoskeletons play the role of mechanoreceptor of the cells. Vocal fold stellate cells (VFSCs) in the human maculae flavae (MFe) are inferred to be involved in the metabolism of extracellular matrices essential for the viscoelasticity of the vocal fold mucosa. Our previous studies have supported the hypothesis that the tension caused by phonation (vocal fold vibration) regulates the behavior of the VFSCs. The microstructure of the intermediate filaments and the expression of their proteins were investigated in VFSCs in the MFe, which had remained unphonated since birth. Three adult vocal fold mucosae that had remained unphonated since birth were investigated by immunohistochemistry and electron microscopy. The intermediate filaments of the VFSCs were fewer in number. The expression of their characteristic proteins (vimentin, desmin, and glial fibrillary acidic protein) was also reduced. Vocal fold vibration seems to affect VFSC morphology and structure, such as cytoskeletal structure and organization. This supports the hypothesis that vocal fold vibration regulates VFSC behavior in the human MFe. In addition to chemical factors, mechanical factors also appear to modulate VFSC behavior.

Cytoskeleton of newborn vocal fold stellate cells.

Vocal fold stellate cells (VFSCs) in the human maculae flavae located at both ends of the vocal fold mucosa are inferred to be involved in the metabolism of extracellular matrices of the vocal fold mucosa. Tension caused by phonation (vocal fold vibration) likely regulates the behavior of the VFSCs in the human maculae flava. Tensile and compressive strains have direct effects on cell morphology and structure, including changes in cytoskeletal structure and organization. Cytoskeletons play a role as mechanoreceptors for the cells. The microstructure of the intermediate filaments and the expression of their characteristic proteins were investigated regarding the human newborn VFSCs. Histopathologic analysis of the human newborn vocal fold. Three newborn vocal fold mucosae were investigated by immunohistochemistry and electron microscopy. The intermediate filaments in the cytoplasm of the newborn VFSCs were few in number. However, their characteristic proteins (vimentin, desmin, GFAP [Glial fibrillary acidic protein], cytokeratin) had already expressed. The function and fate of VFSCs are regulated by various microenvironmental factors. Not only chemical factors but also mechanical factors could also modulate VFSC behaviors. The cytoskeletal structure of the newborn VFSCs is under development. And the newborn VFSCs have not acquired mechanical regulation. N/A.

The Lymphocyte Story – an Overview of Selected Highlights on the in Vitro Activation of Human Lymphocytes in Space

Since the first flight of humans into space it is known that space flight affects the immune system; especially a weakening of the reactivity of T-lymphocytes after flight has been observed. In an in vitro experiment, proposed by Augusto Cogoli and flown in Spacelab-1 in 1983, the activation of T-lymphocytes was found to be strongly inhibited in microgravity. This surprising result triggered extended investigations in space and on the ground by us and other research teams. T-cells are that subpopulation of lymphocytes responsible for the activation of the specific immune system. The mechanism of T-cell activation is very complex; 3 different signals are required as well as an interaction between T-lymphocytes and monocytes. Cell motility based on a continuous rearrangement of the cytoskeletal network within the cell is essential for cell-cell contacts. The objective of all our experiments performed on different platforms in space as well as in simulated microgravity on ground was to understand and explain the dysfunction of the cell activation under reduced gravity conditions. On sounding rockets we have studied the influence of microgravity on the delivery of the first signal, the motility of lymphocytes as well as changes in the cytoskeletal structure and early gene expression. On long term missions we investigated many aspects of the delivery of the 2 nd and 3 rd signal, including motility and aggregate formation of lymphocytes, interaction of lymphocytes with monocytes, motility of monocytes and changes in different cytoskeletal structures.

Role of Airway Recruitment and Derecruitment in Lung Injury

The mechanical forces generated during the ventilation of patients with acute lung injury causes significant lung damage and inflammation. Low-volume ventilation protocols are commonly used to prevent stretch-related injury that occurs at high lung volumes. However, the cyclic closure and reopening of pulmonary airways at low lung volumes, i.e., derecruitment and recruitment, also causes significant lung damage and inflammation. In this review, we provide an overview of how biomedical engineering techniques are being used to elucidate the complex physiological and biomechanical mechanisms responsible for cellular injury during recruitment/derecruitment. We focus on the development of multiscale, multiphysics computational models of cell deformation and injury during airway reopening. These models, and the corresponding in vitro experiments, have been used to both elucidate the basic mechanisms responsible for recruitment/derecruitment injury and to develop alternative therapies that make the epithelium more resistant to injury. For example, models and experiments indicate that fluidization of the cytoskeleton is cytoprotective and that changes in cytoskeletal structure and cell mechanics can be used to mitigate the mechanotransduction of oscillatory pressure into inflammatory signaling. The continued application of biomedical engineering techniques to the problem of recruitment/derecruitment injury may therefore lead to novel and more effective therapies.

Minimal Peroxide Exposure of Neuronal Cells Induces Multifaceted Adaptive Responses

Oxidative exposure of cells occurs naturally and may be associated with cellular damage and dysfunction. Protracted low level oxidative exposure can induce accumulated cell disruption, affecting multiple cellular functions. Accumulated oxidative exposure has also been proposed as one of the potential hallmarks of the physiological/pathophysiological aging process. We investigated the multifactorial effects of long-term minimal peroxide exposure upon SH-SY5Y neural cells to understand how they respond to the continued presence of oxidative stressors. We show that minimal protracted oxidative stresses induce complex molecular and physiological alterations in cell functionality. Upon chronic exposure to minimal doses of hydrogen peroxide, SH-SY5Y cells displayed a multifactorial response to the stressor. To fully appreciate the peroxide-mediated cellular effects, we assessed these adaptive effects at the genomic, proteomic and cellular signal processing level. Combined analyses of these multiple levels of investigation revealed a complex cellular adaptive response to the protracted peroxide exposure. This adaptive response involved changes in cytoskeletal structure, energy metabolic shifts towards glycolysis and selective alterations in transmembrane receptor activity. Our analyses of the global responses to chronic stressor exposure, at multiple biological levels, revealed a viable neural phenotype in-part reminiscent of aged or damaged neural tissue. Our paradigm indicates how cellular physiology can subtly change in different contexts and potentially aid the appreciation of stress response adaptations.

Effects of micrometric titanium particles on osteoblast attachment and cytoskeleton architecture

Titanium (Ti) and its alloys are widely used in biomedical devices as bone tissue replacements due to their advantageous bulk mechanical properties and biocompatibility. It is known that particles released from Ti-based implants impair essential functions of osteoblasts, which for survival require attachment to specific extracellular matrix proteins at the bone surface. This study investigates whether Ti particles of micrometric sizes affect the osteoblast attachment machinery. Exposure of human osteoblastic Saos-2 cells to Ti particles impaired their adhesion strength, migration and proliferation. Attenuation of these functions was associated with reduced cell spreading, cell membrane disruptions and loss of cell shape. Cell exposure to Ti particles led to changes in cytoskeletal structures, including reduced ventral stress fibers combined with a disorderly arrangement of β-tubulin and acetylated α-tubulin fibers. Cytoskeleton disassembly was associated with a reduction in overall cell adhesion area, characterized by fewer centrally localized focal adhesions and shorter focal contacts at the periphery. Paxillin adaptor protein redistributed to peripheral corner regions, colocalizing with poorly organized actin fibers at attachment sites. Total focal adhesion kinase (FAK) protein amounts, as well as its degree of phosphorylation on the active form p-FAK (Tyr-397), decreased, which was accompanied by a lesser extent of co-localization with paxillin in focal contacts. On the other hand, p-FAK (Tyr-407), an inhibitory form of FAK, accumulated in the focal contacts of Ti-treated cells. Pyk2 phosphorylated on Tyr-402 colocalized with paxillin in focal contacts of untreated cells, while it was barely detected upon exposure to particles. In summary, changes in the phosphorylation states of both FAK and Pyk2 tyrosine kinases at focal contacts underlie impaired bone-forming cell attachment after exposure to Ti particles of micrometric sizes.

Estudio del Efecto de Campos Magnéticos en Citoesqueleto de Osteoblastos Humanos

En este trabajo se presentan los resultados obtenidos de la aplicación de Campos Magnéticos (CM) tanto pulsados (CMP) como estáticos (CME) en la morfología de osteoblastos humanos. El efecto de dichos campos ha sido medido por medio del análisis de la estructura de la β-tubulina, la cual es una proteína que forma parte del citoesqueleto celular. El campo magnético aplicado fue de 0.65 mT en el caso del CMP y de 0.5 mT en el caso del CME. La aplicación de los CM provoca alteración en el patrón de distribución normal de las redes de microtúbulos, dando lugar a la formación de agregados fluorescentes en la región cortical de la membrana celular. Las observaciones obtenidas con respecto a loscambios morfológicos de los osteoblastos, indican claramente que éstos son sensibles a la estimulación con CM, alterando su actividad celular a través de cambios en la estructuradel citoesqueleto celular.

Influence of cytoskeletal structure and mechanics on epithelial cell injury during cyclic airway reopening

Although patients with acute respiratory distress syndrome require mechanical ventilation, these ventilators often exacerbate the existing lung injury. For example, the cyclic closure and reopening of fluid-filled airways during ventilation can cause epithelial cell (EpC) necrosis and barrier disruption. Although much work has focused on minimizing the injurious mechanical forces generated during ventilation, an alternative approach is to make the EpC less susceptible to injury by altering the cell's intrinsic biomechanical/biostructural properties. In this study, we hypothesized that alterations in cytoskeletal structure and mechanics can be used to reduce the cell's susceptibility to injury during airway reopening. EpC were treated with jasplakinolide to stabilize actin filaments or latrunculin A to depolymerize actin and then exposed to cyclic airway reopening conditions at room temperature using a previously developed in vitro cell culture model. Actin stabilization did not affect cell viability but significantly improved cell adhesion primarily due to the development of more numerous focal adhesions. Surprisingly, actin depolymerization significantly improved both cell viability and cell adhesion but weakened focal adhesions. Optical tweezer based measurements of the EpC's micromechanical properties indicate that although latrunculin-treated cells are softer, they also have increased viscous damping properties. To further investigate the effect of "fluidization" on cell injury, experiments were also conducted at 37 degrees C. Although cells held at 37 degrees C exhibited no changes in cytoskeletal structure, they did exhibit increased viscous damping properties and improved cell viability. We conclude that fluidization of the actin cytoskeleton makes the EpC less susceptible to the injurious mechanical forces generated during cyclic airway reopening.

The expression and prognostic value of the guanine nucleotide exchange factors (GEFs) Trio, Vav1 and TIAM-1 in human breast cancer.

BackgroundDevelopment of metastasis in breast cancer is a multi-step process comprising changes in cytoskeletal structure and gene expression of tumour cells leading to changes in cell adhesion and motility. The Rho GTPase proteins, which function as guanine nucleotide regulated binary switches, govern a variety of cellular processes including cell motility and migration, changes in cell adhesion as well as actin cytoskeletal reorganisation and gene expression/transcription. One group of activators which regulate the Rho-GTPases is the guanine nucleotide exchange factors (GEFs), and this study looked at three such GEFs, Trio, Vav1 and TIAM-1. The purpose of this study was to investigate the expression of these GEFs, in human breast cancer and assess the affect on clinical outcome.MethodsSpecimens of fresh, frozen breast tumour tissue (n = 113) and normal background tissue (n = 30) were processed for quantitative PCR analysis. The expression and levels of expression of Trio, Vav1 and TIAM-1 were analysed using RT-PCR and real-time Q-PCR respectively. Sections were also immunostained with Trio and Tiam-1 antibodies.ResultsTumour tissue exhibited high levels of all three Rho activators Trio, Vav1 and TIAM-1 compared with normal background breast tissue, reaching a level of significance for the GEF Trio (p = 0.013). Trio levels also increased significantly in patients with a poor prognostic index (p = 0.04).Levels of TIAM-1 were significantly higher in tumour tissue from patients who died from breast cancer compared with those who survived (p = 0.04). No significant correlation was found between tumour grade and histology types.ConclusionHigh expression levels of Trio, Vav1 and TIAM-1 were seen in breast tumours, especially in those with poor prognosis. This suggests that aberrant regulation of Rho family activities by GEFs may have an important prognostic value in breast cancer.

Rho signaling and mechanical control of vascular development

To discuss how mechanical cues and Rho signaling contribute to control of vascular development and hematopoiesis. Rho guanine trinucleotide phosphatases are ubiquitious regulators of cytoskeletal structure and tension generation. Recent work shows that Rho-dependent mechanical interactions between cells and extracellular matrix regulate cell fate switching in capillary endothelial cells and megakaryocytes in vitro, as well as angiogenesis, vascular permeability, leukocyte migration and platelet formation in vivo. Signaling pathways that link integrins and tension-dependent changes in cytoskeletal structure to Rho have also begun to be delineated. Mechanical force generation by cells and simultaneous sensing of these physical forces play critical roles in vascular development by estimating whether individual cells will grow, differentiate, move or undergo apoptosis in the local tissue microenvironment. Future work in the vascular field therefore needs to incorporate physical control mechanisms into existing biochemical concepts of cell and tissue regulation.