The Scar/WAVE complex drives actin nucleation during cell migration. Interestingly, the same complex is important in forming membrane ruffles during macropinocytosis, a process mediating nutrient uptake and membrane receptor trafficking. Mammalian CYRI-B is a recently described negative regulator of the Scar/WAVE complex by RAC1 sequestration, but its other paralogue, CYRI-A, has not been characterized. Here, we implicate CYRI-A as a key regulator of macropinosome formation and integrin internalization. We find that CYRI-A is transiently recruited to nascent macropinosomes, dependent on PI3K and RAC1 activity. CYRI-A recruitment precedes RAB5A recruitment but follows sharply after RAC1 and actin signaling, consistent with it being a local inhibitor of actin polymerization. Depletion of both CYRI-A and -B results in enhanced surface expression of the α5β1 integrin via reduced internalization. CYRI depletion enhanced migration, invasion, and anchorage-independent growth in 3D. Thus, CYRI-A is a dynamic regulator of macropinocytosis, functioning together with CYRI-B to regulate integrin trafficking. Read More…

Liver disease represents an increasing cause of global morbidity and mortality. Currently, liver transplant is the only treatment curative for end-stage liver disease. Donor organs cannot meet the demand and therefore scalable treatments and new disease models are required to improve clinical intervention. Pluripotent stem cells represent a renewable source of human tissue. Recent advances in three-dimensional cell culture have provided the field with more complex systems that better mimic liver physiology and function. Despite these improvements, current cell-based models are variable in performance and expensive to manufacture at scale. This is due, in part, to the use of poorly defined or cross-species materials within the process, severely affecting technology translation. To address this issue, we have developed an automated and economical platform to produce liver tissue at scale for modelling disease and small molecule screening. Stem cell derived liver spheres were formed by combining hepatic progenitors with endothelial cells and stellate cells, in the ratios found within the liver. The resulting tissue permitted the study of human liver biology ‘in the dish’ and could be scaled for screening. In summary, we have developed an automated differentiation system that permits reliable self-assembly of human liver tissue for biomedical application. Going forward we believe that this technology will not only serve as an in vitro resource, and may have an important role to play in supporting failing liver function in humans. Read More…

Pancreatic ductal adenocarcinoma is particularly metastatic, with dismal survival rates and few treatment options. Stiff fibrotic stroma is a hallmark of pancreatic tumours, but how stromal mechanosensing affects metastasis is still unclear. Here, we show that mechanical changes in the pancreatic cancer cell environment affect not only adhesion and migration, but also ATP/ADP and ATP/AMP ratios. Unbiased metabolomic analysis reveals that the creatine–phosphagen ATP-recycling system is a major mechanosensitive target. This system depends on arginine flux through the urea cycle, which is reflected by the increased incorporation of carbon and nitrogen from L-arginine into creatine and phosphocreatine on stiff matrix. We identify that CKB is a mechanosensitive transcriptional target of YAP, and thus it increases phosphocreatine production. We further demonstrate that the creatine–phosphagen system has a role in invasive migration, chemotaxis and liver metastasis of cancer cells. Read More…

Rac1 is a major regulator of actin dynamics, with GTP-bound Rac1 promoting actin assembly via the Scar/WAVE complex. CYRI competes with Scar/WAVE for interaction with Rac1 in a feedback loop regulating actin dynamics. Here, we reveal the nature of the CYRI-Rac1 interaction, through crystal structures of CYRI-B lacking the N-terminal helix (CYRI-BΔN) and the CYRI-BΔN:Rac1Q61L complex, providing the molecular basis for CYRI-B regulation of the Scar/WAVE complex. We reveal CYRI-B as having two subdomains – an N-terminal Rac1 binding subdomain with a unique Rac1-effector interface and a C-terminal Ratchet subdomain that undergoes conformational changes induced by Rac1 binding. Finally, we show that the CYRI protein family, CYRI-A and CYRI-B can produce an autoinhibited hetero- or homodimers, adding an additional layer of regulation to Rac1 signaling. Read More…

Pancreatic ductal adenocarcinoma is one of the most invasive and metastatic cancers and has a dismal 5-year survival rate. We show that N-WASP drives pancreatic cancer metastasis, with roles in both chemotaxis and matrix remodeling. lysophosphatidic acid, a signaling lipid abundant in blood and ascites fluid, is both a mitogen and chemoattractant for cancer cells. Pancreatic cancer cells break lysophosphatidic acid down as they respond to it, setting up a self-generated gradient driving tumor egress. N-WASP-depleted cells do not recognize lysophosphatidic acid gradients, leading to altered RhoA activation, decreased contractility and traction forces, and reduced metastasis. We describe a signaling loop whereby N-WASP and the endocytic adapter SNX18 promote lysophosphatidic acid-induced RhoA-mediated contractility and force generation by controlling lysophosphatidic acid receptor recycling and preventing degradation. This chemotactic loop drives collagen remodeling, tumor invasion, and metastasis and could be an important target against pancreatic cancer spread. Read More…

Actin-based protrusions are reinforced through positive feedback, but it is unclear what restricts their size, or limits positive signals when they retract or split. We identify an evolutionarily conserved regulator of actin-based protrusion: CYRI (CYFIP-related Rac interactor) also known as Fam49 (family of unknown function 49). CYRI binds activated Rac1 via a domain of unknown function (DUF1394) shared with CYFIP, defining DUF1394 as a Rac1-binding module. CYRI-depleted cells have broad lamellipodia enriched in Scar/WAVE, but reduced protrusion–retraction dynamics. Pseudopods induced by optogenetic Rac1 activation in CYRI-depleted cells are larger and longer lived. Conversely, CYRI overexpression suppresses recruitment of active Scar/WAVE to the cell edge, resulting in short-lived, unproductive protrusions. CYRI thus focuses protrusion signals and regulates pseudopod complexity by inhibiting Scar/WAVE-induced actin polymerization. It thus behaves like a ‘local inhibitor’ as predicted in widely accepted mathematical models, but not previously identified in cells. CYRI therefore regulates chemotaxis, cell migration and epithelial polarization by controlling the polarity and plasticity of protrusions. Read More…

Macropinocytosis is known as nonselective drinking of the cellular milieu for scavenging of nutrients and uptake of extracellular fluids. It has recently gained attention in cancer research, as targeting the uptake of nutrients or chemotherapeutic particles by tumor cells holds promise for new therapeutic interventions. Macropinocytic cups form from actin-based protrusions that coalesce into cups and collapse into large vesicles containing cargoes and transmembrane receptors, which can then be degraded in lysosomes or recycled back to the plasma membrane. How macropinocytosis is triggered and the various ways that cells utilize cargoes are topics of much interest. Here, we discuss emerging evidence that cargo or membrane receptor uptake in macropinocytosis may in some cases be selective and may substantially alter cell surface receptor trafficking and display. Read More…

Cells use actin-based protrusions not only to migrate, but also to sample their environment and take up liquids and particles, including nutrients, antigens and pathogens. Lamellipodia are sheet-like actin-based protrusions involved in sensing the substratum and directing cell migration. Related structures, macropinocytic cups, arise from lamellipodia ruffles and can take in large gulps of the surrounding medium. How cells regulate the balance between using lamellipodia for migration and macropinocytosis is not yet well understood. We recently identified CYRI proteins as RAC1-binding regulators of the dynamics of lamellipodia and macropinocytic events. This review discusses recent advances in our understanding of how cells regulate the balance between eating and walking by repurposing their actin cytoskeletons in response to environmental cues. Read More…

Dissemination of malignant cells from primary tumours to metastatic sites is a key step in cancer progression. Disseminated tumour cells preferentially settle in specific target organs, and the success of such metastases depends on dynamic interactions between cancer cells and the microenvironments they encounter at secondary sites. Two emerging concepts concerning the biology of metastasis are that organ-specific microenvironments influence the fate of disseminated cancer cells, and that cancer cell-extracellular matrix interactions have important roles at all stages of the metastatic cascade. The extracellular matrix is the complex and dynamic non-cellular component of tissues that provides a physical scaffold and conveys essential adhesive and paracrine signals for a tissue’s function. Here, we focus on how extracellular matrix dynamics contribute to liver metastases – a common and deadly event. We discuss how matrix components of the healthy and premetastatic liver support early seeding of disseminated cancer cells, and how the matrix derived from both cancer and liver contributes to the changes in niche composition as metastasis progresses. We also highlight the technical developments that are providing new insights into the stochastic, dynamic and multifaceted roles of the liver extracellular matrix in permitting and sustaining metastasis. An understanding of the contribution of the extracellular matrix to different stages of metastasis may well pave the way to targeted and effective therapies against metastatic disease. Read More…

Tumours evolve to cope with environmental stresses or challenges such as nutrient starvation, depletion of survival factors, and unbalanced mechanical forces. The uncontrolled growth and aberrant deregulation of core cell homeostatic pathways induced by genetic mutations create an environment of stress. Here, we explore how the adaptations of tumours to the changing environment can drive changes in the motility machinery of cells, affecting migration, invasion, and metastasis. Tumour cells can invade individually or collectively, or they can be extruded out of the surrounding epithelium. These mechanisms are thought to be modifications of normal processes occurring during development or tissue repair. Therefore, tumours may activate these pathways in response to environmental stresses, enabling them to survive in hostile environments and spread to distant sites. Read More…