ESCRT-0 component HRS and actin polymerization factor WASH reside in adjacent endosomal domains. MacDonald et al. show that HRS controls WASH localization and recycling of WASH-dependent transmembrane cargo. Cargo binding to endosomal actin thus acts as sorting signal to oppose ubiquitin-mediated degradation.
The perinuclear actin cap is an important cytoskeletal structure that regulates nuclear morphology and re-orientation during front-rear polarisation. The mechanisms regulating the actin cap are currently poorly understood. Here, we demonstrate that STEF/TIAM2, a Rac1 selective guanine nucleotide exchange factor, localises at the nuclear envelope, co-localising with the key perinuclear proteins Nesprin-2G and Non-muscle myosin IIB (NMMIIB), where it regulates perinuclear Rac1 activity. We show that STEF depletion reduces apical perinuclear actin cables (a phenotype rescued by targeting active Rac1 to the nuclear envelope), increases nuclear height and impairs nuclear re-orientation. STEF down-regulation also reduces perinuclear pMLC and decreases myosin-generated tension at the nuclear envelope, suggesting that STEF-mediated Rac1 activity regulates NMMIIB activity to promote stabilisation of the perinuclear actin cap. Finally, STEF depletion decreases nuclear stiffness and reduces expression of TAZ-regulated genes, indicating an alteration in mechanosensing pathways as a consequence of disruption of the actin cap.
Alcohol abuse is among the top causes of preventable death, generating considerable financial, health, and societal burdens. Paradoxically, alcohol...
Cell invasion and metastasis is a multi-step process, initialised through the acquisition of a migratory phenotype and the ability to move through differing and complex 3D extracellular environments. In this study we set out to identify the parameters required for invasive cell migration in 3D environments. Cells interact with the extracellular matrix via transmembranespanning integrin adhesion complexes, which are well characterised in cells plated on 2D surfaces, yet much less is known about them in cells embedded in 3D matrices. We establish a technique to determine the composition of cell matrix adhesion complexes of invasive breast cancer cells in 3D matrices and on 2D surfaces and we identify an interaction complex enriched in 3D adhesive sites required for 3D invasive migration. Depletion of β-PIX-Myosin18A (Myo18A) abolishes cancer cell invasion, without negatively affecting matrix degradation, Rho GTPase signalling, or protrusion formation in collagen matrices. Instead, in a mechanism only seen in cells moving through 3D matrix, β-PIX and Myo18A drive the polarised recruitment of non-muscle Myosin 2A (NM2A) to the tips of protrusions. This recruitment of NM2A is required for the creation of an NM2A-NM2B isoform gradient, which ranges from the protrusion to the nucleus. We observe a requirement for active force transmission to the nucleus during invasive migration that is needed to pull the nucleus forward. We postulate that the establishment of the NM2A-NM2B actomyosin gradient facilitates the coupling of cell-matrix interactions at the protrusive cell front with nuclear movement, enabling effective invasive migration and front-rear cell polarity.
The mechanical properties of the extracellular matrix within tumours control multiple cellular functions that drive cancer invasion and metastasis. However, the mechanisms controlling microenvironmental force sensation and transmission, and how these regulate transcriptional reprogramming and invasion, are unclear. Our aim was to understand how mechanical inputs are transmitted bidirectionally and translated into biochemical and transcriptional outputs to drive breast cancer progression. We reveal that adhesion receptor and growth factor receptor crosstalk regulates a bidirectional feedback mechanism co-ordinating force-dependent transcriptional regulation and invasion.Integrin αVβ6 drives invasion in a range of carcinomas and is a potential therapeutic target. αVβ6 exhibits unique biophysical properties that promote force-generation and increase matrix rigidity. We employed an inter-disciplinary approach incorporating proteomics, biophysical techniques and multimodal live-cell imaging to dissect the role of αVβ6-EGFR crosstalk on transmission of mechanical signals bidirectionally between the extracellular matrix and nucleus.We show that αVβ6 expression correlates with poor prognosis in triple-negative breast cancer (TNBC) and drives invasion of TNBC cells. Moreover, our data show that a complex regulatory mechanism exists involving crosstalk between αVβ6 integrin and EGFR that impacts matrix stiffness, force transmission to the nucleus, transcriptional reprogramming and microenvironment rigidity. αVβ6 engagement triggers EGFR & MAPK signalling and αVβ6-EGFR crosstalk regulates mutual receptor trafficking mechanisms. Consequently, EGF stimulation suppresses αVβ6-mediated force-application on the matrix and nuclear shuttling of force-dependent transcriptional co-activators YAP/TAZ. Finally, we show that crosstalk between αVβ6 & EGFR regulates TNBC invasion. We propose a model whereby αVβ6-EGFR crosstalk regulates matrix stiffening, but also the transmission of extracellular forces into the cell in order to co-ordinate transcriptional reprogramming and invasion. To exploit adhesion receptors and receptor tyrosine kinases therapeutically, it will be essential to understand the integration of their signalling functions and how crosstalk mechanisms influence invasion and the response of tumours to molecular therapeutics. Regulation of TP53 Activity through Phosphorylation Regulation of TP53 ActivityRNA Polymerase II Transcription Gene expression (Transcription) Transcriptional Regulation by TP53 Generic Transcription Pathway Costimulation by the CD28 family CD28 dependent PI3K/Akt signaling CD28 co-stimulation VEGFR2 mediated vascular permeability Intracellular signaling by second messengers PIP3 activates AKT signaling Constitutive Signaling by AKT1 E17K in Cancer AKT-mediated inactivation of FOXO1A Regulation of TP53 Activity through Association with Co-factors AKT phosphorylates targets in the nucleus RUNX2 regulates genes involved in cell migration AKT phosphorylates targets in the cytosol Downregulation of ERBB2:...
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