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ArXiv May 2024Crossbridge binding, state transitions, and force in active muscle is dependent on the radial spacing between the myosin-containing thick filament and the...
Crossbridge binding, state transitions, and force in active muscle is dependent on the radial spacing between the myosin-containing thick filament and the actin-containing thin filament in the filament lattice. This radial lattice spacing has been previously shown through spatially explicit modeling and experimental efforts to greatly affect quasi-static, isometric, force production in muscle. It has recently been suggested that this radial spacing might also be able to drive differences in mechanical function, or net work, under dynamic oscillations like those which occur in muscles . However, previous spatially explicit models either had no radial spacing dependence, meaning the lattice spacing could not be investigated, or did include radial spacing dependence but could not reproduce net work during dynamic oscillations and only investigated isometric contractions. Here we show the first spatially explicit model to include radial crossbridge dependence which can produce mechanical function similar to real muscle. Using this spatially explicit model of a half sarcomere, we show that when oscillated at strain amplitudes and frequencies like those in the hawk moth , mechanical function (net work) does depend on the lattice spacing. In addition, since the trajectory of lattice spacing changes during dynamic oscillation can vary from organism to organism, we can prescribe a trajectory of lattice spacing changes in the spatially explicit half sarcomere model and investigate the extent to which the time course of lattice spacing changes can affect mechanical function. We simulated a half sarcomere undergoing dynamic oscillations and prescribed the Poisson's ratio of the lattice to be either 0 (constant lattice spacing) or 0.5 (isovolumetric lattice spacing changes). We also simulated net work using lattice spacing data taken from which has a variable Poisson's ratio. Our simulation results indicate that the lattice spacing can change the mechanical function of muscle, and that in some cases a 1 nm difference can switch the net work of the half sarcomere model from positive (motor-like) to negative (brake-like).
PubMed: 38855552
DOI: No ID Found -
Theranostics 2024Skin cells actively metabolize nutrients to ensure cell proliferation and differentiation. Psoriasis is an immune-disorder-related skin disease with hyperproliferation...
Skin cells actively metabolize nutrients to ensure cell proliferation and differentiation. Psoriasis is an immune-disorder-related skin disease with hyperproliferation in epidermal keratinocytes and is increasingly recognized to be associated with metabolic disturbance. However, the metabolic adaptations and underlying mechanisms of epidermal hyperproliferation in psoriatic skin remain largely unknown. Here, we explored the role of metabolic competition in epidermal cell proliferation and differentiation in psoriatic skin. Bulk- and single-cell RNA-sequencing, spatial transcriptomics, and glucose uptake experiments were used to analyze the metabolic differences in epidermal cells in psoriasis. Functional validation and was done using imiquimod-like mouse models and inflammatory organoid models. We observed the highly proliferative basal cells in psoriasis act as the winners of the metabolic competition to uptake glucose from suprabasal cells. Using single-cell metabolic analysis, we found that the "winner cells" promote OXPHOS pathway upregulation by COX7B and lead to increased ROS through glucose metabolism, thereby promoting the hyperproliferation of basal cells in psoriasis. Also, to prevent toxic damage from ROS, basal cells activate the glutathione metabolic pathway to increase their antioxidant capacity to assist in psoriasis progression. We further found that COX7B promotes psoriasis development by modulating the activity of the PPAR signaling pathway by bulk RNA-seq analysis. We also observed glucose starvation and high expression of SLC7A11 that causes suprabasal cell disulfide stress and affects the actin cytoskeleton, leading to immature differentiation of suprabasal cells in psoriatic skin. Our study demonstrates the essential role of cellular metabolic competition for skin tissue homeostasis.
Topics: Psoriasis; Glucose; Humans; Animals; Mice; Keratinocytes; Cell Proliferation; Cell Differentiation; Disease Models, Animal; Single-Cell Analysis; Epidermal Cells; Reactive Oxygen Species; Energy Metabolism; Epidermis; Imiquimod; Male
PubMed: 38855186
DOI: 10.7150/thno.93764 -
Neurobiology of Disease Aug 2024Periventricular nodular heterotopia (PNH), the most common brain malformation diagnosed in adulthood, is characterized by the presence of neuronal nodules along the...
Periventricular nodular heterotopia (PNH), the most common brain malformation diagnosed in adulthood, is characterized by the presence of neuronal nodules along the ventricular walls. PNH is mainly associated with mutations in the FLNA gene - encoding an actin-binding protein - and patients often develop epilepsy. However, the molecular mechanisms underlying the neuronal failure still remain elusive. It has been hypothesized that dysfunctional cortical circuitry, rather than ectopic neurons, may explain the clinical manifestations. To address this issue, we depleted FLNA from cortical pyramidal neurons of a conditional Flna mice by timed in utero electroporation of Cre recombinase. We found that FLNA regulates dendritogenesis and spinogenesis thus promoting an appropriate excitatory/inhibitory inputs balance. We demonstrated that FLNA modulates RAC1 and cofilin activity through its interaction with the Rho-GTPase Activating Protein 24 (ARHGAP24). Collectively, we disclose an uncharacterized role of FLNA and provide strong support for neural circuit dysfunction being a consequence of FLNA mutations.
Topics: Animals; Filamins; rac1 GTP-Binding Protein; Mice; Cerebral Cortex; Actin Depolymerizing Factors; Pyramidal Cells; Neurogenesis; GTPase-Activating Proteins; Neurons; Mice, Transgenic; Periventricular Nodular Heterotopia; Neuropeptides
PubMed: 38852754
DOI: 10.1016/j.nbd.2024.106558 -
Cell Communication and Signaling : CCS Jun 2024Abnormally expressed BCR/ABL protein serves as the basis for the development of chronic myeloid leukaemia (CML). The F-actin binding domain (FABD), which is a crucial...
BACKGROUND
Abnormally expressed BCR/ABL protein serves as the basis for the development of chronic myeloid leukaemia (CML). The F-actin binding domain (FABD), which is a crucial region of the BCR/ABL fusion protein, is also located at the carboxyl end of the c-ABL protein and regulates the kinase activity of c-ABL. However, the precise function of this domain in BCR/ABL remains uncertain.
METHODS
The FABD-deficient adenovirus vectors Ad-BCR/ABL△FABD, wild-type Ad-BCR/ABL and the control vector Adtrack were constructed, and 32D cells were infected with these adenoviruses separately. The effects of FABD deletion on the proliferation and apoptosis of 32D cells were evaluated by a CCK-8 assay, colony formation assay, flow cytometry and DAPI staining. The levels of phosphorylated BCR/ABL, p73, and their downstream signalling molecules were detected by western blot. The intracellular localization and interaction of BCR/ABL with the cytoskeleton-related protein F-actin were identified by immunofluorescence and co-IP. The effect of FABD deletion on BCR/ABL carcinogenesis in vivo was explored in CML-like mouse models. The degree of leukaemic cell infiltration was observed by Wright‒Giemsa staining and haematoxylin and eosin (HE) staining.
RESULTS
We report that the loss of FABD weakened the proliferation-promoting ability of BCR/ABL, accompanied by the downregulation of BCR/ABL downstream signals. Moreover, the deletion of FABD resulted in a change in the localization of BCR/ABL from the cytoplasm to the nucleus, accompanied by an increase in cell apoptosis due to the upregulation of p73 and its downstream proapoptotic factors. Furthermore, we discovered that the absence of FABD alleviated leukaemic cell infiltration induced by BCR/ABL in mice.
CONCLUSIONS
These findings reveal that the deletion of FABD diminished the carcinogenic potential of BCR/ABL both in vitro and in vivo. This study provides further insight into the function of the FABD domain in BCR/ABL.
Topics: Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Fusion Proteins, bcr-abl; Animals; Humans; Mice; Cell Proliferation; Apoptosis; Actins; Carcinogenesis; Protein Domains; Cell Line, Tumor
PubMed: 38849885
DOI: 10.1186/s12964-024-01694-8 -
Bone Research Jun 2024DNAX-associated protein 12 kD size (DAP12) is a dominant immunoreceptor tyrosine-based activation motif (ITAM)-signaling adaptor that activates costimulatory signals...
DNAX-associated protein 12 kD size (DAP12) is a dominant immunoreceptor tyrosine-based activation motif (ITAM)-signaling adaptor that activates costimulatory signals essential for osteoclastogenesis. Although several DAP12-associated receptors (DARs) have been identified in osteoclasts, including triggering receptor expressed on myeloid cells 2 (TREM-2), C-type lectin member 5 A (CLEC5A), and sialic acid-binding Ig-like lectin (Siglec)-15, their precise role in the development of osteoclasts and bone remodeling remain poorly understood. In this study, mice deficient in Trem-2, Clec5a, Siglec-15 were generated. In addition, mice double deficient in these DAR genes and FcεRI gamma chain (FcR)γ, an alternative ITAM adaptor to DAP12, were generated. Bone mass analysis was conducted on all mice. Notably, Siglec-15 deficient mice and Siglec-15/FcRγ double deficient mice exhibited mild and severe osteopetrosis respectively. In contrast, other DAR deficient mice showed normal bone phenotype. Likewise, osteoclasts from Siglec-15 deficient mice failed to form an actin ring, suggesting that Siglec-15 promotes bone resorption principally by modulating the cytoskeletal organization of osteoclasts. Furthermore, biochemical analysis revealed that Sigelc-15 activates macrophage colony-stimulating factor (M-CSF)-induced Ras-associated protein-1 (RAP1)/Ras-related C3 botulinum toxin substrate 1 (Rac1) pathway through formation of a complex with p130CAS and CrkII, leading to cytoskeletal remodeling of osteoclasts. Our data provide genetic and biochemical evidence that Siglec-15 facilitates M-CSF-induced cytoskeletal remodeling of the osteoclasts.
Topics: Animals; Osteoclasts; Signal Transduction; Macrophage Colony-Stimulating Factor; rap1 GTP-Binding Proteins; Mice; Cytoskeleton; Mice, Knockout; Mice, Inbred C57BL; Membrane Proteins; rac GTP-Binding Proteins; Membrane Glycoproteins; Receptors, Immunologic; Immunoglobulins
PubMed: 38849345
DOI: 10.1038/s41413-024-00340-w -
Scientific Reports Jun 2024The proteasome-associated deubiquitinase USP14 is a potential drug target. Using an inducible USP14 knockout system in colon cancer cells, we found that USP14 depletion...
The proteasome-associated deubiquitinase USP14 is a potential drug target. Using an inducible USP14 knockout system in colon cancer cells, we found that USP14 depletion impedes cellular proliferation, induces cell cycle arrest, and leads to a senescence-like phenotype. Transcriptomic analysis revealed altered gene expression related to cell division and cellular differentiation. USP14 knockout cells also exhibited changes in morphology, actin distribution, and expression of actin cytoskeletal components. Increased ubiquitin turnover was observed, offset by upregulation of polyubiquitin genes UBB and UBC. Pharmacological inhibition of USP14 with IU1 increased ubiquitin turnover but did not affect cellular growth or morphology. BioGRID data identified USP14 interactors linked to actin cytoskeleton remodeling, DNA damage repair, mRNA splicing, and translation. In conclusion, USP14 loss in colon cancer cells induces a transient quiescent cancer phenotype not replicated by pharmacologic inhibition of its deubiquitinating activity.
Topics: Humans; Cellular Senescence; Colorectal Neoplasms; Cell Proliferation; Ubiquitin Thiolesterase; Cell Line, Tumor; Phenotype; Proteasome Endopeptidase Complex; Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Ubiquitin
PubMed: 38844605
DOI: 10.1038/s41598-024-63791-5 -
Biophysics Reviews Jun 2024Cell migration is a fundamental process for life and is highly dependent on the dynamical and mechanical properties of the cytoskeleton. Intensive physical and... (Review)
Review
Cell migration is a fundamental process for life and is highly dependent on the dynamical and mechanical properties of the cytoskeleton. Intensive physical and biochemical crosstalk among actin, microtubules, and intermediate filaments ensures their coordination to facilitate and enable migration. In this review, we discuss the different mechanical aspects that govern cell migration and provide, for each mechanical aspect, a novel perspective by juxtaposing two complementary approaches to the biophysical study of cytoskeletal crosstalk: live-cell studies (often referred to as top-down studies) and cell-free studies (often referred to as bottom-up studies). We summarize the main findings from both experimental approaches, and we provide our perspective on bridging the two perspectives to address the open questions of how cytoskeletal crosstalk governs cell migration and makes cells move.
PubMed: 38840976
DOI: 10.1063/5.0198119 -
BioRxiv : the Preprint Server For... Jun 20242',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) is an abundant constituent of central nervous system non-compact myelin, frequently used as a marker antigen for...
2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) is an abundant constituent of central nervous system non-compact myelin, frequently used as a marker antigen for myelinating cells. The catalytic activity of CNPase, the 3'-hydrolysis of 2',3'-cyclic nucleotides, is well characterised , but the function of CNPase remains unclear. CNPase interacts with the actin cytoskeleton to counteract the developmental closure of cytoplasmic channels that travel through compact myelin; its enzymatic activity may be involved in adenosine metabolism and RNA degradation. We developed a set of high-affinity nanobodies recognizing the phosphodiesterase domain of CNPase, and the crystal structures of each complex show that the five nanobodies have distinct epitopes. One of the nanobodies bound deep into the CNPase active site and acted as an inhibitor. Moreover, the nanobodies were characterised in imaging applications and as intrabodies, expressed in mammalian cells, such as primary oligodendrocytes. Fluorescently labelled nanobodies functioned in imaging of teased nerve fibers and whole brain tissue sections, as well as super-resolution microscopy. These anti-CNPase nanobodies provide new tools for structural and functional biology of myelination, including high-resolution imaging of nerve tissue.
PubMed: 38826303
DOI: 10.1101/2024.05.25.595513 -
Molecules and Cells May 2024The actin-based cytoskeleton is considered a fundamental driving force for cell differentiation and development. Destrin (Dstn), a member of the actin-depolymerizing...
The actin-based cytoskeleton is considered a fundamental driving force for cell differentiation and development. Destrin (Dstn), a member of the actin-depolymerizing factor family, regulates actin dynamics by treadmilling actin filaments and increasing globular actin pools. However, the specific developmental roles of dstn have yet to be fully elucidated. Here, we investigated the physiological functions of dstn during early embryonic development using Xenopus laevis as an experimental model organism. dstn is expressed in anterior neural tissue and neural plate during Xenopus embryogenesis. Depleting dstn promoted morphants with short body axes and small heads. Moreover, dstn inhibition extended the neural plate region, impairing cell migration and distribution during neurulation. In addition to the neural plate, dstn knockdown perturbed neural crest cell migration. Our data suggest new insights for understanding the roles of actin dynamics in embryonic neural development, simultaneously presenting a new challenge for studying the complex networks governing cell migration involving actin dynamics.
PubMed: 38825188
DOI: 10.1016/j.mocell.2024.100076 -
JACS Au May 2024The actin cytoskeleton and its elaborate interplay with the plasma membrane participate in and control numerous biological processes in eukaryotic cells. Malfunction of...
The actin cytoskeleton and its elaborate interplay with the plasma membrane participate in and control numerous biological processes in eukaryotic cells. Malfunction of actin networks and changes in their dynamics are related to various diseases, from actin myopathies to uncontrolled cell growth and tumorigenesis. Importantly, the biophysical and mechanical properties of actin and its assemblies are deeply intertwined with the biological functions of the cytoskeleton. Novel tools to study actin and its associated biophysical features are, therefore, of prime importance. Here we develop a new approach which exploits fluorescence lifetime imaging microscopy (FLIM) and environmentally sensitive fluorophores termed molecular rotors, acting as quantitative microviscosity sensors, to monitor dynamic viscoelastic properties of both actin structures and lipid membranes. In order to reproduce a minimal actin cortex in synthetic cell models, we encapsulated and attached actin networks to the lipid bilayer of giant unilamellar vesicles (GUVs). Using a cyanine-based molecular rotor, DiSC(3), we show that different types of actin bundles are characterized by distinct packing, which can be spatially resolved using FLIM. Similarly, we show that a lipid bilayer-localized molecular rotor can monitor the effects of attaching cross-linked actin networks to the lipid membrane, revealing an increase in membrane viscosity upon actin attachment. Our approach bypasses constraints associated with existing methods for actin imaging, many of which lack the capability for direct visualization of biophysical properties. It can therefore contribute to a deeper understanding of the role that actin plays in fundamental biological processes and help elucidate the underlying biophysics of actin-related diseases.
PubMed: 38818078
DOI: 10.1021/jacsau.4c00237