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JCI Insight Oct 2019Increased fibrosis is a characteristic remodeling response to biomechanical and neurohumoral stress and a determinant of cardiac mechanical and electrical dysfunction in...
Increased fibrosis is a characteristic remodeling response to biomechanical and neurohumoral stress and a determinant of cardiac mechanical and electrical dysfunction in disease. Stress-induced activation of cardiac fibroblasts (CFs) is a critical step in the fibrotic response, although the precise sequence of events underlying activation of these critical cells in vivo remain unclear. Here, we tested the hypothesis that a βIV-spectrin/STAT3 complex is essential for maintenance of a quiescent phenotype (basal nonactivated state) in CFs. We reported increased fibrosis, decreased cardiac function, and electrical impulse conduction defects in genetic and acquired mouse models of βIV-spectrin deficiency. Loss of βIV-spectrin function promoted STAT3 nuclear accumulation and transcriptional activity, and it altered gene expression and CF activation. Furthermore, we demonstrate that a quiescent phenotype may be restored in βIV-spectrin-deficient fibroblasts by expressing a βIV-spectrin fragment including the STAT3-binding domain or through pharmacological STAT3 inhibition. We found that in vivo STAT3 inhibition abrogates fibrosis and cardiac dysfunction in the setting of global βIV-spectrin deficiency. Finally, we demonstrate that fibroblast-specific deletion of βIV-spectrin is sufficient to induce fibrosis and decreased cardiac function. We propose that the βIV-spectrin/STAT3 complex is a determinant of fibroblast phenotype and fibrosis, with implications for remodeling response in cardiovascular disease (CVD).
Topics: Animals; Cardiovascular Diseases; Disease Models, Animal; Female; Fibroblasts; Fibrosis; Heart Ventricles; Humans; Male; Mice; Mice, Knockout; STAT3 Transcription Factor; Spectrin; Ventricular Remodeling
PubMed: 31550236
DOI: 10.1172/jci.insight.131046 -
Cells Aug 2021The 20-60 μm axon initial segment (AIS) is proximally located at the interface between the axon and cell body. AIS has characteristic molecular and structural... (Review)
Review
The 20-60 μm axon initial segment (AIS) is proximally located at the interface between the axon and cell body. AIS has characteristic molecular and structural properties regulated by the crucial protein, ankyrin-G. The AIS contains a high density of Na channels relative to the cell body, which allows low thresholds for the initiation of action potential (AP). Molecular and physiological studies have shown that the AIS is also a key domain for the control of neuronal excitability by homeostatic mechanisms. The AIS has high plasticity in normal developmental processes and pathological activities, such as injury, neurodegeneration, and neurodevelopmental disorders (NDDs). In the first half of this review, we provide an overview of the molecular, structural, and ion-channel characteristics of AIS, AIS regulation through axo-axonic synapses, and axo-glial interactions. In the second half, to understand the relationship between NDDs and AIS, we discuss the activity-dependent plasticity of AIS, the human mutation of AIS regulatory genes, and the pathophysiological role of an abnormal AIS in NDD model animals and patients. We propose that the AIS may provide a potentially valuable structural biomarker in response to abnormal network activity in vivo as well as a new treatment concept at the neural circuit level.
Topics: Action Potentials; Ankyrins; Axon Initial Segment; Humans; Ion Channels; Mutation; Neurodevelopmental Disorders; Neuroglia; Neuronal Plasticity; Spectrin; Synapses
PubMed: 34440880
DOI: 10.3390/cells10082110 -
Cellular and Molecular Life Sciences :... Jul 2021The identification of the membrane periodic skeleton (MPS), composed of a periodic lattice of actin rings interconnected by spectrin tetramers, was enabled by the... (Review)
Review
The identification of the membrane periodic skeleton (MPS), composed of a periodic lattice of actin rings interconnected by spectrin tetramers, was enabled by the development of super-resolution microscopy, and brought a new exciting perspective to our view of neuronal biology. This exquisite cytoskeleton arrangement plays an important role on mechanisms regulating neuronal (dys)function. The MPS was initially thought to provide mainly for axonal mechanical stability. Since its discovery, the importance of the MPS in multiple aspects of neuronal biology has, however, emerged. These comprise its capacity to act as a signaling platform, regulate axon diameter-with important consequences on the efficiency of axonal transport and electrophysiological properties- participate in the assembly and function of the axon initial segment, and control axon microtubule stability. Recently, MPS disassembly has also surfaced as an early player in the course of axon degeneration. Here, we will discuss the current knowledge on the role of the MPS in axonal physiology and disease.
Topics: Animals; Axonal Transport; Axons; Cell Membrane; Cytoskeleton; Humans; Spectrin
PubMed: 34085116
DOI: 10.1007/s00018-021-03867-x -
Scientific Reports Mar 2021The neuronal membrane-associated periodic spectrin skeleton (MPS) contributes to neuronal development, remodeling, and organization. Post-translational modifications...
The neuronal membrane-associated periodic spectrin skeleton (MPS) contributes to neuronal development, remodeling, and organization. Post-translational modifications impinge on spectrin, the major component of the MPS, but their role remains poorly understood. One modification targeting spectrin is cleavage by calpains, a family of calcium-activated proteases. Spectrin cleavage is regulated by activated calpain, but also by the calcium-dependent binding of calmodulin (CaM) to spectrin. The physiologic significance of this balance between calpain activation and substrate-level regulation of spectrin cleavage is unknown. We report a strain of C57BL/6J mice harboring a single αII spectrin point mutation (Sptan1 c.3293G > A:p.R1098Q) with reduced CaM affinity and intrinsically enhanced sensitivity to calpain proteolysis. Homozygotes are embryonic lethal. Newborn heterozygotes of either gender appear normal, but soon develop a progressive ataxia characterized biochemically by accelerated calpain-mediated spectrin cleavage and morphologically by disruption of axonal and dendritic integrity and global neurodegeneration. Molecular modeling predicts unconstrained exposure of the mutant spectrin's calpain-cleavage site. These results reveal the critical importance of substrate-level regulation of spectrin cleavage for the maintenance of neuronal integrity. Given that excessive activation of calpain proteases is a common feature of neurodegenerative disease and traumatic encephalopathy, we propose that damage to the spectrin MPS may contribute to the neuropathology of many disorders.
Topics: Animals; Calpain; Cerebellar Ataxia; Cerebellum; Mice; Mice, Inbred C57BL; Point Mutation; Protein Binding; Proteolysis; Spectrin
PubMed: 33790315
DOI: 10.1038/s41598-021-86470-1 -
PloS One 2016The kidney displays specialized regions devoted to filtration, selective reabsorption, and electrolyte and metabolite trafficking. The polarized membrane pumps,...
The kidney displays specialized regions devoted to filtration, selective reabsorption, and electrolyte and metabolite trafficking. The polarized membrane pumps, channels, and transporters responsible for these functions have been exhaustively studied. Less examined are the contributions of spectrin and its adapter ankyrin to this exquisite functional topography, despite their established contributions in other tissues to cellular organization. We have examined in the rodent kidney the expression and distribution of all spectrins and ankyrins by qPCR, Western blotting, immunofluorescent and immuno electron microscopy. Four of the seven spectrins (αΙΙ, βΙ, βΙΙ, and βΙΙΙ) are expressed in the kidney, as are two of the three ankyrins (G and B). The levels and distribution of these proteins vary widely over the nephron. αΙΙ/βΙΙ is the most abundant spectrin, found in glomerular endothelial cells; on the basolateral membrane and cytoplasmic vesicles in proximal tubule cells and in the thick ascending loop of Henle; and less so in the distal nephron. βΙΙΙ spectrin largely replaces βΙΙ spectrin in podocytes, Bowman's capsule, and throughout the distal tubule and collecting ducts. βΙ spectrin is only marginally expressed; its low abundance hinders a reliable determination of its distribution. Ankyrin G is the most abundant ankyrin, found in capillary endothelial cells and all tubular segments. Ankyrin B populates Bowman's capsule, podocytes, the ascending thick loop of Henle, and the distal convoluted tubule. Comparison to the distribution of renal protein 4.1 isoforms and various membrane proteins indicates a complex relationship between the spectrin scaffold, its adapters, and various membrane proteins. While some proteins (e.g. ankyrin B, βΙΙΙ spectrin, and aquaporin 2) tend to share a similar distribution, there is no simple mapping of different spectrins or ankyrins to most membrane proteins. The implications of this data are discussed.
Topics: Animals; Ankyrins; Blotting, Western; Cytoskeleton; Exons; Kidney; Membrane Proteins; Mice; Mice, Inbred C57BL; Microscopy, Fluorescence; Microscopy, Immunoelectron; Polymerase Chain Reaction; Protein Isoforms; RNA, Messenger; Specific Pathogen-Free Organisms; Spectrin
PubMed: 26727517
DOI: 10.1371/journal.pone.0142687 -
Cellular Physiology and Biochemistry :... 2017Old human red blood cells (RBCs) have a reduced surface area with respect to young RBCs. If this decrease occurred through the release of vesicles similar to the...
BACKGROUND
Old human red blood cells (RBCs) have a reduced surface area with respect to young RBCs. If this decrease occurred through the release of vesicles similar to the spectrin-free vesicles that are shed in vitro under different experimental conditions or during storage, there would be no decrease of membrane-skeleton, but only of lipid bilayer surface area, during RBC ageing in vivo. However, we observed a decrease in spectrin and other membrane-skeletal proteins in old RBCs. Because RBCs contain components of the ubiquitin-proteasome system and other hydrolytic systems for protein degradation, we asked whether increased membrane-skeleton fragments could be detected in older RBCs.
METHODS
Four different anti-spectrin antibodies and an antibody anti-ubiquitin conjugates were used to analyse, by Western blotting, fragments of spectrin and other proteins in RBCs of different age separated in density gradients and characterized for their protein 4.1a/4.1b ratio as a cell age parameter.
RESULTS
spectrin fragments do exist in RBCs of all ages, they represent a minute fraction of all spectrin, are membrane-bound and not cytoplasmic and do not increase with cell age. Besides spectrin, other membrane-skeletal components decrease with cell age.
CONCLUSION
Observed results challenge the commonly accepted view that decrease in cell membrane throughout RBC life in vivo occurs via the release of spectrin-free vesicles.
Topics: Cellular Senescence; Cytoskeleton; Erythrocyte Membrane; Erythrocytes; Exosomes; Humans; Membrane Proteins; Protein Multimerization; Spectrin; Ubiquitination
PubMed: 28668958
DOI: 10.1159/000478769 -
MBio Apr 2024Remodeling the erythrocyte membrane and skeleton by the malarial parasite is closely associated with intraerythrocytic development. However, the mechanisms underlying...
UNLABELLED
Remodeling the erythrocyte membrane and skeleton by the malarial parasite is closely associated with intraerythrocytic development. However, the mechanisms underlying this association remain unclear. In this study, we present evidence that erythrocytic α-spectrin, but not β-spectrin, was dynamically ubiquitinated and progressively degraded during the intraerythrocytic development of from the ring to the schizont stage. We further observed an upregulated expression of phosphatidylinositol 3-kinase (PfPI3K) in the infected red blood cells during the intraerythrocytic development of the parasite. The data indicated that PfPI3K phosphorylated and activated erythrocytic ubiquitin-protein ligase, leading to increased α-spectrin ubiquitination and degradation during development. We further revealed that inhibition of the activity of PfPI3K impaired development and infectivity in mice. These findings collectively unveil an important mechanism of PfPI3K-ubiquitin-mediated degradation of α-spectrin during the intraerythrocytic development of species. Proteins in the PfPI3K regulatory pathway are novel targets for effective treatment of severe malaria.
IMPORTANCE
is the causative agent of severe malaria that causes millions of deaths globally. The parasite invades human red blood cells and induces a cascade of alterations in erythrocytes for development and proliferation. Remodeling the host erythrocytic cytoskeleton is a necessary process during parasitization, but its regulatory mechanisms remain to be elucidated. In this study, we observed that erythrocytic α-spectrin is selectively degraded after invasion, while β-spectrin remained intact. We found that the α-spectrin chain was profoundly ubiquitinated by E3 ubiquitin ligase and degraded by the 26S proteasome. E3 ubiquitin ligase activity was regulated by phosphatidylinositol 3-kinase (PfPI3K) signaling. Additionally, blocking the PfPI3K-ubiquitin-proteasome pathway in -infected red blood cells reduced parasite proliferation and infectivity. This study deepens our understanding of the regulatory mechanisms of host and malarial parasite interactions and paves the way for the exploration of novel antimalarial drugs.
Topics: Humans; Animals; Mice; Plasmodium falciparum; Spectrin; Erythrocytes; Malaria, Falciparum; Ubiquitin; Phosphatidylinositol 3-Kinase; Ubiquitin-Protein Ligases
PubMed: 38470053
DOI: 10.1128/mbio.03510-23 -
Experimental Biology and Medicine... Nov 2019We provide a review of Spectrin isoform function in the cytoplasm, the nucleus, the cell surface, and in intracellular signaling. We then discuss the importance of... (Review)
Review
UNLABELLED
We provide a review of Spectrin isoform function in the cytoplasm, the nucleus, the cell surface, and in intracellular signaling. We then discuss the importance of Spectrin’s E2/E3 chimeric ubiquitin conjugating and ligating activity in maintaining cellular homeostasis. Finally we present spectrin isoform subunit specific human diseases. We have created the Spectrinome, from the Human Proteome, Human Reactome and Human Atlas data and demonstrated how it can be a useful tool in visualizing and understanding spectrins myriad of cellular functions.
IMPACT STATEMENT
Spectrin was for the first 12 years after its discovery thought to be found only in erythrocytes. In 1981, Goodman and colleagues found that spectrin-like molecules were ubiquitously found in non-erythroid cells leading to a great multitude of publications over the next thirty eight years. The discovery of multiple spectrin isoforms found associated with every cellular compartment, and representing 2-3% of cellular protein, has brought us to today’s understanding that spectrin is a scaffolding protein, with its own E2/E3 chimeric ubiquitin conjugating ligating activity that is involved in virtually every cellular function. We cover the history, localized functions of spectrin isoforms, human diseases caused by mutations, and provide the spectrinome: a useful tool for understanding the myriad of functions for one of the most important proteins in all eukaryotic cells.
Topics: Animals; Cytoplasm; Humans; Nuclear Matrix-Associated Proteins; Protein Isoforms; Proteomics; Spectrin
PubMed: 31483159
DOI: 10.1177/1535370219867269 -
PloS One 2015Invadosomes are actin-rich adhesion structures involved in tissue invasion and extracellular matrix (ECM) remodelling. αII-Spectrin, an ubiquitous scaffolding component...
Invadosomes are actin-rich adhesion structures involved in tissue invasion and extracellular matrix (ECM) remodelling. αII-Spectrin, an ubiquitous scaffolding component of the membrane skeleton and a partner of actin regulators (ABI1, VASP and WASL), accumulates highly and specifically in the invadosomes of multiple cell types, such as mouse embryonic fibroblasts (MEFs) expressing SrcY527F, the constitutively active form of Src or activated HMEC-1 endothelial cells. FRAP and live-imaging analysis revealed that αII-spectrin is a highly dynamic component of invadosomes as actin present in the structures core. Knockdown of αII-spectrin expression destabilizes invadosomes and reduces the ability of the remaining invadosomes to digest the ECM and to promote invasion. The ECM degradation defect observed in spectrin-depleted-cells is associated with highly dynamic and unstable invadosome rings. Moreover, FRAP measurement showed the specific involvement of αII-spectrin in the regulation of the mobile/immobile β3-integrin ratio in invadosomes. Our findings suggest that spectrin could regulate invadosome function and maturation by modulating integrin mobility in the membrane, allowing the normal processes of adhesion, invasion and matrix degradation. Altogether, these data highlight a new function for spectrins in the stability of invadosomes and the coupling between actin regulation and ECM degradation.
Topics: Animals; Cell Line; Extracellular Matrix; Gene Knockdown Techniques; Humans; Integrin beta3; Mice; Molecular Imaging; Mutation; Podosomes; Protein Stability; Spectrin
PubMed: 25830635
DOI: 10.1371/journal.pone.0120781 -
Proceedings of the National Academy of... Jul 2019βII-spectrin is the generally expressed member of the β-spectrin family of elongated polypeptides that form micrometer-scale networks associated with plasma membranes....
βII-spectrin is the generally expressed member of the β-spectrin family of elongated polypeptides that form micrometer-scale networks associated with plasma membranes. We addressed in vivo functions of βII-spectrin in neurons by knockout of βII-spectrin in mouse neural progenitors. βII-spectrin deficiency caused severe defects in long-range axonal connectivity and axonal degeneration. βII-spectrin-null neurons exhibited reduced axon growth, loss of actin-spectrin-based periodic membrane skeleton, and impaired bidirectional axonal transport of synaptic cargo. We found that βII-spectrin associates with KIF3A, KIF5B, KIF1A, and dynactin, implicating spectrin in the coupling of motors and synaptic cargo. βII-spectrin required phosphoinositide lipid binding to promote axonal transport and restore axon growth. Knockout of ankyrin-B (AnkB), a βII-spectrin partner, primarily impaired retrograde organelle transport, while double knockout of βII-spectrin and AnkB nearly eliminated transport. Thus, βII-spectrin promotes both axon growth and axon stability through establishing the actin-spectrin-based membrane-associated periodic skeleton as well as enabling axonal transport of synaptic cargo.
Topics: Animals; Axons; Brain; Cell Membrane; Connectome; Mice; Mice, Knockout; Nerve Tissue Proteins; Spectrin
PubMed: 31209033
DOI: 10.1073/pnas.1820649116