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Clinical and Translational Medicine Sep 2023The roles of circRNA and N6-methyladenosine (m A) methylation in Crohn's disease (CD) have drawn much attention. Therefore, this investigation aimed to discover how the...
BACKGROUND
The roles of circRNA and N6-methyladenosine (m A) methylation in Crohn's disease (CD) have drawn much attention. Therefore, this investigation aimed to discover how the m A modification of circRNAs contributes to CD progression.
METHODS
The study performed circRNA sequencing on colon samples from four CD patients and four normal controls (NCs) to screen for dysregulated circRNAs. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to validate the candidate circRNA expression and determine its correlation to CD-associated inflammatory indicators. In vivo and in vitro investigations were conducted to examine the functions and pathways of circPRKAR1B in CD, besides investigating the m A modification role in circRNA expression modulation.
RESULTS
The RNA-seq revealed that hsa_circ_0008039 (circPRKAR1B) was the most significant upregulated circRNA and was identified as the candidate circRNA for further examinations. Relative circPRKAR1B expression was significantly upregulated in CD colon tissues and closely related to CD-associated inflammatory indices. The circPRKAR1B expression and function were regulated by methyltransferase-like 3 (METTL3)-mediated m A methylation. In vitro studies indicated that circPRKAR1B promoted pyroptosis mediated by NLRP3 inflammasome (NLRP3; nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3) and impaired autophagy by interacting with the RNA-binding protein (RBP) SPTBN1, (SPTBN1; spectrin beta, non-erythrocytic 1). The in vivo investigations revealed the treatment effects of si-circPRKAR1B and si-METTL3 in colitis models of IL-10-deficient mice.
CONCLUSION
Our study reveals that METTL3-mediated m A modification of circPRKAR1B promotes Crohn's colitis by aggravating NLRP3 inflammasome-mediated pyroptosis via autophagy impairment in colonic epithelial cells.
Topics: Animals; Mice; Autophagy; Colitis; Crohn Disease; Inflammasomes; NLR Family, Pyrin Domain-Containing 3 Protein; Pyroptosis; RNA, Circular
PubMed: 37679886
DOI: 10.1002/ctm2.1405 -
Biomolecules Feb 2024The protein 4.1R is an essential component of the erythrocyte membrane skeleton, serving as a key structural element and contributing to the regulation of the membrane's... (Review)
Review
The protein 4.1R is an essential component of the erythrocyte membrane skeleton, serving as a key structural element and contributing to the regulation of the membrane's physical properties, including mechanical stability and deformability, through its interaction with spectrin-actin. Recent research has uncovered additional roles of 4.1R beyond its function as a linker between the plasma membrane and the membrane skeleton. It has been found to play a crucial role in various biological processes, such as cell fate determination, cell cycle regulation, cell proliferation, and cell motility. Additionally, 4.1R has been implicated in cancer, with numerous studies demonstrating its potential as a diagnostic and prognostic biomarker for tumors. In this review, we provide an updated overview of the gene and protein structure of 4.1R, as well as its cellular functions in both physiological and pathological contexts.
Topics: Membrane Proteins; Cytoskeletal Proteins; Spectrin; Actins; Erythrocyte Membrane
PubMed: 38397451
DOI: 10.3390/biom14020214 -
Acta Neuropathologica Communications Jul 2023Oligodendrocyte (OL) injury and loss are central features of evolving lesions in multiple sclerosis. Potential causative mechanisms of OL loss include metabolic stress...
Oligodendrocyte (OL) injury and loss are central features of evolving lesions in multiple sclerosis. Potential causative mechanisms of OL loss include metabolic stress within the lesion microenvironment. Here we use the injury response of primary human OLs (hOLs) to metabolic stress (reduced glucose/nutrients) in vitro to help define the basis for the in situ features of OLs in cases of MS. Under metabolic stress in vitro, we detected reduction in ATP levels per cell that precede changes in survival. Autophagy was initially activated, although ATP levels were not altered by inhibitors (chloroquine) or activators (Torin-1). Prolonged stress resulted in autophagy failure, documented by non-fusion of autophagosomes and lysosomes. Consistent with our in vitro results, we detected higher expression of LC3, a marker of autophagosomes in OLs, in MS lesions compared to controls. Both in vitro and in situ, we observe a reduction in nuclear size of remaining OLs. Prolonged stress resulted in increased ROS and cleavage of spectrin, a target of Ca-dependent proteases. Cell death was however not prevented by inhibitors of ferroptosis or MPT-driven necrosis, the regulated cell death (RCD) pathways most likely to be activated by metabolic stress. hOLs have decreased expression of VDAC1, VDAC2, and of genes regulating iron accumulation and cyclophilin. RNA sequencing analyses did not identify activation of these RCD pathways in vitro or in MS cases. We conclude that this distinct response of hOLs, including resistance to RCD, reflects the combined impact of autophagy failure, increased ROS, and calcium influx, resulting in metabolic collapse and degeneration of cellular structural integrity. Defining the basis of OL injury and death provides guidance for development of neuro-protective strategies.
Topics: Humans; Multiple Sclerosis; Reactive Oxygen Species; Oligodendroglia; Cell Death; Multiple Sclerosis, Chronic Progressive; Adenosine Triphosphate
PubMed: 37408029
DOI: 10.1186/s40478-023-01601-1 -
Nature Communications Oct 2023E-cadherin is an essential cell‒cell adhesion protein that mediates canonical cadherin-catenin complex formation in epithelial lateral membranes. Ankyrin-G (AnkG), a...
E-cadherin is an essential cell‒cell adhesion protein that mediates canonical cadherin-catenin complex formation in epithelial lateral membranes. Ankyrin-G (AnkG), a scaffold protein linking membrane proteins to the spectrin-based cytoskeleton, coordinates with E-cadherin to maintain epithelial cell polarity. However, the molecular mechanisms governing this complex formation and its relationships with the cadherin-catenin complex remain elusive. Here, we report that AnkG employs a promiscuous manner to encapsulate three discrete sites of E-cadherin by the same region, a dynamic mechanism that is distinct from the canonical 1:1 molar ratio previously described for other AnkG or E-cadherin-mediated complexes. Moreover, we demonstrate that AnkG-binding-deficient E-cadherin exhibited defective accumulation at the lateral membranes and show that disruption of interactions resulted in cell polarity malfunction. Finally, we demonstrate that E-cadherin is capable of simultaneously anchoring to AnkG and β-catenin, providing mechanistic insights into the functional orchestration of the ankyrin-spectrin complex with the cadherin-catenin complex. Collectively, our results show that complex formation between E-cadherin and AnkG is dynamic, which enables the maintenance of epithelial cell polarity by ensuring faithful targeting of the adhesion molecule-scaffold protein complex, thus providing molecular mechanisms for essential E-cadherin-mediated complex assembly at cell‒cell junctions.
Topics: Ankyrins; Cadherins; Cell Adhesion; Cell Polarity; Epithelial Cells; Spectrin; Humans
PubMed: 37891324
DOI: 10.1038/s41467-023-42628-1 -
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 -
Redox Biology Apr 2024The function of SLC7A11 in the process of ferroptosis is well-established, as it regulates the synthesis of glutathione (GSH), thereby influencing tumor development...
The function of SLC7A11 in the process of ferroptosis is well-established, as it regulates the synthesis of glutathione (GSH), thereby influencing tumor development along with drug resistance in non-small cell lung cancer (NSCLC). However, the determinants governing SLC7A11's membrane trafficking and localization remain unknown. Our study identified SPTBN2 as a ferroptosis suppressor, enhancing NSCLC cells resistance to ferroptosis inducers. Mechanistically, SPTBN2, through its CH domain, interacted with SLC7A11 and connected it with the motor protein Arp1, thus facilitating the membrane localization of SLC7A11 - a prerequisite for its role as System Xc, which mediates cystine uptake and GSH synthesis. Consequently, SPTBN2 suppressed ferroptosis through preserving the functional activity of System Xc on the membrane. Moreover, Inhibiting SPTBN2 increased the sensitivity of NSCLC cells to cisplatin through ferroptosis induction, both in vitro and in vivo. Using Abrine as a potential SPTBN2 inhibitor, its efficacy in promoting ferroptosis and sensitizing NSCLC cells to cisplatin was validated. Collectively, SPTBN2 is a potential therapeutic target for addressing ferroptosis dysfunction and cisplatin resistance in NSCLC.
Topics: Humans; Amino Acid Transport System y+; Biological Transport; Carcinoma, Non-Small-Cell Lung; Cisplatin; Ferroptosis; Glutathione; Lung Neoplasms; Spectrin
PubMed: 38241838
DOI: 10.1016/j.redox.2024.103039 -
Pesticide Biochemistry and Physiology Nov 2023The female reproductive potential plays a crucial role in reproduction, population dynamics and population maintenance. However, the function of endogenous genes in...
The female reproductive potential plays a crucial role in reproduction, population dynamics and population maintenance. However, the function of endogenous genes in undifferentiated germ cells has been largely unknown in Bactrocera dorsalis. In this study, the conservative analysis showed that α-Spectrin shared a similarity in B. dorsalis and other dipteral flies. Further, the differential expression of α-Spectrin was examined in B. dorsalis by RT-qPCR, and the expression pattern of α-Spectrin protein was identified in female adult ovaries by using immunostaining. During the development of ovary, the change on the number of undifferentiated germ cells was also characterized and analyzed. To understand the function of α-Spectrin in B. dorsalis ovary, the RNAi-based knockdown was conducted, and the RNAi efficiency was examined by RT-qPCR, western blot and bioassay. The results revealed that the α-Spectrin dsRNA could strikingly decrease the expression level of α-Spectrin in ovaries and diminish oviposition and ovary size as a consequence of downregulation of α-Spectrin. Overall, our study facilitates reproductive research on the function of conservative genes in B. dorsalis ovary, which may provide a new insight into seeking novel target genes for pest management control.
Topics: Animals; Female; RNA Interference; Spectrin; Reproduction; Tephritidae
PubMed: 37945250
DOI: 10.1016/j.pestbp.2023.105611 -
Biology Open Sep 2023Robust expression of shortened, functional dystrophin provided impetus to develop adeno-associated virus (AAV)-based constructs for clinical application. Because several...
Robust expression of shortened, functional dystrophin provided impetus to develop adeno-associated virus (AAV)-based constructs for clinical application. Because several cassettes are being tested in clinical trials, this study compared the efficacies of four shortened dystrophin-promoter combinations with implications for outcomes in clinical trials: MHCK7 or MCK promoter with a shortened dystrophin transgene containing the N-terminus and spectrin repeats R1, R2, R3 and R24 (rAAVrh74.MHCK7.micro-dystrophin and rAAVrh74.MCK.micro-dystrophin, respectively); shortened dystrophin construct containing the neuronal nitric oxide (nNOS) binding site (rAAVrh74.MHCK7.DV.mini-dystrophin); and shortened dystrophin containing the C-terminus (rAAVrh74.MHCK7.micro-dystrophin.Cterm). Functional and histological benefit were examined at 4 weeks following intramuscular delivery in mdx mice. rAAVrh74.MHCK7.micro-dystrophin provided the most robust transgene expression and significantly increased specific force output in the tibialis anterior muscle. Muscle environment was normalized (i.e. reductions in central nucleation), indicating functional and histological advantages of rAAVrh74.MHCK7.micro-dystrophin. Thus, promoter choice and transgene design are critical for optimal dystrophin expression/distribution for maximal functional improvement.
Topics: Mice; Animals; Dystrophin; Muscular Dystrophy, Duchenne; Mice, Inbred mdx; Dependovirus; Actin Cytoskeleton; Disease Models, Animal
PubMed: 37670674
DOI: 10.1242/bio.059797 -
Cardiovascular Research Jun 2024βII spectrin is a cytoskeletal protein known to be tightly linked to heart development and cardiovascular electrophysiology. However, the roles of βII spectrin in...
AIMS
βII spectrin is a cytoskeletal protein known to be tightly linked to heart development and cardiovascular electrophysiology. However, the roles of βII spectrin in cardiac contractile function and pathological post-myocardial infarction remodeling remain unclear. Here, we investigated whether and how βII spectrin, the most common isoform of non-erythrocytic spectrin in cardiomyocytes, is involved in cardiac contractile function and ischemia/reperfusion (I/R) injury.
METHODS AND RESULTS
We observed that the levels of serum βII spectrin breakdown products (βII SBDPs) were significantly increased in patients with acute myocardial infarction (AMI). Concordantly, βII spectrin was degraded into βII SBDPs by calpain in mouse hearts after I/R injury. Using tamoxifen-inducible cardiac-specific βII spectrin knockout mice, we found that deletion of βII spectrin in the adult heart resulted in spontaneous development of cardiac contractile dysfunction, cardiac hypertrophy and fibrosis at 5 weeks after tamoxifen treatment. Moreover, at 1 week after tamoxifen treatment, although spontaneous cardiac dysfunction in cardiac-specific βII spectrin knockout mice had not developed, deletion of βII spectrin in the heart exacerbated I/R-induced cardiomyocyte death and heart failure. Furthermore, restoration of βII spectrin expression via adenoviral small activating RNA (saRNA) delivery into the heart reduced I/R injury. Immunoprecipitation coupled with mass spectrometry (IP-LC-MS/MS) analyses and functional studies revealed that βII spectrin is indispensable for mitochondrial complex I activity and respiratory function. Mechanistically, βII spectrin promotes translocation of NADH:ubiquinone oxidoreductase 75 kDa Fe-S protein 1 (NDUFS1) from the cytosol to mitochondria by crosslinking with actin filaments (F-actin) to maintain F-actin stability.
CONCLUSION
βII spectrin is an essential cytoskeletal element for preserving mitochondrial homeostasis and cardiac function. Defects in βII spectrin exacerbate cardiac I/R injury.
PubMed: 38832923
DOI: 10.1093/cvr/cvae116 -
The Journal of Neuroscience : the... Jul 2023Neural circuit assembly is a multistep process where synaptic partners are often born at distinct developmental stages, and yet they must find each other and form...
Neural circuit assembly is a multistep process where synaptic partners are often born at distinct developmental stages, and yet they must find each other and form precise synaptic connections with one another. This developmental process often relies on late-born neurons extending their processes to the appropriate layer to find and make synaptic connections to their early-born targets. The molecular mechanism responsible for the integration of late-born neurons into an emerging neural circuit remains unclear. Here, we uncovered a new role for the cytoskeletal protein βII-spectrin in properly positioning presynaptic and postsynaptic neurons to the developing synaptic layer. Loss of βII-spectrin disrupts retinal lamination, leads to synaptic connectivity defects, and results in impaired visual function in both male and female mice. Together, these findings highlight a new function of βII-spectrin in assembling neural circuits in the mouse outer retina. Neurons that assemble into a functional circuit are often integrated at different developmental time points. However, the molecular mechanism that guides the precise positioning of neuronal processes to the correct layer for synapse formation is relatively unknown. Here, we show a new role for the cytoskeletal scaffolding protein, βII-spectrin in the developing retina. βII-spectrin is required to position presynaptic and postsynaptic neurons to the nascent synaptic layer in the mouse outer retina. Loss of βII-spectrin disrupts positioning of neuronal processes, alters synaptic connectivity, and impairs visual function.
Topics: Male; Mice; Female; Animals; Spectrin; Cytoskeletal Proteins; Neurons; Cytoskeleton
PubMed: 37369589
DOI: 10.1523/JNEUROSCI.0063-23.2023