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Brain, Behavior, and Immunity Nov 2023The pathophysiology of autism spectrum disorder (ASD) involves genetic and environmental factors. Mounting evidence demonstrates a role for the gut microbiome in ASD,...
BACKGROUND
The pathophysiology of autism spectrum disorder (ASD) involves genetic and environmental factors. Mounting evidence demonstrates a role for the gut microbiome in ASD, with signaling via short-chain fatty acids (SCFA) as one mechanism. Here, we utilize mice carrying deletion to exons 4-22 of Shank3 (Shank3) to model gene by microbiome interactions in ASD. We identify SCFA acetate as a mediator of gut-brain interactions and show acetate supplementation reverses social deficits concomitant with alterations to medial prefrontal cortex (mPFC) transcriptional regulation independent of microbiome status.
METHODS
Shank3 and wild-type (Wt) littermates were divided into control, Antibiotic (Abx), Acetate and Abx + Acetate groups upon weaning. After six weeks, animals underwent behavioral testing. Molecular analysis including 16S and metagenomic sequencing, metabolomic and transcriptional profiling were conducted. Additionally, targeted serum metabolomic data from Phelan McDermid Syndrome (PMS) patients (who are heterozygous for the Shank3 gene) were leveraged to assess levels of SCFA's relative to ASD clinical measures.
RESULTS
Shank3 mice were found to display social deficits, dysregulated gut microbiome and decreased cecal levels of acetate - effects exacerbated by Abx treatment. RNA-sequencing of mPFC showed unique gene expression signature induced by microbiome depletion in the Shank3 mice. Oral treatment with acetate reverses social deficits and results in marked changes in gene expression enriched for synaptic signaling, pathways among others, even in Abx treated mice. Clinical data showed sex specific correlations between levels of acetate and hyperactivity scores.
CONCLUSION
These results suggest a key role for the gut microbiome and the neuroactive metabolite acetate in regulating ASD-like behaviors.
Topics: Humans; Male; Female; Mice; Animals; Autism Spectrum Disorder; Nerve Tissue Proteins; Prefrontal Cortex; Acetates; Dietary Supplements; Microfilament Proteins
PubMed: 37657643
DOI: 10.1016/j.bbi.2023.08.020 -
EBioMedicine Mar 2024Tertiary Lymphoid Structures (TLS) correlate with positive outcomes in patients with NSCLC and the efficacy of immune checkpoint blockade (ICB) in cancer. The actin... (Observational Study)
Observational Study
BACKGROUND
Tertiary Lymphoid Structures (TLS) correlate with positive outcomes in patients with NSCLC and the efficacy of immune checkpoint blockade (ICB) in cancer. The actin regulatory protein hMENA undergoes tissue-specific splicing, producing the epithelial hMENA linked to favorable prognosis in early NSCLC, and the mesenchymal hMENAΔv6 found in invasive cancer cells and pro-tumoral cancer-associated fibroblasts (CAFs). This study investigates how hMENA isoforms in tumor cells and CAFs relate to TLS presence, localization and impact on patient outcomes and ICB response.
METHODS
Methods involved RNA-SEQ on NSCLC cells with depleted hMENA isoforms. A retrospective observational study assessed tissues from surgically treated N0 patients with NSCLC, using immunohistochemistry for tumoral and stromal hMENA isoforms, fibronectin, and TLS presence. ICB-treated patient tumors were analyzed using Nanostring nCounter and GeoMx spatial transcriptomics. Multiparametric flow cytometry characterized B cells and tissue-resident memory T cells (T). Survival and ICB response were estimated in the cohort and validated using bioinformatics pipelines in different datasets.
FINDINGS
Findings indicate that hMENA in NSCLC cells upregulates the TLS regulator LTβR, decreases fibronectin, and favors CXCL13 production by T. Conversely, hMENAΔv6 in CAFs inhibits LTβR-related NF-kB pathway, reduces CXCL13 secretion, and promotes fibronectin production. These patterns are validated in N0 NSCLC tumors, where hMENA11a expression, CAF hMENAΔv6, and stromal fibronectin are associated with intratumoral TLS, linked to memory B cells and predictive of longer survival. The hMENA isoform pattern, fibronectin, and LTβR expression broadly predict ICB response in tumors where TLS indicates an anti-tumor immune response.
INTERPRETATION
This study uncovers hMENA alternative splicing as an unexplored contributor to TLS-related Tumor Immune Microenvironment (TIME) and a promising biomarker for clinical outcomes and likely ICB responsiveness in N0 patients with NSCLC.
FUNDING
This work is supported by AIRC (IG 19822), ACC (RCR-2019-23669120), CAL.HUB.RIA Ministero Salute PNRR-POS T4, "Ricerca Corrente" granted by the Italian Ministry of Health.
Topics: Humans; Lung Neoplasms; Fibronectins; Immune Checkpoint Inhibitors; Tertiary Lymphoid Structures; Microfilament Proteins; Cell Line, Tumor; Protein Isoforms; Carcinoma, Non-Small-Cell Lung; Tumor Microenvironment
PubMed: 38340557
DOI: 10.1016/j.ebiom.2024.105003 -
Cell Death and Differentiation Feb 2024Actin-binding LIM protein 1 (ABLIM1), a member of the LIM-domain protein family, has been reported as a suppressor in several tumors whereas its role in colorectal...
Actin-binding LIM protein 1 (ABLIM1), a member of the LIM-domain protein family, has been reported as a suppressor in several tumors whereas its role in colorectal cancer (CRC) remains unknown. In this study, we find that ABLIM1 is up-regulated in CRC patients and high levels of ABLIM1 predict short disease-free survival time. Knock-down of ABLIM1 in CRC cell lines by lenti-virus leads to inhibited cell proliferation, migration, and invasion capabilities in vitro and impaired growth of tumor xenografts and liver metastasis lesions in vivo, while ABLIM1 overexpression accelerates tumor growth and invasion in vitro. Mechanistically, we uncover that ABLIM1 activates the NF-ĸB/CCL-20 signaling through modulating IĸBα ubiquitination and proteasomal-mediated degradation. Further co-immunoprecipitation, in vivo and in vitro ubiquitination assays reveal ABLIM1 as a novel ubiquitin E3 ligase binding to IĸBα. Interestingly, The E3 ligase catalysis activity of ABLIM1 depends on its 402-778aa rather than its LIM domains and its interaction with IĸBα relies on the HP domain. Our findings delineate the oncogenic role of ABLIM1 in CRC progression and reveal it as a novel E3 ligase targeting IĸBα, providing new insights into the regulation of NF-ĸB signaling in tumors.
Topics: Humans; Cell Line, Tumor; Colorectal Neoplasms; LIM Domain Proteins; Microfilament Proteins; NF-kappa B; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination
PubMed: 38228802
DOI: 10.1038/s41418-024-01256-y -
Scientific Reports Aug 2023Mounting evidence has found that tumor microenvironment (TME) plays an important role in the tumor progression of lung adenocarcinoma (LUAD). However, the roles of tumor...
Mounting evidence has found that tumor microenvironment (TME) plays an important role in the tumor progression of lung adenocarcinoma (LUAD). However, the roles of tumor microenvironment-related genes in immunotherapy and clinical outcomes remain unclear. In this study, 6 TME-related genes (PLK1, LDHA, FURIN, FSCN1, RAB27B, and MS4A1) were identified to construct the prognostic model. The established risk scores were able to predict outcomes at 1, 3, and 5 years with greater accuracy than previously known models. Moreover, the risk score was closely associated with immune cell infiltration and the immunoregulatory genes including T cell exhaustion markers. In conclusion, the TME risk score can function as an independent prognostic biomarker and a predictor for evaluating immunotherapy response in LUAD patients, which provides recommendations for improving patients' response to immunotherapy and promoting personalized tumor immunotherapy in the future.
Topics: Humans; Tumor Microenvironment; Prognosis; Adenocarcinoma of Lung; CD3 Complex; Immunotherapy; Lung Neoplasms; Carrier Proteins; Microfilament Proteins
PubMed: 37604869
DOI: 10.1038/s41598-023-40980-2 -
Biophysical Journal Sep 2023
Topics: Pseudopodia; Actins
PubMed: 37311456
DOI: 10.1016/j.bpj.2023.05.030 -
Cells Nov 2023The TEM8 protein represents an emerging biomarker in many solid tumor histologies. Given the various roles it plays in oncogenesis, including but not limited to... (Review)
Review
The TEM8 protein represents an emerging biomarker in many solid tumor histologies. Given the various roles it plays in oncogenesis, including but not limited to angiogenesis, epithelial-to-mesenchymal transition, and cell migration, TEM8 has recently served and will continue to serve as the target of novel oncologic therapies. We review herein the role of TEM8 in oncogenesis. We review its normal function, highlight the additional roles it plays in the tumor microenvironment, and synthesize pre-clinical and clinical data currently available. We underline the protein's prognostic and predictive abilities in various solid tumors by (1) highlighting its association with more aggressive disease biology and poor clinical outcomes and (2) assessing its associated clinical trial landscape. Finally, we offer future directions for clinical studies involving TEM8, including incorporating pre-clinical agents into clinical trials and combining previously tested oncologic therapies with currently available treatments, such as immunotherapy.
Topics: Humans; Receptors, Cell Surface; Neoplasm Proteins; Microfilament Proteins; Neoplasms; Cell Transformation, Neoplastic; Carcinogenesis; Biology; Tumor Microenvironment
PubMed: 37998358
DOI: 10.3390/cells12222623 -
Sleep Jul 2023Shank3 is a shared risk gene for autism spectrum disorders and schizophrenia. Sleep defects have been characterized for autism models with Shank3 mutations; however,...
Shank3 is a shared risk gene for autism spectrum disorders and schizophrenia. Sleep defects have been characterized for autism models with Shank3 mutations; however, evidence has been lacking for the potential sleep defects caused by Shank3 mutation associated with schizophrenia and how early in development these defects may occur. Here we characterized the sleep architecture of adolescent mice carrying a schizophrenia-linked, R1117X mutation in Shank3. We further employed GRABDA dopamine sensor and fiber photometry to record dopamine release in the nucleus accumbens during sleep/wake states. Our results show that homozygous mutant R1117X mice have significantly reduced sleep in the dark phase during adolescence, altered electroencephalogram power, especially during the rapid-eye-movement sleep, and dopamine hyperactivity during sleep but not during wakefulness. Further analyses suggest that these adolescent defects in sleep architecture and dopaminergic neuromodulation tightly correlate with the social novelty preference later in adulthood and predict adult social performance during same-sex social interactions. Our results provide novel insights into the sleep phenotypes in mouse models of schizophrenia and the potential use of developmental sleep as a predictive metric for adult social symptoms. Together with recent studies in other Shank3 models, our work underscores the idea that Shank3-involved circuit disruptions may be one of the shared pathologies in certain types of schizophrenia and autism. Future research is needed to establish the causal relationship among adolescent sleep defects, dopaminergic dysregulation, and adult behavioral changes in Shank3 mutation animals and other models.
Topics: Mice; Animals; Schizophrenia; Dopamine; Nerve Tissue Proteins; Mutation; Autism Spectrum Disorder; Sleep; Microfilament Proteins
PubMed: 37144901
DOI: 10.1093/sleep/zsad131 -
Scientific Reports Oct 2023CLIC5 belongs to a family of ion channels with six members reported so far. In vertebrates, the CLIC5 gene encodes two different isoforms, CLIC5A and CLIC5B. In addition...
CLIC5 belongs to a family of ion channels with six members reported so far. In vertebrates, the CLIC5 gene encodes two different isoforms, CLIC5A and CLIC5B. In addition to its ion channel activity, there is evidence for further functions of CLIC5A, such as the remodeling of the actin cytoskeleton during the formation of a functional glomerulus in the vertebrate kidney. However, its specific role is still incompletely understood and a specific functional role for CLIC5B has not been described yet. Here we report our findings on the differential expression and functions of Clic5a and Clic5b during zebrafish kidney development. Whole-mount in situ hybridization studies revealed specific expression of clic5a in the eye and pronephric glomerulus, and clic5b is expressed in the gut, liver and the pronephric tubules. Clic5 immunostainings revealed that Clic5b is localized in the cilia. Whereas knockdown of Clic5a resulted in leakiness of the glomerular filtration barrier, Clic5b deficient embryos displayed defective ciliogenesis, leading to ciliopathy-associated phenotypes such as ventral body curvature, otolith deposition defects, altered left-right asymmetry and formation of hydrocephalus and pronephric cysts. In addition, Clic5 deficiency resulted in dysregulation of cilia-dependent Wnt signalling pathway components. Mechanistically, we identified a Clic5-dependent activation of the membrane-cytoskeletal linker proteins Ezrin/Radixin/Moesin (ERM) in the pronephric tubules of zebrafish. In conclusion, our in vivo data demonstrates a novel role for Clic5 in regulating essential ciliary functions and identified Clic5 as a positive regulator of ERM phosphorylation.
Topics: Animals; Actin Cytoskeleton; Chloride Channels; Chlorides; Cilia; Kidney Glomerulus; Microfilament Proteins; Zebrafish
PubMed: 37848494
DOI: 10.1038/s41598-023-44235-y -
Journal of Cell Science Jan 2024The actin cytoskeleton performs multiple cellular functions, and as such, actin polymerization must be tightly regulated. We previously demonstrated that reversible,...
The actin cytoskeleton performs multiple cellular functions, and as such, actin polymerization must be tightly regulated. We previously demonstrated that reversible, non-degradative ubiquitylation regulates the function of the actin polymerase VASP in developing neurons. However, the underlying mechanism of how ubiquitylation impacts VASP activity was unknown. Here, we show that mimicking multi-monoubiquitylation of VASP at K240 and K286 negatively regulates VASP interactions with actin. Using in vitro biochemical assays, we demonstrate the reduced ability of multi-monoubiquitylated VASP to bind, bundle, and elongate actin filaments. However, multi-monoubiquitylated VASP maintained the ability to bind and protect barbed ends from capping protein. Finally, we demonstrate the electroporation of recombinant multi-monoubiquitylated VASP protein altered cell spreading morphology. Collectively, these results suggest a mechanism in which ubiquitylation controls VASP-mediated actin dynamics.
Topics: Actin Cytoskeleton; Actins; Microfilament Proteins; Neurons; Phosphoproteins
PubMed: 38277158
DOI: 10.1242/jcs.261527 -
Proceedings of the National Academy of... Aug 2023Arp2/3 complex generates branched actin networks that drive fundamental processes such as cell motility and cytokinesis. The complex comprises seven proteins, including...
Arp2/3 complex generates branched actin networks that drive fundamental processes such as cell motility and cytokinesis. The complex comprises seven proteins, including actin-related proteins (Arps) 2 and 3 and five scaffolding proteins (ArpC1-ArpC5) that mediate interactions with a pre-existing (mother) actin filament at the branch junction. Arp2/3 complex exists in two main conformations, inactive with the Arps interacting end-to-end and active with the Arps interacting side-by-side like subunits of the short-pitch helix of the actin filament. Several cofactors drive the transition toward the active state, including ATP binding to the Arps, WASP-family nucleation-promoting factors (NPFs), actin monomers, and binding of Arp2/3 complex to the mother filament. The precise contribution of each cofactor to activation is poorly understood. We report the 3.32-Å resolution cryo-electron microscopy structure of a transition state of Arp2/3 complex activation with bound constitutively dimeric NPF. Arp2/3 complex-binding region of the NPF N-WASP was fused C-terminally to the α and β subunits of the CapZ heterodimer. One arm of the NPF dimer binds Arp2 and the other binds actin and Arp3. The conformation of the complex is intermediate between those of inactive and active Arp2/3 complex. Arp2, Arp3, and actin also adopt intermediate conformations between monomeric (G-actin) and filamentous (F-actin) states, but only actin hydrolyzes ATP. In solution, the transition complex is kinetically shifted toward the short-pitch conformation and has higher affinity for F-actin than inactive Arp2/3 complex. The results reveal how all the activating cofactors contribute in a coordinated manner toward Arp2/3 complex activation.
Topics: Protein Binding; Models, Molecular; Actins; Protein Multimerization; Protein Subunits; Humans; Animals; Mice
PubMed: 37549294
DOI: 10.1073/pnas.2306165120