The cholesterol biosynthesis enzyme FAXDC2 couples Wnt/β-catenin to RTK/MAPK signaling

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Abstract

Wnts, cholesterol, and MAPK signaling are essential for development and adult homeostasis. Here we report for the first time that fatty acid hydroxylase domain containing 2 (FAXDC2), a previously uncharacterized enzyme, functions as a methyl sterol oxidase catalyzing C4 demethylation in the Kandutsch-Russell branch of the cholesterol biosynthesis pathway. FAXDC2, a paralog of MSMO1, regulates the abundance of specific C4-methyl sterols lophenol and dihydro-TMAS. Highlighting its clinical relevance, FAXDC2 is repressed in Wnt/β-catenin high cancer xenografts, in a mouse genetic model of Wnt activation, and in human colorectal cancers. Moreover, in primary human colorectal cancers, the sterol lophenol, regulated by FAXDC2, accumulates in the cancerous tissues and not in adjacent normal tissues. FAXDC2 links Wnts to RTK/MAPK signaling. Wnt inhibition drives increased recycling of RTKs and activation of the MAPK pathway, and this requires FAXDC2. Blocking Wnt signaling in Wnt-high cancers causes both differentiation and senescence; and this is prevented by knockout of FAXDC2. Our data shows the integration of three ancient pathways, Wnts, cholesterol synthesis, and RTK/MAPK signaling, in cellular proliferation and differentiation.

Authors

Babita Madan, Shawn R. Wadia, Siddhi Patnaik, Nathan Harmston, Emile K.W. Tan, Iain Bee Huat Tan, W. David Nes, Enrico Petretto, David M. Virshup

Disassembly of the TRIM56-ATR complex promotes cytoDNA/cGAS/STING axis-dependent intervertebral disc inflammatory degeneration

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Abstract

Given the leading cause of disability worldwide, low back pain (LBP) is recognized as a pivotal socio-economic challenge to the aging population, which is importantly attributed to intervertebral disc degeneration (IVDD). Elastic nucleus pulposus (NP) tissue is essential for the maintenance of IVD structural and functional integrity. The accumulation of senescent NP cells with inflammatory hypersecretory phenotype due to aging and other damaged factors is a distinctive hallmark of IVDD initiation and progression. In this study, we revealed a mechanism of IVDD progression in which aberrant genomic DNA damage promoted NP cell inflammatory senescence via activation of the cGAS-STING axis but not AIM2 inflammasome assembly. ATR deficiency destroyed genomic integrity and led to cytosolic mislocalization of genomic DNA, which acted as a powerful driver of cGAS-STING axis-dependent inflammatory phenotype acquisition during NP cell senescence. Mechanically, the disassembly of ATR-TRIM56 complex with the enzyme activity liberation of USP5 and TRIM25 drove change in ATR ubiquitination, with ATR switching from K63-linked modification to K48-linked modification, promoting ubiquitin-proteasome-dependent dynamic instability of ATR protein during NP cell senescent progression. Importantly, an engineered extracellular vesicle (EV)-based strategy for delivering ATR-overexpressing plasmid cargo efficiently diminished DNA damage-associated NP cell senescence and substantially mitigated IVDD progression, indicating promising targets and efficient approaches for ameliorating the impact of IVDD.

Authors

Weifeng Zhang, Gaocai Li, Xingyu Zhou, Huaizhen Liang, Bide Tong, Di Wu, Kevin Yang, Yu Song, Bingjin Wang, Zhiwei Liao, Liang Ma, Wencan Ke, Xiaoguang Zhang, Jie Lei, Chunchi Lei, Xiaobo Feng, Kun Wang, Kangcheng Zhao, Cao Yang

Targeting mitochondrial dynamics of morphine-responsive dopaminergic neurons ameliorates opiate withdrawal

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Abstract

Converging studies demonstrate the dysfunction of the dopaminergic neurons following chronic opioid administration. However, the therapeutic strategies targeting opioid-responsive dopaminergic ensembles that contribute to the development of opioid withdrawal remain to be elucidated. Here, we used the neuronal activity-dependent Tet-Off system to label dopaminergic ensembles in response to initial morphine exposure (Mor-Ens) in the ventral tegmental area (VTA). Fiber optic photometry recording and transcriptome analysis revealed downregulated spontaneous activity, dysregulated mitochondrial respiratory, ultrastructure, and oxidoreductase signal pathways after chronic morphine administration in these dopaminergic ensembles. Mitochondrial fragmentation and the decreased mitochondrial fusion gene mitofusin 1 (Mfn1) were found in these ensembles after prolonged opioid withdrawal. Restoration of Mfn1 in the dopaminergic Mor-Ens attenuated excessive oxidative stress and the development of opioid withdrawal. Administration of Mdivi-1, a mitochondrial fission inhibitor, ameliorated the mitochondrial fragmentation and maladaptation of the neuronal plasticity in these Mor-Ens, accompanied by attenuated development of opioid withdrawal after chronic morphine administration, without affecting the analgesic effect of morphine. These findings highlighted the plastic architecture of mitochondria as a potential therapeutic target for opioid analgesic-induced substance use disorders.

Authors

Changyou Jiang, Han Huang, Xiao Yang, Qiumin Le, Xing Liu, Lan Ma, Feifei Wang

Sterol biosynthesis regulates TLR signaling and the innate immune response in Smith-Lemli-Opitz syndrome model

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Abstract

Authors

Kristin Gabor, Emily V. Mesev, Jennifer Madenspacher, Julie M. Meacham, Prashant Rai, Sookjin Moon, Christopher A. Wassif, Saame Raza Shaikh, Charles J. Tucker, Peer W. Karmaus, Simona Bianconi, Forbes D. Porter, Michael B. Fessler

Tumor-educated Gr1+CD11b+ cells drive breast cancer metastasis via OSM/IL6-JAK-induced cancer cell plasticity

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Abstract

Cancer cell plasticity contributes to therapy resistance and metastasis, which represent the main causes of cancer-related death, including in breast cancer. The tumor microenvironment drives cancer cell plasticity and metastasis, and unravelling the underlying cues may provide novel strategies to manage metastatic disease. Using breast cancer experimental models and transcriptomic analyses, we showed that stem cell antigen-1 positive (SCA1+) murine breast cancer cells enriched during tumor progression and metastasis had higher in vitro cancer stem cell-like properties, enhanced in vivo metastatic ability, and generated tumors rich in Gr1high Ly6G+CD11b+ cells. In turn, tumor-educated Gr1+CD11b+(Tu-Gr1+CD11b+) cells rapidly and transiently converted low metastatic SCA1- cells into highly metastatic SCA1+ cells via secreted OSM and IL6. JAK inhibition prevented OSM/IL6-induced SCA1+ population enrichment while OSM/IL6 depletion suppressed Tu-Gr1+CD11b+-induced SCA1+ population enrichment in vitro and metastasis in vivo. Moreover, chemotherapy-selected highly metastatic 4T1 cells maintained high SCA1+ positivity through autocrine IL6 production and in vitro JAK inhibition blunted SCA1 positivity and metastatic capacity. Importantly, Tu-Gr1+CD11b+ cells invoked a gene signature in tumor cells predicting shorter OS, RFS and lung metastasis in breast cancer patients. Collectively, our data identified OSM/IL6-JAK as a clinically relevant paracrine/autocrine axis instigating breast cancer cell plasticity and triggering metastasis.

Authors

Sanam Peyvandi, Manon Bulliard, Alev Yilmaz, Annamaria Kauzlaric, Rachel Marcone, Lisa Haerri, Oriana Coquoz, Yu-Ting Huang, Nathalie Duffey, Laetitia Gafner, Girieca Lorusso, Nadine Fournier, Qiang Lan, Curzio Rüegg

APOL1-mediated monovalent cation transport contributes to APOL1-mediated podocytopathy in kidney disease

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Abstract

Two coding variants of apolipoprotein L1 (APOL1) called G1 and G2 explain much of the excess risk of kidney disease in African Americans. While various cytotoxic phenotypes have been reported in experimental models, the proximal mechanism by which G1 and G2 cause kidney disease is poorly understood. Here, we leveraged three experimental models and a recently reported small molecule blocker of APOL1 protein, VX-147, to identify the upstream mechanism of G1-induced cytotoxicity. In HEK293 cells, we demonstrated that G1-mediated Na+ import/K+ efflux triggered activation of G protein-coupled receptor (GPCR)-IP3-mediated calcium release from the endoplasmic reticulum (ER), impaired mitochondrial ATP production, and impaired translation, which were all reversed by VX-147. In human podocyte-like epithelial cells (HUPEC), we demonstrated that G1 caused cytotoxicity that was again reversible by VX-147. Finally, in podocytes isolated from APOL1 G1 transgenic mice, we showed that Interferon gamma (IFNγ)-mediated induction of G1 caused K+ efflux, activation of GPCR-IP3 signaling, and inhibition of translation, podocyte injury, and proteinuria, all reversed by VX-147. Together, these results establish APOL1-mediated Na+/K+ transport as the proximal driver of APOL1-mediated kidney disease.

Authors

Somenath Datta, Brett M. Antonio, Nathan H. Zahler, Jonathan W. Theile, Doug Krafte, Hengtao Zhang, Paul B. Rosenberg, Alec B. Chaves, Deborah M. Muoio, Guofang Zhang, Daniel Silas, Guojie Li, Karen Soldano, Sarah Nystrom, Davis Ferreira, Sara E. Miller, James R. Bain, Michael J. Muehlbauer, Olga Ilkayeva, Thomas C. Becker, Hans-Ewald Hohmeier, Christopher B. Newgard, Opeyemi A. Olabisi

Contralateral second dose improves antibody responses to a two-dose mRNA vaccination regimen

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Abstract

BACKGROUND. Vaccination is typically administered without regard to site of prior vaccination but this factor may substantially impact downstream immune responses. METHODS. We assessed serological responses to initial COVID-19 vaccination in baseline seronegative adults who received second–dose boosters in the ipsilateral or contralateral arm relative to initial vaccination. We measured serum SARS-CoV2 spike-specific Ig, RBD-specific IgG, SARS-CoV-2-nucleocapsid-specific IgG, and neutralizing antibody titers against SARS-CoV-2.D614G (early strain) and SARS-CoV-2.B.1.1.529 (Omicron) at approximately 0.6, 8, and 14 months after boosting. RESULTS. In 947 individuals, contralateral boosting was associated with higher spike-specific serum Ig, and this effect increased over time from a 1.1-fold to a 1.4-fold increase by 14 months (P < 0.001). A similar pattern was seen for RBD-specific IgG. Among 54 pairs matched for age, gender and relevant time intervals, arm groups had similar antibody levels at W2 but contralateral boosting resulted in significantly higher binding and neutralizing antibody titers at W3 and W4, with progressive increase over time, ranging from 1.3-fold (total Ig, P = 0.007) to 4.0-fold (pseudovirus neutralization to B.1.1.529 P < 0.001). CONCLUSIONS. In previously unexposed adults receiving an initial vaccine series with the BNT162b2 mRNA COVID-19 vaccine, contralateral boosting substantially increases antibody magnitude and breadth at times beyond 3 weeks after vaccination. This effect should be considered during arm selection in the context of multi-dose vaccine regimens.

Authors

Sedigheh Fazli, Archana Thomas, Abram E. Estrada, Hiro A.P. Ross, David Xthona Lee, Steven Kazmierczak, Mark K. Slifka, David Montefiori, William B. Messer, Marcel E. Curlin

Glucocorticoid receptor-dependent therapeutic efficacy of tauroursodeoxycholic acid in preclinical models of Spinocerebellar ataxia type 3

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Abstract

Spinocerebellar ataxia type 3 (SCA3) is an adult-onset neurodegenerative disease caused by a polyglutamine expansion in the ataxin-3 (ATXN3) gene. No effective treatment is available for this disorder, other than symptom-directed approaches. Bile acids have shown therapeutic efficacy in neurodegenerative disease models. Here, we pinpointed tauroursodeoxycholic acid (TUDCA) as an efficient therapeutic, improving the motor and neuropathological phenotype of SCA3 nematode and mouse models. Surprisingly, transcriptomic and functional in vivo data showed that TUDCA acts in neuronal tissue through the glucocorticoid receptor (GR), but independently of its canonical receptor, the FXR. TUDCA was predicted to bind to the GR, similarly to corticosteroid molecules. GR levels were decreased in disease-affected brain regions, likely due to increased protein degradation as a consequence of ATXN3 dysfunction, being restored by TUDCA treatment. Analysis of a SCA3 clinical cohort showed intriguing correlations between the peripheral expression of GR and the predicted age at disease onset, in pre-symptomatic subjects, and of FKBP5 expression with disease progression, suggesting this pathway as a potential source of biomarkers for future study. We have established a novel in vivo mechanism for the neuroprotective effects of TUDCA in SCA3, and propose this readily available drug for clinical trials in SCA3 patients.

Authors

Sara Duarte-Silva, Jorge Diogo Da Silva, Daniela Monteiro-Fernandes, Marta Daniela Costa, Andreia Neves-Carvalho, Mafalda Raposo, Carina Soares-Cunha, Joana S. Correia, Gonçalo Nogueira-Gonçalves, Henrique S. Fernandes, Stéphanie Oliveira, Ana Rita Ferreira-Fernandes, Fernando Rodrigues, Joana Pereira-Sousa, Daniela Vilasboas-Campos, Sara Guerreiro, Jonas Campos, Liliana Meireles-Costa, Cecilia M.P. Rodrigues, Stephanie Cabantous, Sérgio F. Sousa, Manuela Lima, Andreia Teixeira-Castro, Patricia Maciel

A membrane-associated phosphoswitch in Rad controls adrenergic regulation of cardiac calcium channels

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Abstract

The ability to fight or flee from a threat relies upon an acute adrenergic surge that augments cardiac output, which is dependent upon increased cardiac contractility and heart rate. This cardiac response depends on β-adrenergic-initiated reversal of the small RGK G-protein Rad-mediated inhibition of voltage-gated calcium channels (CaV) acting through the Cavβ subunit. Here, we investigate how Rad couples phosphorylation to augmented Ca2+ influx and increased cardiac contraction. We show that reversal requires phosphorylation of Ser272 and Ser300 within Rad’s polybasic, hydrophobic C-terminal domain (CTD). Phosphorylation of Ser25 and Ser38 in Rad’s N-terminal domain (NTD) alone is ineffective. Phosphorylation of Ser272 and Ser300 or the addition of four Asp to the CTD reduces Rad’s association with the negatively charged, cytoplasmic plasmalemmal surface and with CaVβ, even in the absence of CaVα, measured here by FRET. Addition of a post-translationally prenylated CAAX motif to Rad’s C-terminus, which constitutively tethers Rad to the membrane, prevents the physiological and biochemical effects of both phosphorylation and Asp-substitution. Thus, dissociation of Rad from the sarcolemma, and consequently from CaVβ, is sufficient for sympathetic up-regulation of Ca2+ currents.

Authors

Arianne Papa, Pedro J. del Rivero Morfin, Bi-Xing Chen, Lin Yang, Alex N. Katchman, Sergey I. Zakharov, Guoxia Liu, Michael S. Bohnen, Vivian Zheng, Moshe Katz, Suraj Subramaniam, Joel A. Hirsch, Sharon Weiss, Nathan Dascal, Arthur Karlin, Geoffrey S. Pitt, Henry M. Colecraft, Manu Ben Johny, Steven O. Marx

Reversible synaptic adaptations in a subpopulation of murine hippocampal neurons following early-life seizures

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Abstract

Early-life seizures (ELS) can cause permanent cognitive deficits and network hyperexcitability, but it is unclear whether ELS induce persistent alterations to specific neuronal populations and if these changes can be targeted to mitigate network dysfunction. We used the targeted recombination of activated populations (TRAP) approach to genetically label neurons activated by kainate-induced ELS in immature mice. The ELS-TRAPed neurons were mainly found in hippocampal CA1, remained uniquely susceptible to reactivation by later-life seizures, and displayed sustained enhancement in α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor (AMPAR)-mediated excitatory synaptic transmission and inward rectification. ELS-TRAPed neurons, but not non-TRAPed surrounding neurons, exhibited enduring decreases in Gria2 mRNA, responsible for encoding the GluA2 subunit of the AMPARs. This was paralleled by decreased synaptic GluA2 protein expression and heightened phosphorylated GluA2 at Ser880 in dendrites, indicative of GluA2 internalization. Consistent with increased GluA2-lacking AMPARs, ELS-TRAPed neurons showed premature silent synapse depletion, impaired long-term potentiation, and impaired long-term depression. In vivo post-seizure treatment with IEM-1460, a GluA2-lacking AMPAR inhibitor, markedly mitigated ELS-induced alterations in TRAPed neurons. These findings show that enduring modifications of AMPARs occur in a subpopulation of ELS-activated neurons, contributing to synaptic dysplasticity and network hyperexcitability, but are reversible with early IEM-1460 intervention.

Authors

Bo Xing, Aaron J. Barbour, Joseph Vithayathil, Xiaofan Li, Sierra Dutko, Jessica Fawcett-Patel, Eunjoo Lancaster, Delia M. Talos, Frances E. Jensen