SPNS1 variants cause multi-organ disease and implicate lysophospholipid transport as critical for mTOR-regulated lipid homeostasis

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Abstract

SPNS1 is a lysosomal transporter mediating the salvage of lysoglycerophospholipids, the degradative products of lysosomal phospholipid catabolism. However, a role of lysolipid transport and salvage in regulating cellular lipid homeostasis and in disease is lacking. Here, we identified two families with biallelic SPNS1 loss-of-function variants that presented primarily with progressive liver and striated muscle injury. Patient fibroblasts accumulated lysophospholipids including lysoplasmalogens and cholesterol in lysosomes with reduced cellular plasmalogens. Notably, SPNS1 deficiency resulted in reduced biogenesis of cytosolic lipid droplets containing triglycerides and cholesteryl esters. Mechanistically, we found that lysophospholipids transported by SPNS1 into the cytosol quantitatively contributed to triglyceride synthesis while lysosomal buildup of lyso-ether-phospholipid inhibited lysosomal cholesterol egress, effects that were enhanced with inhibition of mTOR. These findings support a gene-disease association and reveal connectivity between lysosomal transport of lysophospholipids and storage of reserve cellular energy as triglyceride and in the regulation of cholesterol homeostasis, processes that become important under nutrient limitation.

Authors

Menglan He, Mei Ding, Michaela Chocholouskova, Cheen Fei Chin, Martin Engvall, Helena Malmgren, Matias Wagner, Marlen C. Lauffer, Jacob Heisinger, May Christine V. Malicdan, Valérie Allamand, Madeleine Durbeej, Angelica M. Delgado-Vega, Thomas Sejersen, Ann Nordgren, Federico Torta, David L. Silver

Marburg virus glycoprotein mRNA vaccine is more protective than a virus-like particle-forming mRNA vaccine

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Abstract

Although virus-like particle (VLPs) vaccines were shown to be effective against several viruses, their advantage over vaccines which include envelope protein only is not completely clear, particularly for mRNA-encoded VLPs. We conducted a side-by-side comparison of the immunogenicity and protective efficacy of mRNA vaccines encoding for the Marburg virus (MARV) full-length GP delivered alone or as a VLP. Electron microscopy confirmed VLP formation when MARV GP and matrix protein VP40 co-expressed. We vaccinated guinea pigs with a two-component mRNA vaccine encoding for GP and VP40 (VLP) or GP alone. At the highest dose, both vaccines protected fully, although the VLP vaccine elicited a slightly lower humoral response than the GP-only group. However, at low doses, GP-only mRNA conferred 100% protection, whereas the VLP exhibited only partial protection. In mice, VLP mRNA induced a moderate preference for GP-specific CD8+ T cells responses, whereas the GP-only mRNA somewhat favored CD4+ T cell responses. Guinea pig whole blood RNA-seq revealed that the VLP vaccine down-regulated genes associated with various biological and metabolic processes, including the NF-κB signaling pathway, whereas the GP-only vaccine upregulated interferon signaling. Overall, the VLP mRNA vaccine was less immunogenic and protective, whereas the GP-only mRNA vaccine conferred robust protection by as little as one µg dose in guinea pigs.

Authors

Chandru Subramani, Michelle N. Meyer, Matthew A. Hyde, Margaret E. Comeaux, Haiping Hao, James E. Crowe Jr., Vsevolod L. Popov, Harshwardhan Thaker, Sunny Himansu, Andrea Carfi, Alexander Bukreyev

Mutations in spliceosomal gene SNW1 cause neurodevelopment disorders with microcephaly

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Abstract

The spliceosome is a critical cellular machinery responsible for pre-mRNA splicing, essential for the proper expression of genes. Mutations in its core components are increasingly linked to neurodevelopmental disorders, such as primary microcephaly. Here, we investigated the role of SNW1, a spliceosomal protein, in splicing integrity and neurodevelopment. We identified nine heterozygous mutations in the SNW1 gene in patients presenting with primary microcephaly. These mutations impaired SNW1's interactions with core spliceosomal proteins, leading to defective RNA splicing and reduced protein functionality. Using Drosophila melanogaster and human embryonic stem cell-derived cerebral organoids models, we demonstrated that SNW1 depletion resulted in significant reductions in neural stem cell proliferation and increased apoptosis. RNA-sequencing revealed disrupted alternative splicing, especially skipping exons, and altered expression of neurodevelopment-associated genes (CENPE, MEF2C, and NRXN2). Our findings provide crucial insights into the molecular mechanisms by which SNW1 dysfunction contributes to neurodevelopmental disorders and underscore the importance of proper spliceosome function in brain development.

Authors

Lei Ji, Jin Yan, Nicole A. Losurdo, Hua Wang, Liangjie Liu, Keyi Li, Zhen Liu, Zhenming Guo, Jing Xu, Adriana Bibo, Decheng Ren, Ke Yang, Yingying Luo, Fengping Yang, Gui Wang, Zhenglong Xiang, Yuan Wang, Huaizhe Zhan, Hu Pan, Juanli Hu, Jianmin Zhong, Rami Abou Jamra, Pia Zacher, Luciana Musante, Flavio Faletra, Paola Costa, Caterina Zanus, Nathalie Couque, Lyse Ruaud, Anna Maria Cueto-González, Hector San Nicolas Fernández, Eduardo Tizzano, Núria Martínez Gil, Xiaorong Liu, Weiping Liao, Layal Abi Farraj, Alden Y. Huang, Liying Zhang, Aparna Murali, Esther Schmuel, Christina S. Han, Kayla King, Weiyue Gu, Pengchao Wang, Kai Li, Nichole Link, Guang He, Shan Bian, Xiao Mao

Tumoral RCOR2 promotes tumor development through dual epigenetic regulation of tumor plasticity and immunogenicity

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Abstract

Gain of plasticity and loss of MHC-II enable tumor cells to evade immune surveillance contributing to tumor development. Here, we showed that the transcriptional corepressor RCOR2 is a key factor that integrates two epigenetic programs surveilling tumor plasticity and immunogenicity. RCOR2 was upregulated predominantly in tumor cells and promoted tumor development in mice through reducing tumor cell death by CD4+/CD8+ T cells and inducing cancer stemness. Mechanistically, RCOR2 repressed RNF43 expression through LSD1-mediated demethylation of histone H3 at lysine 4 to induce activation of Wnt/β-catenin and tumor stemness. Simultaneously, RCOR2 inhibited CIITA expression through HDAC1/2-mediated deacetylation of histone H4 at lysine 16, leading to MHC-II silencing in tumor cells and subsequent impairment of CD4+/CD8+ T cell immunosurveillance, thereby promoting immune evasion. RCOR2 loss potentiated anti-PD-1 therapy in mouse models of cancer and correlated with better response to anti-PD-1 therapy in human patients. Collectively, these findings uncover a “two birds with one stone” effect for RCOR2, highlighting its potential as a valuable target for improved cancer therapy.

Authors

Lei Bao, Ming Zhu, Maowu Luo, Ashwani Kumar, Yan Peng, Chao Xing, Yingfei Wang, Weibo Luo

Cyclin D1 overexpression induces replication stress and microhomology-mediated end-joining dependence in mantle cell lymphoma

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Abstract

Oncogene expression can cause replication stress (RS), leading to DNA double-strand breaks (DSB) that require repair through pathways such as homologous recombination, non-homologous end-joining, and microhomology-mediated end-joining (MMEJ). Cyclin D1 (encoded by CCND1) is a well-known oncoprotein overexpressed in cancer; however, its role in RS is unknown. Using mantle cell lymphoma (MCL) as a naturally occurring model of cyclin D1 overexpression, we examined its impact on RS and DSB-repair mechanisms. Cyclin D1 overexpression elevated RS, increased DNA damage, especially during mitosis, and caused specific upregulation of MMEJ. Furthermore, cyclin D1 activates the polymerase theta (POLQ) transcription by binding its promoter loci, driving POLΘ-mediated MMEJ that is essential to withstand cyclin D1-induced RS. Moreover, concurrent ATM deficiency further intensifies RS, enhances POLQ expression and heightens reliance on MMEJ mediated DNA damage repair. Consequently, inhibition of POLΘ in cyclin D1-overexpressed settings further exacerbates RS, causing single-strand DNA gap accumulations and chromosomal instability, ultimately leading to apoptosis, an effect amplified in ATM-deficient cells. Targeting MMEJ via POLΘ inhibition is, therefore, an effective strategy in the context of cyclin D1 overexpression and ATM deficiency and may provide a unique therapeutic approach for treating MCL and other malignancies characterized by similar alterations.

Authors

Jithma Abeykoon, Shuhei Asada, Guangli Zhu, Yuna Hirohashi, Lisa Moreau, Divya R. Iyer, Sirisha Mukkavalli, Kalindi Parmar, Gabriella Zambrano, Lige Jiang, Dongni Yi, Michelle Manske, Kimberly Gwin, Rebecca L. King, James R. Cerhan, Xiaosheng Wu, Zhenkun Lou, Geoffrey I. Shapiro, Thomas Witzig, Alan D'Andrea

Hepcidin sustains Kupffer cell immune defense against bloodstream bacterial infection via gut-derived metabolites in mice

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Abstract

Bloodstream bacterial infections cause one-third of deaths from bacterial infections, and eradication of circulating bacteria is essential to prevent disseminated infections. We here found that hepcidin, the master regulator of systemic iron homeostasis, affected Kupffer cell (KC) immune defense against bloodstream bacterial infections by modulating the gut commensal bacteria-derived tryptophan derivative indole-3-propionic acid (IPA). Hepcidin deficiency impaired bacterial capture by KCs and exacerbated systemic bacterial dissemination through morphological changes in KCs. Gut microbiota depletion and fecal microbiota transplantation revealed that the gut microbiota mediated the alteration of KCs volume. Mechanistically, hepcidin deficiency led to a decreased abundance of the IPA-producing commensal Lactobacillus intestinalis and a concomitant reduction in the gut-to-liver shuttling of its metabolite IPA. IPA supplementation or Lactobacillus intestinalis colonization restored the KC volume and hepatic immune defense against bloodstream bacterial infection in hepcidin-deficient mice. Moreover, hepcidin levels in patients with bacteremia were associated with days of antibiotic usage and hospitalization. Collectively, our findings described a previously unappreciated role of hepcidin in sustaining KC-mediated hepatic defense against bloodstream bacterial infections through the gut commensal Lactobacillus intestinalis and its tryptophan derivative IPA. More importantly, restoring the crosstalk between the gut microbiota and liver through IPA-inspired therapies may offer a promising strategy for enhancing the host defense against bloodstream bacterial infections in those with low hepcidin levels and a high risk for bacterial infections.

Authors

Yihang Pan, Lihua Shen, Zehua Wu, Xueke Wang, Xiwang Liu, Yan Zhang, Qinyu Luo, Sijin Liu, Xiangming Fang, Qiang Shu, Qixing Chen

Blocking immune checkpoint LAIR1 with antibody blockade or 3-in-1 CAR T cells enhances antitumor response

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Abstract

Tumor-associated macrophages (TAMs) are abundant in the tumor microenvironment (TME) and dampen the immune response, negatively affecting patient survival. Therefore, targeting TAMs could address the limitations of current cancer treatments. However, drug development in this area remains limited. The Leukocyte-associated Immunoglobulin-like Receptor-1 (LAIR1), also called CD305, is prominently expressed on the surface of TAMs. We have uncovered a previously unrecognized immunosuppressive LAIR1 → Factor XIII A (FXIII-A) → Collagen IV pathway across various cancer types. Inhibition of LAIR1, either through knockout (Lair1–/–), antibody blockade (aLAIR1), or a chimeric antigen receptor (CAR) design (3-in-1 CAR by combining tumor targeting, T cell trafficking, and remodeling of the immunosuppressive TME in one CAR construct) provides enhanced antitumor response. LAIR1 inhibition enhances peripheral and intratumoral CD8 memory T-cell populations, induces a phenotypic shift of M2-like Macrophages towards M1, and normalizes tumor collagen IV and structural components in the TME, facilitating effective tumor-T cell interactions and tumor suppression. Enhanced antitumor responses were observed when Lair1–/– or aLAIR1 was used alone or combined with CAR T cells or when the 3-in-1 CAR T cells were used solely in chemotherapy-radiation-PD-1 blockade-resistant tumor models. These findings position LAIR1 inhibition as a promising strategy for cancer immunotherapies.

Authors

Haipeng Tao, Dongjiang Chen, Changlin Yang, Duy T. Nguyen, Georges Abboud, Ruixuan Liu, Tianyi Liu, Avirup Chakraborty, Alicia Y. Hou, Nicole A. Petit, Muhammad Abbas, Robert W. Davis, Janie Zhang, Christina Von Roemeling, Mohammed O. Gbadamosi, Linchun Jin, Tongjun Gu, Tuo Lin, Pengchen Wang, Alfonso Pepe, Diego Ivan Pedro, Hector R. Mendez-Gomez, Chao Xie, Aida Karachi, Frances Weidert, Dan Jin, Chenggang Wang, Kaytora Long-James, Elizabeth K. Molchan, Paul Castillo, John A. Ligon, Ashley P. Ghiaseddin, Elias J. Sayour, Maryam Rahman, Loic P. Deleyrolle, Betty Y.S. Kim, Duane A. Mitchell, W. Gregory Sawyer, Jianping Huang

Iron supplementation alleviates pathologies in a mouse model of facioscapulohumeral muscular dystrophy

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Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is a genetic muscle disease caused by ectopic expression of the toxic protein DUX4, resulting in muscle weakness. However, the mechanism by which DUX4 exerts its toxicity remains unclear. In this study, we observed abnormal iron accumulation in muscles of patients with FSHD and in muscle-specific DUX4-expressing (DUX4-Tg) mice. Treatment with iron chelators, an iron-deficient diet, and genetic modifications inhibiting intracellular uptake of iron did not improve but rather exacerbated FSHD pathology in DUX4-Tg mice. Unexpectedly, however, iron supplementation, either from a high-iron diet or intravenous iron administration, resulted in remarkable improvement in grip strength and running performance in DUX4-Tg mice. Iron supplementation suppressed abnormal iron accumulation and the ferroptosis-related pathway involving increased lipid peroxidation in DUX4-Tg muscle. Muscle-specific DUX4 expression led to retinal vasculopathy, a part of FSHD pathology, which was prevented by iron administration. Furthermore, high-throughput compound screening of the ferroptosis pathway identified drug candidates including Ferrostatin-1 (Fer-1), a potent inhibitor of lipid peroxidation. Treatment with Fer-1 dramatically improved physical function in DUX4-Tg mice. Our findings demonstrate that DUX4-provoked toxicity is involved in the activation of the ferroptosis-related pathway and that supplementary iron could be a promising and readily available therapeutic option for FSHD.

Authors

Kodai Nakamura, Huascar-Pedro Ortuste-Quiroga, Naoki Horii, Shin Fujimaki, Toshiro Moroishi, Keiichi I. Nakayama, Shinjiro Hino, Yoshihiko Saito, Ichizo Nishino, Yusuke Ono

The human glucocorticoid receptor variant rs6190 increases blood cholesterol and promotes atherosclerosis

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Abstract

Elevated cholesterol poses cardiovascular risks. The glucocorticoid receptor (GR) harbors a still undefined role in cholesterol regulation. Here, we report that a coding single nucleotide polymorphism (SNP) in the gene en-coding the GR, rs6190, associated with increased cholesterol in women according to UK Biobank and All Of Us datasets. In SNP-genocopying mice, we found that the SNP enhanced hepatic GR activity to transactivate Pcsk9 and Bhlhe40, negative regulators of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) re-ceptors respectively. In mice, the SNP was sufficient to elevate circulating cholesterol across all lipoprotein frac-tions and the risk and severity of atherosclerotic lesions on the pro-atherogenic hAPOE*2/*2 background. The SNP effect on atherosclerosis was blocked by in vivo liver knockdown of Pcsk9 and Bhlhe40. Also, corti-costerone and testosterone were protective against the mutant GR program in cholesterol and atherosclerosis in male mice, while the SNP effect was additive to estrogen loss in females. Remarkably, we found that the mu-tant GR program was conserved in human hepatocyte-like cells using CRISPR-engineered, SNP-genocopying human induced pluripotent stem cells (hiPSCs). Taken together, our study leverages a non-rare human variant to uncover a novel GR-dependent mechanism contributing to atherogenic risk, particularly in women.

Authors

Hima Bindu Durumutla, April Haller, Greta Noble, Ashok Daniel Prabakaran, Kevin McFarland, Hannah Latimer, Akanksha Rajput, Olukunle Akinborewa, Bahram Namjou-Khales, David Y. Hui, Mattia Quattrocelli

Cingulate retinoic acid signaling regulates neuropathic pain and comorbid anxiodepression via extracellular matrix homeostasis

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Abstract

Neuropathic pain is often comorbid with affective disorders. Synaptic plasticity in anterior cingulate cortex (ACC) is assumed to be a crucial interface for pain perception and emotion. Laminin β1 (LAMB1), a key element of extracellular matrix (ECM) in ACC was recently revealed to convey extracellular alterations to intracellular synaptic plasticity and underlie neuropathic pain and aversive emotion. However, it remains elusive what triggers activity-dependent changes of LAMB1 and ECM remodeling after nerve injury. Here, we uncovered a key role of retinoic acid (RA)/RARB signaling in neuropathic pain and associated anxiodepression via regulation of ECM homeostasis. We showed that nerve injury reduced RA level in the serum and ACC in mice and human, which brought about downregulation of its corresponding receptor, RARB. Overexpressing RARB relieved pain hypersensitivity and comorbid anxiodepression, while silencing RARB exacerbated pain sensitivity and induced anxiodepression. Further mechanistic analysis revealed that RARB maintained ECM homeostasis via transcriptional regulation of LAMB1, reversing abnormal synaptic plasticity and eventually improved neuropathic pain and aversive emotion. Taken together with our previous study, we revealed an intracellular-extracellular-intracellular feedforward regulatory network in modulating pain plasticity. Moreover, we identified cingulate RA/RARB signaling as a promising therapeutic target for treatment of neuropathic pain and associated anxiodepression.

Authors

Zhen-Zhen Li, Wan-Neng Liu, Ke-Xin Liu, Zhi-Wei Dou, Rui Zhao, Yun Chen, Meng-Meng Wang, Tao-Zhi Wang, Fei Wang, Wen-Juan Han, Wen-Guang Chu, Xing-Xing Zheng, Rou-Gang Xie, Hua Yuan, Xiao-Fan Jiang, Xiao-Long Sun, Ceng Luo, Shengxi Wu