Research
Abstract
X-linked dominant incontinentia pigmenti (IP) and X-linked recessive anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) are caused by loss-of-function and hypomorphic NEMO mutations, respectively. We describe a European mother with mild IP and a Japanese mother without IP, whose three boys with EDA-ID died of immunodeficiency. We identify the same private variant in an intron of IKBKG/NEMO, IVS4+866 C>T, which was inherited from and occurred de novo in the European and Japanese mothers, respectively. This mutation creates a new splicing donor site, giving rise to a 44-nucleotide pseudo-exon generating a frameshift. Its leakiness accounts for NF-κB activation being impaired, but not abolished in the boys’ cells. However, aberrant splicing rates differ between cell types, with WT NEMO mRNA and protein levels ranging from barely detectable in leukocytes to residual amounts in iPSC-derived macrophages, and higher levels in fibroblasts and iPSC-derived neuronal precursor cells. Finally, SRSF6 binds to the pseudo-exon, facilitating its inclusion. Moreover, SRSF6 knockdown or CLK inhibition restores WT NEMO expression and function in mutant cells. A recurrent deep intronic splicing mutation in IKBKG/NEMO underlies a purely quantitative NEMO defect in males that is most severe in leukocytes and can be rescued by the inhibition of SRSF6 or CLK.
Authors
Bertrand Boisson, Yoshitaka Honda, Masahiko Ajiro, Jacinta Bustamante, Matthieu Bendavid, Andrew R. Gennery, Yuri Kawasaki, Jose Ichishima, Mitsujiro Osawa, Hiroshi Nihira, Takeshi Shiba, Takayuki Tanaka, Maya Chrabieh, Benedetta Bigio, Hong Hur, Yuval Itan, Yupu Liang, Satoshi Okada, Kazushi Izawa, Ryuta Nishikomori, Osamu Ohara, Toshio Heike, Laurent Abel, Anne Puel, Megumu K. Saito, Jean-Laurent Casanova, Masatoshi Hagiwara, Takahiro Yasumi
Abstract
Hereditary angioedema (HAE) is an autosomal dominant disease characterized by recurrent edema attacks associated with morbidity and mortality. HAE results from variations in the SERPING1 gene encoding C1 inhibitor (C1INH), a serine protease inhibitor (serpin). Reduced plasma levels of C1INH lead to enhanced activation of the contact system triggering high levels of bradykinin and increased vascular permeability, but the cellular mechanisms leading to low C1INH levels (20-30% of normal) in heterozygous HAE type I patients remain obscure. Here, we showed that C1INH encoded by a subset of HAE-causing SERPING1 alleles affected secretion of normal C1INH protein in a dominant negative fashion by triggering formation of protein-protein interactions between normal and mutant C1INH leading to creation of larger intracellular C1INH aggregates that were trapped in the endoplasmic reticulum (ER). Notably, intracellular aggregation of C1INH and ER abnormality were observed in fibroblasts from a heterozygous carrier of a dominant negative SERPING1 gene variant, but the condition was ameliorated by viral delivery of the SERPING1 gene. Collectively, our data link abnormal accumulation of serpins, a hallmark of serpinopathies, with dominant negative disease mechanisms affecting C1INH plasma levels in HAE type I patients and may pave the way for new treatments of HAE.
Authors
Didde Haslund, Laura Barrett Ryø, Sara Seidelin Majidi, Iben Kløvgaard Rose, Kristian Alsbjerg Skipper, Tue Fryland, Anja Bille Bohn, Claus Koch, Martin K. Thomsen, Yaseelan Palarasah, Thomas J. Corydon, Anette Bygum, Lene N. Nejsum, Jacob Giehm Mikkelsen
Abstract
The negatively charged sugar sialic acid (Sia) occupies the outermost position in the bulk of cell surface glycans. Lack of sialylated glycans due to genetic ablation of the Sia activating enzyme CMP-sialic acid synthase (CMAS) resulted in embryonic lethality around day 9.5 post coitum (E9.5) in mice. Developmental failure was caused by complement activation on trophoblasts in Cmas-/- implants accompanied by infiltration of maternal neutrophils at the fetal-maternal interface, intrauterine growth restriction, impaired placental development and a thickened Reichert’s membrane. This phenotype, which shared features with complement-recepter-1 related protein Y (Crry) depletion, was rescued in E8.5 Cmas-/- mice upon injection of cobra venom factor resulting in exhaustion of the maternal complement component C3. Here we show that Sia is dispensable for early development of the embryo proper, but pivotal for fetal-maternal immune homeostasis during pregnancy, i.e. for protecting the allograft implant against attack by the maternal innate immune system. Finally, embryos devoid of cell surface sialylation suffered from malnutrition due to inadequate placentation as secondary effect.
Authors
Markus Abeln, Iris Albers, Ulrike Peters-Bernard, Kerstin Flächsig-Schulz, Elina Kats, Andreas Kispert, Stephen Tomlinson, Rita Gerardy-Schahn, Anja Münster-Kühnel, Birgit Weinhold
Abstract
Aldehyde dehydrogenase 2 (ALDH2) is a mitochondrial enzyme detoxifying acetaldehyde and endogenous lipid aldehydes; previous studies suggest a protective role of ALDH2 against cardiovascular disease (CVD). Around 40% of East Asians carrying a single nucleotide polymorphism (SNP) ALDH2 rs671 have increased incidences of CVD. However, the role of ALDH2 in CVD beyond alcohol consumption remains poorly defined. Here we report that ALDH2/LDLR DKO mice have decreased atherosclerosis compared to LDLR KO mice, whereas ALDH2/APOpoE DKO have increased atherosclerosis, suggesting an unexpected interaction of ALDH2 with LDLR. Further studies demonstrate that in the absence of LDLR, AMPK phosphorylates ALDH2 at threonine 356 and enables its nuclear translocation. Nuclear ALDH2 interacts with HDAC3 and represses transcription of a lysosomal proton pump protein ATP6Vv0Ee2, critical for maintaining lysosomal function, autophagy and degradation of oxLDL. Interestingly, an interaction of cytosolic LDLR C-terminus with AMPK blocks ALDH2 phosphorylation and subsequent nuclear translocation, whereas ALDH2 rs671 mutant in human macrophages attenuates this interaction, which releases ALDH2 to nucleus to suppress ATP6Vv0Ee2 expression, resulting in increased foam cells due to impaired lysosomal function. Our studies reveal a novel role of ALDH2 and LDLR in atherosclerosis and provide a molecular mechanism by which ALDH2 rs671 SNP increases CVD.
Authors
Shanshan Zhong, Luxiao Li, Yu-Lei Zhang, Lili Zhang, Jianhong Lu, Shuyuan Guo, Ningning Liang, Jing Ge, Mingjiang Zhu, Yongzhen Tao, Yun-Cheng Wu, Huiyong Yin
Abstract
Notch signaling regulates differentiation of the pancreatic endocrine lineage during embryogenesis, but the role of Notch in mature β cells is unclear. We found that islets derived from lean mice show modest β cell Notch activity, which increases in obesity and in response to high glucose. This response appeared maladaptive, as mice with β cell-specific deficient Notch transcriptional activity (β-Rbpj, β-DNMAML) showed improved glucose tolerance when subjected to high-fat diet feeding. Conversely, mice with β cell-specific expression of constitutively-active Notch1 (β-NICD) had a progressive loss of β cell maturity, due to proteasomal degradation of MafA, leading to impaired glucose-stimulated insulin secretion and glucose intolerance with aging or obesity. Surprisingly, Notch-active β cells had increased proliferative capacity, leading to increased but dysfunctional β cell mass. These studies demonstrate a dynamic role for Notch in developed β cells to simultaneously regulate β cell function and proliferation.
Authors
Alberto Bartolome, Changyu Zhu, Lori Sussel, Utpal B. Pajvani
Abstract
Levothyroxine (LT4) is a form of thyroid hormone used to treat hypothyroidism. In the brain, T4 is converted to the active form T3 by the type 2 deiodinase (D2). Thus, it is intriguing that carriers of the Thr92Ala polymorphism in the D2 gene (DIO2) exhibit clinical improvement when liothyronine (LT3) is added to LT4 therapy. Here we report that D2 is a cargo protein in endoplasmic reticulum Golgi intermediary compartment (ERGIC) vesicles, recycling between ER and Golgi. The Thr92 to Ala substitution (Ala92-D2) caused ER stress and activated the unfolded protein response (UPR); Ala92-D2 accumulated in the trans-Golgi and generated less T3, all of which was restored by eliminating ER stress with the chemical chaperone 4-phenyl butyric acid (4-PBA). An Ala92-Dio2 polymorphism-carrying mouse exhibited UPR and hypothyroidism in distinct brain areas. The mouse refrained from physical activity, slept more and required additional time to memorize objects. Enhancing T3 signaling in the brain with LT3 improved cognition, whereas restoring proteostasis with 4-PBA eliminated the Ala92-Dio2 phenotype. In contrast, primary hypothyroidism intensified the Ala92-Dio2 phenotype, with only partial response to LT4 therapy. Disruption of cellular proteostasis and reduced Ala92-D2 activity may explain the failure of LT4 therapy in carriers of Thr92Ala-DIO2.
Authors
Sungro Jo, Tatiana L. Fonseca, Barbara M.L. Da Costa Bocco, Gustavo W. Fernandes, Elizabeth A. McAninch, Anaysa P. Bolin, Rodrigo R. Da Conceição, Joao Pedro W.S. De Castro, Daniele L. Ignacio, Péter Egri, Dorottya Németh, Csaba Fekete, Maria Martha Bernardi, Victoria D. Leitch, Naila S. Mannan, Katharine F. Curry, Natalie C. Butterfield, J.H. Duncan Bassett, Graham R. Williams, Balázs Gereben, Miriam O. Ribeiro, Antonio C. Bianco
Abstract
Iron-related disorders are among the most prevalent diseases worldwide. Systemic iron homeostasis requires hepcidin, a hepatic-derived hormone that controls iron mobilization through its molecular target, ferroportin (FPN), the only known mammalian iron exporter. This pathway is perturbed in diseases that cause iron overload. Additionally, intestinal HIF-2α is essential for the local absorptive response to systemic iron deficiency and iron overload. Our data demonstrate a hetero-tissue crosstalk mechanism, where liver hepcidin regulated intestinal HIF-2α in iron deficiency, anemia, and iron overload. We show that FPN controlled cell autonomous iron efflux to regulate the activity of iron-dependent, intestinal prolyl hydroxylase domain enzymes to stabilize HIF-2α. Pharmacological blockade of HIF-2α using a clinically relevant and highly specific inhibitor successfully treated iron overload in a mouse model. These findings demonstrate a molecular link between liver hepcidin and intestinal HIF-2α that controls physiological iron uptake and drives iron hyperabsorption during iron overload.
Authors
Andrew J. Schwartz, Nupur K. Das, Sadeesh K. Ramakrishnan, Chesta Jain, Mladen Jurkovic, Jun Wu, Elizabeta Nemeth, Samira Lakhal-Littleton, Justin A. Colacino, Yatrik M. Shah
Abstract
The pathogenesis of ischemic diseases remains unclear. Here we demonstrate the induction of microRNA-668 (mir-668) in ischemic acute kidney injury (AKI) in human patients, mice, and renal tubular cells. The induction was HIF-1dependant as HIF-1-deficiency in cells and kidney proximal tubules attenuated mir-668 expression. We further identified a functional HIF-1 binding site in mir-668 gene promoter. Anti-mir-668 increased apoptosis in renal tubular cells and enhanced ischemic AKI in mice, whereas mir-668 mimic was protective. Mechanistically, anti-mir-668 induced mitochondrial fragmentation, whereas mir-668 blocked mitochondrial fragmentation during hypoxia. We analyzed mir-668 target genes through immunoprecipitation of microRNA-induced silencing complexes followed by RNA deep sequencing and identified 124 protein-coding genes as likely targets of mir-668. Among these genes, only Mitochondrial Protein 18 KDa (MTP18) has been implicated in mitochondrial dynamics. In renal cells and mouse kidneys, mir-668 mimic suppressed MTP18, whereas anti-mir-668 increased MTP18 expression. Luciferase microRNA target reporter assay further verified MTP18 as a direct target of mir-668. In renal tubular cells, knockdown of MTP18 suppressed mitochondrial fragmentation and apoptosis. Together, the results suggest that mir-668 is induced via HIF-1 in ischemic AKI and, upon induction, mir-668 represses MTP18 to preserve mitochondrial dynamics for renal tubular cell survival and kidney protection.
Authors
Qingqing Wei, Haipeng Sun, Shuwei Song, Yong Liu, Pengyuan Liu, Man J. Livingston, Jianwen Wang, Mingyu Liang, Qing-Sheng Mi, Yuqing Huo, N. Stanley Nahman, Changlin Mei, Zheng Dong
Abstract
Mutations in CDCA7 and HELLS that respectively encode a CXXC-type zinc finger protein and a SNF2 family chromatin remodeler cause immunodeficiency, centromeric instability, facial anomalies (ICF) syndrome type 3 and 4, respectively. Here, we demonstrate that classical non-homologous end joining (C-NHEJ) proteins Ku80 and Ku70, as well as HELLS coimmunoprecipitated with CDCA7. The coimmunoprecipitation of the repair proteins was sensitive to nuclease treatment and an ICF3 mutation in CDCA7 that impairs its chromatin binding. The functional importance of these interactions was strongly suggested by the compromised C-NHEJ activity and significant delay in Ku80 accumulation at DNA damage sites in CDCA7 and HELLS deficient HEK293 cells. Consistent with the repair defect, these cells displayed increased apoptosis, abnormal chromosome segregation, aneuploidy, centrosome amplification, and significant accumulation of γH2AX signals. Although less prominent, cells mutated for the other ICF genes DNMT3B and ZBTB24 (responsible for ICF type 1 and 2, respectively) showed similar defects. Importantly, lymphoblastoid cells from ICF patients shared the same changes detected in the mutant HEK293 cells to varying degrees. Although the C-NHEJ defect alone did not cause CG hypomethylation, CDCA7 and HELLS are involved in maintaining CG methylation at centromeric and pericentromeric repeats. The defect in C-NHEJ may account for some common features of ICF cells, including centromeric instability, abnormal chromosome segregation, and apoptosis.
Authors
Motoko Unoki, Hironori Funabiki, Guillaume Velasco, Claire Francastel, Hiroyuki Sasaki
Abstract
Glioblastoma is highly enriched with macrophages, and osteopontin (OPN) expression levels correlate with glioma grade and the degree of macrophage infiltration, thus we studied whether OPN plays a crucial role in immune modulation. Quantitative PCR, immune blotting, and ELISA were used to determine OPN expression. Knockdown of OPN was achieved using complementary siRNA, shRNA and CRISPR/CAS9 techniques followed by a series of in vitro functional migration and immunological assays. OPN gene-deficient mice were used to examine the roles of non-tumor-derived OPN on survival of mice harboring intracranial gliomas. Patients with mesenchymal GBM show high OPN expression, a negative survival prognosticator. OPN is a potent chemokine for macrophages, and its blockade significantly impaired the ability of glioma cells to recruit macrophages. Integrin αVβ5 (ITGαVβ5) is highly expressed on glioblastoma-infiltrating macrophages and constitutes a major OPN receptor. OPN maintains the M2 macrophage gene signature and phenotype. Both tumor-derived OPN and host-derived OPN was critical for glioma development. OPN deficiency in either the innate immune or glioma cells demonstrated a marked reduction of M2 macrophages and elevated T cell effector activity infiltrating the glioma. Furthermore, OPN deficiency in the glioma cells sensitized them to direct CD8+ T cell cytotoxicity. OPN can be exploited as an immune modulatory target, with efficacious therapeutic results using systemically administered OPN-4-1BB bispecific aptamers, increasing median survival time by 68% (P < 0.05). OPN is an important chemokine for recruiting macrophages to glioblastoma, mediates crosstalk between tumor cells and the innate immune system, and can be exploited as a therapeutic target.
Authors
Jun Wei, Anantha Marisetty, Brett Schrand, Konrad Gabrusiewicz, Yuuri Hashimoto, Martina Ott, Zacharia Grami, Ling-Yuan Kong, Xiaoyang Ling, Hillary G. Caruso, Shouhao Zhou, Y. Alan Wang, Gregory N. Fuller, Jason T. Huse, Eli Gilboa, Nannan Kang, Xingxu Huang, Roel Verhaak, Shulin Li, Amy B. Heimberger
No posts were found with this tag.
Copyright © 2026 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)