Biology subjects

Hermine, O.

Publications and source records attributed to Hermine, O..

14 recordsLinked to original sources

bioRxiv2026

Masitinib is an oral, brain penetrant inhibitor of microglial and mast cell activity with neuroprotective potential in progressive forms of multiple sclerosis

IntroductionProgressive multiple sclerosis (MS), including primary progressive MS (PPMS) and non-active secondary progressive MS (nSPMS), remains an unmet need, as few treatments target innate immune pathways. Masitinib (AB1010) is a selective tyrosine kinase inhibitor that targets c-Kit and colony-stimulating factor 1 receptor pathways. This mechanism disrupts mast cell-microglia interactions, key innate immune effectors in progressive MS pathogenesis, reducing neuroinflammation and neuronal damage. In the phase 3 AB07002 trial, masitinib (4.5 mg/kg/d) over 96 weeks met its primary endpoint. Comparable signals in PPMS and nSPMS indicated masitinib benefited both phenotypes. Secondary analyses showed that masitinib lowered the progression to wheelchair dependence (EDSS [≥]7.0, 12 weeks) and reduced the 12-week confirmed EDSS progression risk by 37% versus placebo. MethodsThis study aimed to confirm that oral masitinib achieves central nervous system (CNS) concentrations sufficient to modulate CSF1R and wild-type c-Kit, thereby underpinning its neuroprotective potential. Male Sprague Dawley rats (n=12, [~]200 g) were administered a single oral dose (30 mg/kg). Plasma and brain samples were collected at 2, 4, 8, and 24 hours post-dose (n=3 per time point). Masitinib (AB1010) and its metabolite (AB3280) were quantified in plasma and brain homogenates using LC-MS/MS. ResultsMasitinib reached a brain Cmax of 223.5 ng/mL ([~]450 nM), exceeding IC50 values for CSF1R and wild-type c-KIT by [~]5-fold and 2-fold, respectively, indicating effective CNS target engagement. The active metabolite AB3280 also achieved brain Cmax levels with full inhibitory activity. Masitinib demonstrated consistent CNS penetration supported by a proportional plasma-to-brain exposure relationship. ConclusionMasitinibs favorable CNS penetration and safety profile, alongside its unique mast cell inhibition, position it as a compelling candidate for progressive MS treatment, either as monotherapy or in combination with other agents. This multifaceted immunomodulatory approach addresses critical unmet needs in progressive MS and supports further clinical development.

neuroscience↗

bioRxiv2025

AXL mediates mast cell survival and resistance to tyrosine kinase inhibitors in mastocytosis

Mastocytosis is a clonal disorder driven by KIT mutations, but resistance to tyrosine kinase inhibitors (TKIs) remains a major challenge. Following the discovery of an AXL L197M mutation in a patient with congenital aggressive mastocytosis, we demonstrated unexpected wild-type AXL expression in neoplastic mast cells (MCs) across mastocytosis subtypes, challenging current views concerning mastocytosis pathophysiology. AXL was undetectable in steady-state MCs but several factors, including IFN- and IFN-{beta}, induced its expression, consistent with the inflammatory nature of mastocytosis and the high interferon levels in patient plasma. Ectopic expression of WT or L197M AXL in the ROSA KIT D816V cell line enhanced proliferation and survival by upregulating pSTAT5, pSTAT3, pFAK, p-p38, survivin and BCL2. Both AXL forms conferred resistance to the KIT inhibitor PKC412/midostaurin by sustaining BCL2, MCL1, and BCL-XL expression while reducing caspase-3 activation. L197M AXL induced slightly stronger resistance to apoptosis than WT, but this difference was not significant. Combined KIT and AXL targeting (PKC412+R428) restored TKI sensitivity by downregulating BCL-XL, Livin and cIAP1, and activating caspase-3, highlighting the therapeutic potential of dual KIT/AXL pathway inhibition. Importantly, neoplastic MCs from a mast cell leukemia patient harboring the KIT F522C mutation and unresponsive to PKC412 strongly expressed AXL and displayed marked in vitro sensitivity to R428 alone, highlighting AXL as a potential therapeutic target in aggressive mastocytosis not driven by KIT D816V. These findings identify AXL as a previously unrecognized driver of malignant MC survival and TKI resistance, and support AXL inhibition as a promising therapeutic strategy in aggressive mastocytosis. Key Points- AXL is aberrantly expressed in neoplastic mast cells, driving survival and resistance to KIT inhibition in mastocytosis. - Dual KIT and AXL inhibition restores TKI sensitivity in KIT-mutant mastocytosis. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/686205v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1569c9eorg.highwire.dtl.DTLVardef@1445b32org.highwire.dtl.DTLVardef@bf3565org.highwire.dtl.DTLVardef@14a94f0_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Neuropilin-1 expression by solid tumors impairs CAR T cell function

Neuropilin-1 (NRP-1) is a versatile transmembrane protein expressed in numerous cell types and tissues, both in health and in disease. In particular, it is expressed by activated T cells, where it has been shown to play an inhibitory role, dampening their response to tumor cells. Chimeric antigen receptor (CAR) T cells have had considerable success in treating cancers such as B cell leukemias, but face several obstacles in the treatment of solid tumors. We hypothesized that NRP-1 may contribute to dampening CAR T cell responses to cancer. While NRP-1 blockade either through neutralizing antibodies or through CRISPR/Cas9-mediated deletion did not improve CAR T cell cytotoxic activity in an in vitro solid tumor model, NRP-1 was found to have a protective effect against CAR T cells when expressed by the tumor cells, in line with previous literature implicating NRP-1 as a pro-tumor factor involved in their growth and invasiveness. We found that NRP-1 was transferred from target cells to CAR T cells via trogocytosis, i.e. the "nibbling" and subsequent display of membrane proteins from one cell by another. CD19, the target antigen in this model, was also transferred to CAR T cells, raising interesting questions about trogocytosis as a marker for effective tumor cell killing through direct interaction. On the contrary, trogocytosis may impede effective CAR T function by promoting fratricide due to their display of target antigen. While no conclusive mechanisms for NRP-1 activity were found, several avenues of research have been opened up to further our understanding of the multifaceted roles of NRP-1 in CAR T cell activity and interactions with their targets.

immunology↗

bioRxiv2025

CLONAL LYMPHOCYTE EXPANSIONS AND JAK-STAT PATHWAY MUTATIONS DEFINE A PATHOGENIC CONTINUUM DRIVING RESISTANCE TO GLUTEN-FREE DIET IN CELIAC DISEASE

Background&AimsDespite recent advances, refractory celiac disease (RCD) poses challenging questions. In type 2 RCD (RCD2), the lack of response to the gluten-free diet is attributed to an intestinal intraepithelial lymphoma carrying driver JAK1 or STAT3 mutations. However, it remains unclear whether these can be safely targeted for therapy. In RCD1, pathogenic insights are still lacking. MethodsDuodenal biopsies and peripheral blood mononuclear cells (PBMCs) from patients with RCD1, RCD2, active CeD, CeD in remission, and controls were analyzed. Lymphocyte populations were characterized using single-cell transcriptomic, genomic, and TCR repertoire profiling. Functional and exome sequencing analyses were performed on patient-derived RCD2 cell lines exposed to JAK inhibitors. ResultsWe show that clonal malignant RCD2 lymphocytes exhibit interpatient similarities but substantial intratumoral heterogeneity, and provide in vitro evidence that JAK inhibitors can select drug-resistant tumor cells, arguing against their use as monotherapy. In RCD1, we identified clonal T-cell expansions harboring mutations that enhance the JAK-STAT pathway. The detection of both RCD2 and a CD4 lymphoproliferation in a patient initially diagnosed with RCD1 further illustrates the diversity of lymphoproliferative outcomes in celiac disease. ConclusionsThese findings suggest that RCD subtypes may share underlying mechanisms driven by clonal evolution and JAK-STAT dysregulation. They also highlight the potential limitations of JAK inhibitor monotherapy and the importance of molecularly informed therapeutic strategies. What You Need to KnowO_ST_ABSBACKGROUND AND CONTEXTC_ST_ABSRefractory celiac disease (RCD) can lead to intestinal lymphoma, but the biological processes driving immune cell transformation and therapy resistance remain incompletely understood. NEW FINDINGSSingle-cell analyses reveal clonal evolution, JAK-STAT pathway dysregulation, and shared molecular features between RCD subtypes, with implications for disease progression and treatment response. LIMITATIONSSample size and reliance on ex vivo models limit generalizability; further in vivo validation of resistance mechanisms is needed to confirm therapeutic implications. CLINICAL RESEARCH RELEVANCEThese findings suggest that RCD subtypes may share underlying mechanisms driven by clonal evolution and JAK-STAT dysregulation. They also highlight the potential limitations of JAK inhibitor monotherapy and the importance of molecularly informed therapeutic strategies. BASIC RESEARCH RELEVANCEThis work uncovers mechanisms of immune cell transformation in chronic intestinal inflammation and provides insight into how tumor heterogeneity and somatic mutations drive disease progression and therapeutic resistance. Lay summaryThis study reveals shared mechanisms in refractory celiac disease subtypes driven by clonal evolution and JAK-STAT activation, highlighting limits of JAK inhibitor monotherapy and the need for personalized treatments.

cancer biology↗

bioRxiv2025

Therapeutic potential of oxygen through M101 transportation in reducing vaso-occlusion and hemolysis in a mouse model of sickle cell disease.

Sickle cell disease (SCD) is a genetic disorder marked by hemoglobin S-induced red blood cell (RBC) sickling, causing hemolysis, vaso-occlusion, and organ damage. Despite its severity, treatment options remain limited. RBC transfusion is a highly effective therapeutic option in SCD, particularly due to its capacity to improve oxygen delivery and reduce sickling by introducing healthy red blood cells, but it is associated with a significant risk of transfusion-related complications. Our study investigates the treatment potential of M101, a novel hemoglobin-based oxygen carrier derived from the marine worm Arenicola marina, in a mouse model of SCD. M101s unique properties, including its high oxygen-carrying capacity and indirect antioxidant outcome, make it a promising candidate for ameliorating SCD pathophysiology. The study demonstrates that M101 administration significantly reduces RBC sickling and hemolysis in SCD mice. Furthermore, M101 infusion significantly prevents microvascular occlusion in liver and lung and attenuates the inflammatory response after vaso-occlusion induction. These findings highlight M101 strong potential as a novel therapeutic strategy for managing or preventing painful vaso-occlusive crises in SCD. Further research is warranted to determine the optimal dosing regimen and to evaluate the long-term safety and efficacy of M101 in humans.

pathology↗

bioRxiv2024

VEXAS anemia is a mosaic erythroblastopenia

VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) is a recently discovered autoinflammatory disorder linked to somatic mutations in the UBA1 gene, resulting in a profound cytoplasm-restricted defect in ubiquitylation. The disease is characterized by a macrocytic anemia that remains poorly understood. To investigate the erythroid lineage in VEXAS, we conducted a comprehensive study combining in vivo assessments of patients mature red cells and marrow erythroblasts, alongside in vitro base-editing models of erythropoiesis. Here we show that mature red cells do not exhibit ubiquitylation defects, and patient-derived bone marrow erythroblasts lack UBA1 mutations beyond the basophilic stage of erythroid differentiation. In vitro base editing of UBA1 variants in CD34+ primary cells resulted in high mortality during early erythroid differentiation, but not during monocytic differentiation. Edited erythroid precursors displayed TP53 overexpression linked to defective ubiquitylation and anomalies in ribosome biogenesis, reminiscent of Diamond-Blackfan anemia. We propose that VEXAS-associated anemia should be considered as a mosaic erythroblastopenia, where the severity of anemia is influenced by the quality and quantity of the UBA1-WT compartment. These insights may aid clinicians in tailoring treatment strategies.

cell biology↗

bioRxiv2024

Proteostasis and metabolic dysfunction in a distinct subset of storage-induced senescent erythrocytes targeted for clearance

Although refrigerated storage slows the metabolism of volunteer donor RBCs, cellular aging still occurs throughout this in vitro process, which is essential in transfusion medicine. Storage-induced microerythrocytes (SMEs) are morphologically-altered senescent RBCs that accumulate during storage and which are cleared from circulation following transfusion. However, the molecular and cellular alterations that trigger clearance of this RBC subset remain to be identified. Using a staining protocol that sorts long-stored SMEs (i.e., CFSEhigh) and morphologically-normal RBCs (CFSElow), these in vitro aged cells were characterized. Metabolomics analysis identified depletion of energy, lipid-repair, and antioxidant metabolites in CFSEhigh RBCs. By redox proteomics, irreversible protein oxidation primarily affected CFSEhigh RBCs. By proteomics, 96 proteins, mostly in the proteostasis family, had relocated to CFSEhigh RBC membranes. CFSEhigh RBCs exhibited decreased proteasome activity and deformability; increased phosphatidylserine exposure, osmotic fragility, and endothelial cell adherence; and were cleared from the circulation during human spleen ex vivo perfusion. Conversely, molecular, cellular, and circulatory properties of long-stored CFSElow RBCs resembled those of short-stored RBCs. CFSEhigh RBCs are morphologically and metabolically altered, have irreversibly oxidized and membrane-relocated proteins, and exhibit decreased proteasome activity. In vitro aging during storage selectively alters metabolism and proteostasis in SMEs, targeting these senescent cells for clearance.

cell biology↗

bioRxiv2024

Masitinib Limits Neuronal Damage, as Measured by Serum Neurofilament Light Chain Concentration, in a Model of Neuroimmune-Driven Neurodegenerative Disease

BackgroundMasitinib is an orally administered tyrosine kinase inhibitor that targets activated cells of the innate neuroimmune system. We have studied the neuroprotective action of masitinib on the manifestations of experimental autoimmune encephalitis (EAE) induced axonal and neuronal damage. EAE is a model of neuroimmune-driven chronic neuroinflammation and therefore highly relevant to masitinibs mechanism of action in neurodegenerative diseases. Importantly, neuronal damage, or prevention thereof, can be rapidly assessed by measuring serum neurofilament light chain (NfL) concentration in EAE-induced mice. MethodsEAE induction was performed in healthy female C57BL/6 mice via active MOG35-55 peptide immunization. Treatments were initiated 14 days post EAE induction. On day-0, 39 mice with established EAE symptoms were randomly assigned to 3 treatment groups (n=13): EAE control, masitinib 50 mg/kg/day (M50), and masitinib 100 mg/kg/day (M100). Treatment started on day-1 and ended on day-15. Blood samples were collected at day-1, day-8 (via tail vein sampling) and day-15 (via intracardiac puncture). Assessments included quantification of serum NfL levels along the disease duration, cytokine quantification at day-15, and clinical assessments. ResultsMasitinib treatment significantly (p<0.0001) limited NfL production with respect to control; specifically, relative change in serum NfL concentration at day-8 was 43% and 60% lower for the M50 and M100 groups, respectively. Likewise, for the assessment of absolute serum NfL at day-8 and day-15, there was a significantly lower NfL concentration for masitinib treatment as compared with control. Furthermore, EAE mice treated with masitinib showed significantly lower concentrations of several well-established pro-inflammatory cytokines relative to control at day-15. A beneficial effect of masitinib on functional performance was also observed, with both M50 and M100 groups showing significantly less relative deterioration in grip strength at day-15 as compared with control (p<0.001). ConclusionThis study is the first demonstration that masitinib, a drug that targets the innate as opposed to the adaptive neuroimmune system, can lower serum NfL levels, and by extension therefore, neuronal damage, in a neuroimmune-driven neurodegenerative disease model. Overall, findings indicate that masitinib has a neuroprotective effect under conditions of chronic neuroinflammation and therefore plausible disease-modifying activity across a broad range of neurodegenerative diseases.

neuroscience↗

bioRxiv2022

Splenic clearance of rigid erythrocytes as an inherited mechanism for splenomegaly and natural resistance to malaria

In malaria-endemic areas, subjects from specific groups like Fulani have a peculiar protection against malaria, with high levels of IgM but also frequent anemia and splenomegaly. The mechanisms underlying this phenotype remain elusive. In Benin, West Africa, we measured the deformability of circulating erythrocytes in genetically distinct groups (including Fulani) living in sympatry, using ektacytometry and microsphiltration, a mimic of how the spleen clears rigid erythrocytes. Compared to non-Fulani, Fulani displayed a higher deformability of circulating erythrocytes, pointing to an enhanced clearance of rigid erythrocytes by the spleen. This phenotype was observed in individuals displaying markers of Plasmodium falciparum infection. The heritability of this new trait was high, with a strong multigenic component. Five of the top 10 genes selected by a population structure-adjusted GWAS, expressed in the spleen, are potentially involved in splenic clearance of erythrocytes (CHERP, MB, PALLD, SPARC, PDE10A), through control of vascular tone, collagen synthesis and macrophage activity. In specific ethnic groups, genetically-controlled processes likely enhance the innate retention of infected and uninfected erythrocytes in the spleen, explaining splenomegaly, anemia, cryptic intrasplenic parasite loads, hyper-IgM, and partial protection against malaria. Beyond malaria-related phenotypes, inherited splenic hyper-filtration of erythrocytes may impact the pathogenesis of other hematologic diseases. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSThe genetic background of individuals influences their susceptibility to infectious diseases. Specific human groups, like the Fulani in Africa, react to malaria parasites (named Plasmodium) in a specific way. Upon infection, Fulani develop a grossly enlarged spleen, and high levels of anti-Plasmodium antibodies in their blood. They also carry smaller numbers of parasites in their blood, and thus are considered partially protected against malaria. The mechanisms underlying this natural protection, different from other natural protective mechanisms such as the sickle cell trait, are not well understood. Malaria impairs the deformability of red blood cells and the spleen is a key organ to controlling red blood cell quality. We have recently demonstrated that red blood cells containing live malaria parasites accumulate intensely in the spleen of subjects with long term exposure to these parasites. Enhanced retention of infected and uninfected red blood cells in the spleen would explain why the spleen is larger and why lower numbers of parasites are left in circulation. We thus explored whether the retention of infected and uninfected red blood cells could explain why Fulani are partially protected against malaria. Because it is unethical to perform spleen puncture or biopsies for research purposes, our explorations were indirect by carefully analyzing the properties of circulating red blood cells in a large number of subjects and by assessing whether observations could be explained by their genetic make-up. Added value of this studyIn more than 500 subjects, we confirmed the high frequency of large spleens in Fulani and, through 2 different methods, we demonstrated an enhanced deformability of their circulating red blood cells, that likely stems from the more efficient removal of the less deformable ones. This enhanced deformability was found to be inheritable based on carefully collected family links and refined analysis of genetic markers. Implications of all the available evidenceOur findings indicate that genes potentially driving the filtration of red blood cells by the spleen likely influence how subjects in specific groups in Africa and elsewhere react to malaria. While most previous hypotheses pointed to conventional immunological mechanisms as the trigger, we propose that a simple physiological mechanism that controls the quality of red blood cells may drive natural protection from malaria even before the intervention of immunological cells. A better understanding of these processes is of great importance in the context of malaria elimination efforts. These findings may also have an impact on the understanding of other red blood cell-related disorders, such as inherited red cell diseases, in which splenic filtration of abnormal red blood cells may precipitate splenic complications.

physiology↗

bioRxiv2022

Mast cell-mediated inflammation relies on insulin-regulated aminopeptidase controlling cytokine export from the Golgi

Upon activation, mast cells rapidly release preformed inflammatory mediators from large cytoplasmic granules via regulated exocytosis. This acute degranulation is followed by a late activation phase involving synthesis and secretion of cytokines, growth factors and other inflammatory molecules via the constitutive pathway that remains ill-defined. Here we describe a role for an insulin-responsive vesicle-like endosomal compartment, marked by insulin-regulated aminopeptidase (IRAP), in the secretion of TNF-- and IL-6 in mast cells and macrophages. IRAP-deficient mice are protected from TNF--dependent kidney injury and inflammatory arthritis. In the absence of IRAP, TNF- fails to be efficiently exported from the Golgi. Subsequently, reduced co-localization of VAMP3+ endosomes with Stx4 was observed, while VAMP8-dependent exocytosis of secretory granules was facilitated. Chemical targeting of IRAP+ endosomes reduced pro-inflammatory cytokine secretion thereby highlighting this compartment as a promising target for the therapeutic control of inflammation.

immunology↗

bioRxiv2021

Serotonin regulates hepcidin expression via a gut-liver axis

Iron is essential to key biological processes of all living organisms. Proper iron levels must be maintained to meet biological needs and prevent toxicity. Given the central role played by the hormone hepcidin in systemic iron homeostasis, extensive research has sought to identify regulators of its expression. Diverse evidence shows the gut to be an essential sensor and regulator of iron homeostasis, independently of other known hepcidin regulators, including bone marrow signals. Here we identify gut-derived serotonin as a key physiological factor in hepcidin regulation. In response to hypoxia, serotonin synthesized and secreted by enterochromaffin cells can act beyond the gut to repress hepcidin expression in the liver, through a 5-HT2B receptor-dependent pathway. Bone marrow transplant experiments clearly indicate the gut is responsible for hepcidin repression. This regulatory system appears to be conserved in humans: a significant negative correlation exists between hepcidin and serotonin levels in the serum of healthy individuals. Our findings imply hepcidin regulation by serotonin is a physiological process, and modulation of the gut serotonergic system may have broad therapeutic implications.

physiology↗

bioRxiv2020

Oncogenetic Landscape Of Lymphomagenesis In Coeliac Disease

ObjectiveEnteropathy-associated T-cell lymphoma (EATL) is a rare but severe complication of celiac disease (CeD), often preceded by low-grade clonal intraepithelial lymphoproliferation, referred to as type II refractory CeD (RCDII). Knowledge on underlying oncogenic mechanisms remains scarce. Here, we analysed and compared the mutational landscape of RCDII and EATL in order to identify genetic drivers of CeD-associated lymphomagenesis. DesignPure populations of RCDII-cells derived from intestinal biopsies (n=9) or sorted from blood (n=2) were analysed by whole exome sequencing, comparative genomic hybridization and RNA-sequencing. Biopsies from RCDII (n=50), EATL (n=19), type I refractory CeD (n=7) and uncomplicated CeD (n=7) were analysed by targeted next-generation sequencing. Moreover, functional in vitro studies and drug testing were performed in RCDII-derived cell lines. Results80% of RCDII and 90% of EATL displayed somatic gain-of-functions mutations in the JAK1-STAT3 pathway, including a remarkable p.G1097 hotspot mutation in the JAK1 kinase-domain in approximately 50% of cases. Other recurrent somatic events were deleterious mutations in NF{kappa}B-regulators TNFAIP3 and TNIP3 and potentially oncogenic mutations in TET2, KMT2D and DDX3X. JAK1 inhibitors and the proteasome inhibitor bortezomib could block survival and proliferation of malignant RCDII-cell lines. ConclusionMutations activating the JAK1-STAT3 pathway appear to be the main drivers of CeD-associated lymphomagenesis. In concert with mutations in negative regulators of NF{kappa}B, they may favour the clonal emergence of malignant lymphocytes in the cytokine-rich coeliac intestine. The identified mutations are attractive therapeutic targets to treat RCDII and block progression towards EATL.

cancer biology↗

bioRxiv2019

A toxic palmitoylation on Cdc42 drives a severe autoinflammatory syndrome

BackgroundAutoinflammatory diseases (AID) result from dysregulation of the first lines of innate immune responses. Recently, development of high throughput genome sequencing technology led to the rapid emergence of important knowledge in the genetic field. About 20 genes have been identified so far in monogenic forms of distinct AID. However, 70-90 % of patients with AID remain without genetic diagnosis.\n\nObjectiveWe report the identification and characterization of a mutation in the C-terminal region of the Rho GTPase Cdc42 in a patient presenting a severe autoinflammatory phenotype.\n\nMethodsWe have analyzed the consequences of the mutation on the subcellular localization of the Cdc42 protein using imaging techniques. Molecular studies were performed using proteomic and biochemical experiments to provide mechanistic bases of the observed defects. Functional assays were also conducted using flow cytometry and cytokine production measurements.\n\nResultsWe show that mutant Cdc42 is trapped in the Golgi apparatus due to the aberrant addition of a palmitate that both enhances the interaction of mutant Cdc42 with Golgi membranes and inhibit its extraction by GDP dissociation inhibitor (GDI), thus impairing its cytosol/membrane shuttling. At the functional level, mutant Cdc42 fails to sustain actin filaments polymerization and induces an exacerbated profile of pro-inflammatory cytokine production due to increased NF-{kappa}B activation.\n\nConclusionsOur study now provides a molecular explanation for mutations that have been identified recently in our AID patient and others in the C-terminal part of Cdc42. Mutations located in this region of Cdc42 impair the intracellular localization of Cdc42, preventing its interaction with the plasma membrane. Thus, our results definitively link mutations in the CDC42 gene to a complex immune-hemato-autoinflammatory phenotype in humans.

genetics↗

bioRxiv2019

Pooled CRISPR Inverse PCR sequencing (PCIP-seq): simultaneous sequencing of retroviral insertion points and the associated provirus in thousands of cells with long reads

Retroviral infections create a large population of cells, each defined by a unique proviral insertion site. Methods based on short-read high throughput sequencing can identify thousands of insertion sites, but the proviruses within remain unobserved. We have developed Pooled CRISPR Inverse PCR sequencing (PCIP-seq), a method that leverages long reads on the Oxford Nanopore MinION platform to sequence the insertion site and its associated provirus. We have applied the technique to three exogenous retroviruses, HTLV-1, HIV-1 and BLV, as well as endogenous retroviruses in both cattle and sheep. The long reads of PCIP-seq improved the accuracy of insertion site identification in repetitive regions of the genome. The high efficiency of the method facilitated the identification of tens of thousands of insertion sites in a single sample. We observed thousands of SNPs and dozens of structural variants within proviruses and uncovered evidence of viral hypermutation, recombination and recurrent selection.

genomics↗