Biology subjects

Jansen, M.

Publications and source records attributed to Jansen, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Spatiotemporal profiling of T cell repertoires in Lynch syndrome reveals convergent immune responses through neoplastic progression

Surveillance of individuals with Lynch syndrome (LS), who are carriers of heterozygous inactivating mismatch repair (MMR) gene mutations that predispose to colorectal cancer, enables the study of tumour-immune coevolution during tumour initiation. Somatic loss of MMR activity in the LS colon leads to the accrual of cancer driver mutations, and also of neoantigens likely to generate an adaptive immune response. However, the spatial and temporal immune dynamics in colon and peripheral blood are uncharacterised. Here we show that the neoplastic evolution of mismatch repair deficient (MMRd) clones leads to large-scale and sustained reorganisation of the T-cell repertoire in colon and peripheral blood. We performed dense spatiotemporal profiling of T cell receptor (TCR) repertoires from 109 colonic biopsies (11 tumour samples, 98 normal bowel) from seven individuals, with eight matched blood repertoires. We additionally analysed 13 colorectal polyp repertoires from eight participants, 286 cross sectional blood repertoires from 242 participants (198 LS and 44 non-LS) and 52 longitudinal peripheral blood samples from 26 individuals with LS. We detected tumour-associated T cell clones, some of which were widely present in normal bowel tissue and also detectable in peripheral blood. Sequence similarity analysis revealed conserved TCR architectures across tissue compartments and recurrent clustering patterns between patients, that were not present in non-Lynch controls, consistent with convergent MMRd-associated antigen-driven immune selection. We identified systemic signatures associated with local tumour immune activity and revealed evidence of structured repertoire remodelling before overt malignancy. Longitudinal profiling revealed frequent turnover of low-abundance clonotypes, whereas highly abundant clonotypes persisted for several years. Collectively, our data defines the local and circulating TCR landscape associated with colonic neoplasia in LS, and provides a framework for LS blood-based immune monitoring, early interception and biomarker development.

immunology↗

bioRxiv2026

HIV-1 Tat protects macrophages from killing by the Exoenzyme U from Pseudomonas aeruginosa

People living with HIV (PLWH) have a higher risk of developing other diseases, such as bacterial pneumonia. While Pseudomonas aeruginosa (PAE) is a common cause of pneumonia in humans, PLWHs are infrequently infected by PAE. The reasons for this relative resistance of PLWH to PAE are not known. The most virulent PAE strains produce ExoU which is a key effector of PAE cytotoxicity. Upon injection into the target cell cytosol by the type III secretion system, ExoU binds to PI(4,5)P2 on the inner leaflet of the plasma membrane. Its phospholipase activity then induces a loss of plasma membrane integrity, rapidly leading to cell death. HIV-Tat is secreted by HIV-infected cells, leading to nanomolar concentrations of Tat in the sera of PLWH, even under antiretroviral therapy. Circulating Tat can be endocytosed by uninfected cells, translocate to the cytosol and bind to PI(4,5)P2 at the plasma membrane. In uninfected cells only, Tat is palmitoylated, enabling Tat to become resident on PI(4,5)P2. We found that Tat can interfere with the recruitment of ExoU by PI(4,5)P2, thereby protecting macrophages from PAE toxicity. HIV Tat could therefore be involved in the relative protection of PLWH against the most aggressive PAE isolates and their ExoU type III effector. Tat nevertheless enhances the toxicity of ExoU-deficient PAE strains toward macrophages. ImportancePeople living with HIV (PLWH) are at risk of developing bacterial pneumonia. A widespread cause of pneumonia is Pseudomonas aeruginosa (PAE) but, for unknown reasons, PLWHs are infrequently infected by PAE. A key effector of PAE cytotoxicity is ExoU that is injected by PAE into the target cell cytosol, then binds to PI(4,5)P2 on the inner leaflet of the plasma membrane. The potent phospholipase activity of ExoU then induces a loss of plasma membrane integrity, rapidly leading to cell death. HIV-Tat is present in the serum of PLWH. Circulating Tat is endocytosed by cells, translocates to the cytosol and bind to PI(4,5)P2 at the plasma membrane with a very high affinity. This study indicates that Tat interferes with the recruitment of ExoU by PI(4,5)P2, thereby protecting macrophages from ExoU+ PAE. HIV Tat could therefore be involved in the relative protection of PLWH against the most aggressive PAE isolates.

microbiology↗

bioRxiv2026

Impact of the MX segment on the biogenesis of α7 nACh receptors

Nicotinic acetylcholine receptors (nAChRs) belong to the pentameric ligand-gated ion channel superfamily (pLGICs). Among them, the neuronal homomeric 7 nAChR is highly permeable to calcium and plays critical roles in synaptic transmission, cell signaling, and inflammation modulation. The biogenesis of 7 nAChRs is enhanced by the chaperone proteins RIC-3 and NACHO. Previously, we reported a motif in the 5-HT3A receptor, another pLGIC, involved in RIC-3 modulation. Residues in this motif are conserved and also found within the L1-MX segment of the 7 nACh subunit. We therefore explored the regulatory roles of these conserved residues in the biogenesis of 7 nAChRs using multiple approaches, including heterologous expression in Xenopus laevis oocytes, mutagenesis, pull-down assays, cell-surface labeling, and two-electrode voltage-clamp (TEVC) recordings. We find that synthetic 7 L1-MX peptide interacts with both RIC-3 and NACHO. In particular, conserved residues W330, R332, and L336 in the L1-MX positively regulates the assembly of 7 oligomers and the biogenesis of 7nAChR. In presence of residues W330, R332, and L336, NACHO promotes an assembly of an 7 pentamer which is resistant to strong denaturing conditions. NACHO-promoted 7 pentamer is also resistant to Endo H enzyme. Sensitivity of the pentamer to moderate temperatures (37 {degrees}C, 45 {degrees}C, and 50 {degrees}C) suggests that NACHO stabilizes the pentamer via non-covalent interactions. In contrast, Ala replacements at these residues disrupt the biogenesis and abolish 7 current. NACHO and RIC-3 co-expression yields partial rescue of functional expression for some Ala replacement constructs. SUMMARYThis work identifies regulatory roles of conserved residues W330, R332, and L336 in the biogenesis of 7 nAChR. This discovery positions MX subdomain as a promising target for future drug development that can minimize adverse effects.

neuroscience↗

bioRxiv2026

RIC-3 Interacts Directly with the 5-HT3A Receptor to Mediate Trafficking Across Subcellular Compartments

The serotonin type 3A (5-HT3A) receptor is a pentameric ligand-gated ion channel (pLGIC) in central and peripheral neurons that conducts sodium and potassium ions upon serotonin binding. 5-HT3 receptors modulate neurotransmission and synaptic plasticity, influencing mood, sleep, appetite, and addiction. Disruptions in serotonin signaling are linked to central nervous system disorders, including schizophrenia, anxiety and depression. Clinically, these receptors are targeted by antagonists to treat chemotherapy-induced nausea and vomiting. The functional surface expression of these channels is regulated by the chaperone protein Resistant to Inhibitors of Cholinesterase 3 (RIC-3) that promotes plasma-membrane expression, maturation, and trafficking of 5-HT3A and nicotinic acetylcholine receptors. Our previous work identified a duplicated RIC-3 binding motif within the 5-HT3A intracellular domain (ICD). However, it was unclear whether this interaction reflected native conditions. Here, we used a recombinant 5-HT3A ICD peptide in peptide-resin pull-down assays to investigate RIC-3 Interactions in plasma membrane (PM) fractions from Xenopus oocytes, endoplasmic reticulum (ER) fractions from SH-SY5Y cells, and mouse brain tissue. Across all tested systems, the 5-HT3A ICD peptide specifically bound RIC-3. Furthermore, RIC-3 knockdown (RIC-3 KD) SH-SY5Y cells showed a marked reduction in peptide binding and decreased surface levels of nAChR7 and 5-HT3A receptors. These results demonstrate RIC-3-5-HT3A ICD interaction in native cellular contexts and support a role for RIC-3 in regulating receptor surface expression and neuronal signaling.

neuroscience↗

bioRxiv2025

PKN2 regulates cell-junctions to limit colitis and colon tumour formation

Intestinal barrier failure is a defining feature of inflammatory bowel disease and a key driver of inflammation-associated colorectal cancer. However, the epithelial mechanisms that preserve barrier stability during inflammatory stress remain incompletely defined. Here, we identify the Rho-regulated kinase PKN2 as a non-redundant safeguard of tight junction integrity and a candidate tumour suppressor in colitis-associated colorectal cancer. Conditional PKN2 deletion in mice sensitised the colon to inflammatory injury and significantly increased adenoma formation in an inflammatory cancer model, with tumour burden tightly correlating with colitis severity. Mechanistically, PKN2 localises to tight junctions and is required to stabilise barrier integrity during injury in mouse and organoid models. PKN2 loss induces transcriptional programs conserved in human inflammatory bowel disease, where reduced PKN2 expression associates with increased disease severity and altered therapeutic response. Notably, PKN2 is encoded within genomic loci previously linked to intestinal inflammation susceptibility and tumour suppression in both humans and mice. Our findings identify PKN2 as a functional effector underlying these phenotypes and demonstrate that even heterozygous loss is sufficient to confer heightened sensitivity to inflammatory injury and tumour initiation, positioning PKN2 as a central regulator of tight junction stability that shapes disease severity, treatment response and cancer risk.

cell biology↗

bioRxiv2025

Retargeted adenoviruses for local IgA and CD47 blocker production as a novel cancer therapy

Despite advances in IgG-based cancer immunotherapy, challenges remain in effectively engaging innate immune responses against solid tumors. Here, IgA antibodies hold promise due to their ability to activate neutrophils and macrophages. We present a novel retargeted adenovirus-mediated approach that transforms cancer cells into "biofactories" for localized production of monomeric or dimeric IgA antibodies and a CD47 blocker to potentiate the effect of IgA antibodies. With our approach tumor cells effectively produced IgA antibodies against tumor antigens such as EGFR or EpCAM and a soluble SIRP-Fc fusion protein, which blocks the CD47-SIRP axis. In a perfused tumor-on-a-chip model, locally produced IgA triggered neutrophil- and macrophage-mediated tumor cell killing, further potentiated by SIRP-Fc co-production. In FcRI-transgenic, tumor-bearing mice, intratumoral adenoviral injection induced strong local IgA and SIRP-Fc expression, immune cell infiltration, and more than 50% tumor volume reduction after a single treatment. We found that dimeric IgA exerts stronger effects than monomeric IgA, which is of particular interest since dimeric IgA necessitates a local production approach. Together, these results demonstrate that adenovirus-mediated, tumor-restricted delivery of IgA antibodies and CD47 blockade effectively engages innate immune mechanisms, providing a promising new avenue to enhance cancer immunotherapy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/686998v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@183dd88org.highwire.dtl.DTLVardef@452624org.highwire.dtl.DTLVardef@1cb10e8org.highwire.dtl.DTLVardef@c2fa85_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Root growth in Arabidopsis depends on the amount of glutathione and not the glutathione redox potential

Cell cycle activity and hence plant growth strictly depends on glutathione homeostasis. Despite compelling evidence for this dependency based on glutathione depletion, the cause for the cell cycle arrest had remained unclear. Cell cycle control may depend on either the absolute amount of glutathione or on the glutathione redox potential (EGSH). To unambiguously distinguish those two options, we characterized an allelic series of six Arabidopsis mutants affected in glutamate-cysteine ligase, which catalyses the first step for the biosynthesis of reduced glutathione (GSH). When grown under the same conditions, even mutants with 20% and 50% of wild-type GSH amounts are slightly stunted. The most severely compromised mutants, zir1 and rml1, were crossed with either gr1, which lacks cyto-nuclear glutathione disulfide reductase and was used to induce a pronounced shift in EGSH, or with bir6, which has a diminished glutathione consumption and thus exhibits slightly increased levels of GSH. Based on theoretical considerations, these levels are not expected to shift the EGSH to any significant extent. Our study shows that deleting GR1 in the zir1 or rml1 background does not result in an obvious phenotypic change. By contrast, deleting BIR6 was sufficient to suppress the growth arrest in rml1 and to attenuate the growth in zir1. These findings demonstrate that root growth is dependent on the availability of sufficient amounts of GSH, and not affected by pronounced changes in EGSH. This insight provides a decisive step towards understanding the mechanisms underpinning the proposed role of glutathione in cell cycle and growth control. HighlightCrosses of GSH-deficient Arabidopsis mutants with mutants affected in glutathione reduction or GSH consumption pinpoint the GSH amount rather than the glutathione redox potential as causative for changes in root growth.

plant biology↗

bioRxiv2025

Mechano-activated chondroprogenitors (MACs) drive intrinsic cartilage repair; a process that is arrested in osteoarthritis

Articular cartilage, an avascular, matrix-rich tissue, is thought to have limited repair, thereby contributing to osteoarthritis (OA), the common degenerative disease of joints. Cartilage regeneration does occur, however, in OA joints that are mechanically off-loaded. Here we show that mechanical stress, through release of matrix-bound growth factors, reprogrammes chondrocytes, the primary cells of cartilage, into mechano-activated chondroprogenitors (MACs). Studying OA joint fluid before and after mechanical off-loading, reveals evidence of chronic MAC activity, which switches back to a chondrogenic one when mechanical stress is removed. Taken together, we conclude that OA is a disease of arrested repair in which mechanical stress signals need to be switched off before full repair can occur. This novel paradigm uncovers exciting new treatment opportunities.

cell biology↗

bioRxiv2025

Myeloid cell-mediated killing of B-ALL by CD38 and CD20IgA antibody variants is enhanced by CD47/SIRPα interference

Enhancing myeloid effector cell recruitment may improve immunotherapy by monoclonal antibodies - including that against acute lymphoblastic leukemia (ALL). To assess expression of target antigens in B-ALL, we compared mRNA profiling of 559 patient leukemia samples across 18 molecular subtypes with that of representative cell lines. The latter served as target cells to compare human IgG1 or IgA2 variants against CD19, CD20 or CD38 in antibody-dependent cellular phagocytosis (ADCP) by macrophages and antibody-dependent cell-mediated cytotoxicity (ADCC) by polymorphonuclear leukocytes (PMN). Interestingly, antibodies against broadly expressed CD19 were negligibly effective in mediating ADCP or ADCC. Antibodies against CD20 or CD38, the former variably expressed across subtypes, triggered ADCP by macrophages both as IgG1 and IgA2. However, PMN mediated ADCC against CD20 or CD38 was only observed with IgA2 variants, but not with respective IgG1 antibodies. Blocking the myeloid checkpoint molecule CD47 with a CD47 antibody or a soluble SIRP-Fc fusion protein enhanced ADCP and ADCC by IgA2 antibodies. The binding site for SIRP on CD47 contains an N-terminal pyroglutamate (pGlu), whose formation is catalyzed by glutaminyl-peptide cyclotransferase like (QPCTL). The direct involvement of pGlu in CD47/SIRP interactions was shown by using engineered CD47 variants. Both CD47 and QPCTL were broadly expressed across BCP-ALL subtypes, indicating QPCTL inhibitors as additional therapeutic option. Importantly, the combination of anti-CD38 IgA2 and CD47 blockade was effective against xenografted B-ALL cells in human FcRI (CD89) transgenic (tg) NXG mice. Together, these studies support the combination of anti-CD38 IgA2 with CD47 interference to improve myeloid effector cell recruitment for immunotherapy of B-ALL. Data sharing statementRNA-Seq data from BCP-ALL patients and B cells from healthy donors are available in the European Genome-Phenome Archive (EGA) accession numbers EGAS00001006107 and EGAS00001007305, respectively. Mass spectrometry data of analyzed proteins will be made available after manuscript acceptance on PRIDE - PRoteomics IDEntifications Database. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=174 HEIGHT=200 SRC="FIGDIR/small/669665v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@15220daorg.highwire.dtl.DTLVardef@eace04org.highwire.dtl.DTLVardef@17b87daorg.highwire.dtl.DTLVardef@1142822_HPS_FORMAT_FIGEXP M_FIG C_FIG Key pointsO_LIIgG1 and IgA2 antibodies against CD38 or CD20 were effective in recruiting macrophages for ADCP, but only IgA2 triggered ADCC by PMN C_LIO_LIMyeloid effector cell activation was enhanced by interfering with the CD47/SIRP axis, especially when IgA2 antibodies were applied C_LI

cancer biology↗

bioRxiv2025

Alterations of the composition and spatial organization of the microenvironment following non-dysplastic Barrett's esophagus through progression to cancer

Barretts esophagus (BE), a metaplastic condition that is the only known precursor for esophageal adenocarcinoma (EAC), is relatively common, but progression to cancer is infrequent. BE is inflamed but the contribution of the immune system to the carcinogenic process is unknown. To this end, we contrasted non-dysplastic metaplasia of BE patients, captured when they did not progress (non-progressors), did subsequently, but had not yet progressed (pre-progressors) or had already progressed to EAC (progressors). Using spatial multiplexed 56-protein analysis, serial laser capture microdissection (LCM) RNAseq and shallow whole genome sequencing, we identified prooncogenic immune neighbourhoods and dysregulated immune cell populations predictive of subsequent progression to EAC. Indeed, spatial analysis revealed that M1 macrophages, regulatory natural killer (NK) cells, neutrophils and altered ratios of intraepithelial CD4+ and CD8+ lymphocytes typify tumor microenvironmental (TME) changes associated with cancer initiation. Spatially derived cell-to-cell interactions revealed progression-specific immune cell interaction signatures predominantly involving M1 macrophages NK cells and plasma cells. Furthermore, LCM RNAseq analysis identified gene expression hot signatures enriched in pre-progression and progression samples. Notably, we also observed a correlation between immune cells and copy number alterations in progressor metaplasia. By exposing coordinated changes in the immune cell landscape in patients at high risk of developing EAC, this multi-omic dataset provides novel diagnostic and therapeutic opportunities

cancer biology↗

bioRxiv2025

Syk activation during FcγR-mediated phagocytosis involves Syk palmitoylation and desulfenylation

The non-receptor Spleen tyrosine kinase Syk acts downstream of several receptors of the immune system such as the Fc{gamma}R. Syk is composed of a kinase domain and two SH2 domains that interact with the bi- phosphorylated ITAMs motifs of the Fc{gamma}R upon phagocytosis. This results in the activation of Syk by auto- phosphorylation, triggering phosphorylation of several downstream targets in a process that will culminate in F-actin polymerization and phagocytosis of the IgG-opsonized target. We found that Syk is S-acylated upon phagocytosis by macrophages. Palmitoylation is performed on a single Syk-Cys by the protein S-acyl transferase DHHC5 that specifically associates with Syk upon phagocytosis. Syk palmitoylation is required for Syk localization to the phagocytic cup, Syk phosphorylation/activation, Cdc42 recruitment to the cup, F-actin polymerization and phagocytosis. We also observed that another Syk-Cys residue is modified by sulfenylation. Mutation of the sulfenylated Cys that belongs to a redox-motif inactivated the Syk catalytic activity and phagocytosis. We found that Syk desulfenylation occurs during phagocytosis. Molecular dynamics studies indicated that desulfenylation increased the mobility and exposure of a loop within the Syk interdomain B, likely facilitating phosphorylation of key Syk-Tyr residues by upstream effectors such as Src kinases. We thus propose an original updated model for Syk activation during Fc{gamma}R-mediated phagocytosis that involves both Syk palmitoylation and desulfenylation.

cell biology↗

bioRxiv2025

HIV-1 Tat favors the multiplication of Mycobacterium tuberculosis and toxoplasma by inhibiting clathrin-mediated endocytosis and autophagy

AbstractHIV-1 and Mycobacterium tuberculosis (Mtb) coinfections are a major public health problem but are not well characterized. HIV-1 Tat is secreted by infected cells, generating nanomolar concentrations of Tat in the sera of people living with HIV. Circulating Tat enters cells, binds to PI(4,5)P2 then undergoes palmitoylation, thereby becoming resident on this phosphoinositide. Here, we found that Tat favors the multiplication of Mtb in macrophages. Moreover, Tat renders zebrafish larvae more sensitive to mycobacterial infection. We found that Tat binding to PI(4,5)P2 and palmitoylation enable Tat to inhibit the recruitment of the AP-2 adaptor, thereby inhibiting clathrin-mediated endocytosis and in turn autophagy. This inhibition prevents the degradation of intracellular pathogens such as Mtb and opsonized Toxoplasma gondii, but also of lipid droplets, thereby facilitating the access of these pathogens to lipids. We thus identified a mechanism enabling HIV Tat to favor the multiplication of intracellular pathogens such as Mtb.

microbiology↗

bioRxiv2024

Epigenome and early selection determine the tumour-immune evolutionary trajectory of colorectal cancer

Immune system control is a major hurdle that cancer evolution must circumvent. The relative timing and evolutionary dynamics of subclones that have escaped immune control remain incompletely characterized, and how immune-mediated selection shapes the epigenome has received little attention. Here, we infer the genome- and epigenome-driven evolutionary dynamics of tumour-immune coevolution within primary colorectal cancers (CRCs). We utilise our existing CRC multi-region multi-omic dataset that we supplement with high-resolution spatially-resolved neoantigen sequencing data and highly multiplexed imaging of the tumour microenvironment (TME). Analysis of somatic chromatin accessibility alterations (SCAAs) reveals frequent somatic loss of accessibility at antigen presenting genes, and that SCAAs contribute to silencing of neoantigens. We observe that strong immune escape and exclusion occur at the outset of CRC formation, and that within tumours, including at the microscopic level of individual tumour glands, additional immune escape alterations have negligible consequences for the immunophenotype of cancer cells. Further minor immuno-editing occurs during local invasion and is associated with TME reorganisation, but that evolutionary bottleneck is relatively weak. Collectively, we show that immune evasion in CRC follows a "Big Bang" evolutionary pattern, whereby genetic, epigenetic and TME-driven immune evasion acquired by the time of transformation defines subsequent cancer-immune evolution.

cancer biology↗

bioRxiv2023

Identification of a binding site for bupropion in Gloeobacter violaceus ligand-gated ion channel

Bupropion is an atypical antidepressant and smoking cessation drug which causes adverse effects such as insomnia, irritability, and anxiety. Bupropion inhibits dopamine and norepinephrine reuptake transporters and eukaryotic cation-conducting pentameric ligand-gated ion channels (pLGICs), such as nicotinic acetylcholine (nACh) and serotonin type 3A (5-HT3A) receptors, at clinically relevant concentrations. However, the binding sites and binding mechanisms of bupropion are still elusive. To further understand the inhibition of pLGICs by bupropion, in this work, using a prokaryotic homologue of pLGICs as a model, we examined the inhibitory potency of bupropion in Gloeobacter violaceus ligand-gated ion channel (GLIC), a proton-gated ion channel. Bupropion inhibited proton-induced currents in GLIC with an inhibitory potency of 14.9 {+/-} 2.0 M, comparable to clinically attainable concentrations previously shown to also modulate eukaryotic pLGICs. Using single amino acid substitutions in GLIC and two-electrode voltage-clamp recordings, we further determined a binding site for bupropion in the lower third of the first transmembrane segment M1 at residue T214. The sidechain of M1 T214 together with additional residues of M1 and also of M3 of the adjacent subunit have previously been shown to contribute to binding of other lipophilic molecules like allopregnanolone and pregnanolone. SIGNIFICANCEGLIC, Gloeobacter ligand-gated ion channel, has been extensively used as a model to identify and understand binding sites and mechanisms for cholesterol, neurosteroids, and anesthetics interacting with neurotransmitter-gated ion channels, such as, GABAA receptors and nicotinic acetylcholine receptors. Recently, increasing evidence has revealed that another neurotransmitter-gated ion channel, the serotonin type 3A receptor, binds to an atypical antidepressant, bupropion, at clinically relevant concentrations. Our work here proposes a binding site for bupropion in GLIC and suggests a new approach to characterize binding mechanisms of bupropion in other neurotransmitter-gated ion channels.

neuroscience↗

bioRxiv2023

A single amino acid substitution in MdLAZY1A dominantly impairs shoot gravitropism in Malus

Plant architecture is one of the most important factors that determines crop yield potential and productivity. In apple (Malus), genetic improvement of tree architecture has been challenging due to a long juvenile phase and their growth as complex trees composed of a distinct scion and a rootstock. To better understand the genetic control of apple tree architecture, the dominant weeping growth phenotype was investigated. We report the identification of MdLAZY1A (MD13G1122400) as the genetic determinant underpinning the Weeping (W) locus that largely controls weeping growth in Malus. MdLAZY1A is one of the four paralogs in apple that are most closely related to AtLAZY1 involved in gravitropism in Arabidopsis. The weeping allele (MdLAZY1A-W) contains a single nucleotide mutation c.584T>C that leads to a leucine to proline (L195P) substitution within a predicted transmembrane domain that co-localizes with Region III, one of the five conserved regions in LAZY1-like proteins. Subcellular localization revealed that MdLAZY1A localizes to the plasma membrane and nucleus in plant cells. Over-expressing the weeping allele in apple cultivar Royal Gala (RG) with standard growth habit impaired its gravitropic response and altered the growth to weeping-like. Suppressing the standard allele (MdLAZY1A-S) by RNA interference (RNAi) in RG similarly changed the branch growth direction to downward. Overall, the L195P mutation in MdLAZY1A is genetically causal for weeping growth, underscoring not only the crucial roles of residue L195 and Region III in MdLAZY1A-mediated gravitropic response, but also a potential DNA base editing target for tree architecture improvement in Malus and other crops.

plant biology↗

bioRxiv2022

Mutation Rate Evolution Drives Immune Escape In Mismatch Repair-Deficient Cancer

Mutation rate optimisation drives evolution and immune evasion of bacteria and lentiviral strains, including HIV. Whether evolving cancer lineages similarly adapt mutation rates to increase tumour cell fitness is unknown. Here, by mapping the clonal topography of mismatch repair-deficient (MMRd) colorectal cancer, we show that genomic MMRd mutability co-evolves with neoantigen selection to drive intratumour diversification and immune escape. Mechanistically, we find that microsatellite instability modulates subclonal DNA repair by toggling two hypermutable mononucleotide homopolymer runs in the mismatch repair genes MSH6 and MSH3 (C8 and A8, respectively) through stochastic frameshift switching. Spontaneous mutation and reversion at these evolvability switches modulates subclonal mutation rate, mutation bias, and clonal HLA diversity during MMRd cancer evolution. Combined experimental and simulation studies demonstrate that subclonal immune selection favours incremental MMR mutations. MMRd cancers thus fuel intratumour heterogeneity by adapting subclonal mutation rate and mutation bias to immune selection, revealing a conserved co-evolutionary arms race between neoantigen selection and adaptive genomic mutability. Our work reveals layers of mutational complexity and microsatellite biology in MMRd cancer evolution previously hidden in bulk analyses.

cancer biology↗

bioRxiv2022

Interaction Interface between 5-HT3A serotonin receptor and chaperone RIC-3

Serotonin or 5-hydroxytryptamine receptors type 3 (5-HT3) belong to the family of pentameric ligand-gated ion channels (pLGICs), which also includes other neurotransmitter-gated ion channels such as nicotinic acetylcholine receptors (nAChRs). pLGICs have been long-standing therapeutic targets for psychiatric disorders such as anxiety, schizophrenia, and addiction, and neurological diseases like Alzheimers and Parkinsons disease. Due to structural conservation and significant sequence similarities of pLGICs extracellular and transmembrane domains across the more than 40 subunits found in humans, clinical trials for drug candidates targeting these two domains have been largely hampered by undesired effects mediated by off-subunit modulation. With the present study, we explore the interaction interface of the 5-HT3A intracellular domain (ICD) with the resistance to inhibitors of choline esterase (RIC-3) protein. Previously, we have shown that RIC-3 directly interacts with the ICD of 5-HT3A subunits. Using a sequential deletion approach, we identified the L1-MX segment of the ICD fused to maltose-binding protein as sufficient for the interaction. For the present study, synthetic L1-MX-based peptides, Ala-scanning, and a pull-down assay identified positions W347, R349, and L353 as critical for binding to RIC-3. In complementary studies with full-length 5-HT3A subunits, the identified Ala substitutions reduced the modulation of functional surface expression by co-expression of RIC-3. Additionally, we found and characterized a duplication of the binding motif at the transition between the ICD MA-helix and transmembrane segment M4. Analogous Ala substitutions at W447, R449, and L454 disrupt MAM4-peptide RIC-3 interactions and reduce modulation of functional surface expression. In summary we identify two binding sites for RIC-3 with a shared duplicated motif in 5-HT3A subunits, one in the MX-helix and one at the MAM4-helix transition.

neuroscience↗

bioRxiv2021

The scratch-digging lifestyle of the Permian "microsaur" Batropetes as a model for the exaptative origin of jumping locomotion in frogs

Recent studies have shown that the Triassic stem-frog Triadobatrachus lacked the ability to jump off, but nonetheless had the forelimb strength to withstand the impact of landing from a jump. We propose a hypothesis to resolve this pseudoparadox: the strengthened forelimbs are former adaptations to forelimb-based digging that later made jumping possible by exaptation. Micro-CT data from a skeleton of Batropetes palatinus reveal thin cortical bone, confirming Batropetes as terrestrial. Combining adaptations to walking and digging, confirmed by statistical analyses, Batropetes is thought to have searched for food in leaf litter or topsoil. We interpret Batropetes as having used one forelimb at a time to shove leaf litter aside. Batropetes may thus represent an analog or possibly a homolog of the digging stage that preceded the origin of Salientia. We discuss the possibility of homology with the digging lifestyles of other "microsaurs" and other amphibians.

paleontology↗