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  • Journal article
    Blight J, Sala K, Atcheson E, Kramer H, El-Turabi A, Coelho Real E, Dahalan F, Bettenbourt P, Dickinson-Craig E, Alves E, Salman A, Janse C, Ashcroft F, Hill A, Reyes-Sandoval A, Blagborough A, Baum Jet al., 2021,

    Dissection-independent production of a Plasmodium sporozoites from whole mosquitoes

    , Life Science Alliance, ISSN: 2575-1077
  • Journal article
    Bonnardel F, Haslam SM, Dell A, Feizi T, Liu Y, Tajadura-Ortega V, Akune Y, Sykes L, Bennett PR, MacIntyre DA, Lisacek F, Imberty Aet al., 2021,

    Proteome-wide prediction of bacterial carbohydrate-binding proteins as a tool for understanding commensal and pathogen colonisation of the vaginal microbiome

    , npj Biofilms and Microbiomes, Vol: 7, Pages: 1-10, ISSN: 2055-5008

    Bacteria use carbohydrate-binding proteins (CBPs), such as lectins and carbohydrate-binding modules (CBMs), to anchor to specific sugars on host surfaces. CBPs in the gut microbiome are well studied, but their roles in the vagina microbiome and involvement in sexually transmitted infections, cervical cancer and preterm birth are largely unknown. We established a classification system for lectins and designed Hidden Markov Model (HMM) profiles for data mining of bacterial genomes, resulting in identification of >100,000 predicted bacterial lectins available at unilectin.eu/bacteria. Genome screening of 90 isolates from 21 vaginal bacterial species shows that those associated with infection and inflammation produce a larger CBPs repertoire, thus enabling them to potentially bind a wider array of glycans in the vagina. Both the number of predicted bacterial CBPs and their specificities correlated with pathogenicity. This study provides new insights into potential mechanisms of colonisation by commensals and potential pathogens of the reproductive tract that underpin health and disease states.

  • Journal article
    Gisriel C, Azai C, Cardona Londono T, 2021,

    Recent advances in the structural diversity of reaction centers

    , Photosynthesis Research, ISSN: 0166-8595
  • Journal article
    Byrne B, Saouros S, Mohan T, Cecchetti C, lehmann S, barritt J, Scull N, Simpson P, Alguel Y, cameron A, jones Aet al., 2021,

    Structural and functional insights into the mechanism of action of plant borate transporters

    , Scientific Reports, ISSN: 2045-2322
  • Journal article
    Weinzierl ROJ, 2021,

    Molecular dynamics simulations of human FOXO3 reveal intrinsically disordered regions spread spatially by intramolecular electrostatic repulsion

    , Biomolecules, Vol: 11, Pages: 1-16, ISSN: 2218-273X

    The human transcription factor FOXO3 (a member of the ‘forkhead’ family of transcription factors) controls a variety of cellular functions that make it a highly relevant target for intervention in anti-cancer and anti-aging therapies. FOXO3 is a mostly intrinsically disordered protein (IDP). Absence of knowledge of its structural properties outside the DNA-binding domain constitutes a considerable obstacle to a better understanding of structure/function relationships. Here, I present extensive molecular dynamics (MD) simulation data based on implicit solvation models of the entire FOXO3/DNA complex, and accelerated MD simulations under explicit solvent conditions of a central region of particular structural interest (FOXO3120–530). A new graphical tool for studying and visualizing the structural diversity of IDPs, the Local Compaction Plot (LCP), is introduced. The simulations confirm the highly disordered nature of FOXO3 and distinguish various degrees of folding propensity. Unexpectedly, two ‘linker’ regions immediately adjacent to the DNA-binding domain are present in a highly extended conformation. This extended conformation is not due to their amino acid composition, but rather is caused by electrostatic repulsion of the domains connected by the linkers. FOXO3 is thus an IDP present in an unusually extended conformation to facilitate interaction with molecular interaction partners.

  • Journal article
    Campos M, Willis K, Rona LDP, Christophides G, Maccallum Ret al., 2021,

    Unravelling population structure heterogeneity within the genome of the malaria vector Anopheles gambiae

    , BMC Genomics, ISSN: 1471-2164
  • Journal article
    Kotta-Loizou I, 2021,

    Mycoviruses and their role in fungal pathogenesis.

    , Curr Opin Microbiol, Vol: 63, Pages: 10-18

    Nowadays, the focus of mycovirology research has expanded from plant pathogenic fungi and mycovirus mediated hypovirulence to include insect and human pathogenic fungi together with a range of mycovirus mediated phenotypes, such as hypervirulence, control of endophytic traits, regulation of metabolite production and drug resistance. In fungus-mycovirus-environmental interactions, the environment and both abiotic and biotic factors play crucial roles in whether and how mycovirus mediated phenotypes are manifest. Mycovirus infections result in alterations in the host transcriptome profile, via protein-protein interactions and triggering of antiviral RNA silencing in the fungus. These alterations, in combination with the environmental factors, may result in desirable phenotypic traits for the host, for us and in some cases for both.

  • Journal article
    Ng BG, Sosicka P, Fenaille F, Harroche A, Vuillaumier-Barrot S, Porterfield M, Xia Z-J, Wagner S, Bamshad MJ, Vergnes-Boiteux M-C, Cholet S, Dalton S, Dell A, Dupré T, Fiore M, Haslam SM, Huguenin Y, Kumagai T, Kulik M, McGoogan K, Michot C, Nickerson DA, Pascreau T, Borgel D, Raymond K, Warad D, University of Washington Center for Mendelian Genomics UW-CMG, Flanagan-Steet H, Steet R, Tiemeyer M, Seta N, Bruneel A, Freeze HHet al., 2021,

    A mutation in SLC37A4 causes a dominantly inherited congenital disorder of glycosylation characterized by liver dysfunction.

    , Am J Hum Genet, Vol: 108, Pages: 1040-1052

    SLC37A4 encodes an endoplasmic reticulum (ER)-localized multitransmembrane protein required for transporting glucose-6-phosphate (Glc-6P) into the ER. Once transported into the ER, Glc-6P is subsequently hydrolyzed by tissue-specific phosphatases to glucose and inorganic phosphate during times of glucose depletion. Pathogenic variants in SLC37A4 cause an established recessive disorder known as glycogen storage disorder 1b characterized by liver and kidney dysfunction with neutropenia. We report seven individuals who presented with liver dysfunction multifactorial coagulation deficiency and cardiac issues and were heterozygous for the same variant, c.1267C>T (p.Arg423∗), in SLC37A4; the affected individuals were from four unrelated families. Serum samples from affected individuals showed profound accumulation of both high mannose and hybrid type N-glycans, while N-glycans in fibroblasts and undifferentiated iPSC were normal. Due to the liver-specific nature of this disorder, we generated a CRISPR base-edited hepatoma cell line harboring the c.1267C>T (p.Arg423∗) variant. These cells replicated the secreted abnormalities seen in serum N-glycosylation, and a portion of the mutant protein appears to relocate to a distinct, non-Golgi compartment, possibly ER exit sites. These cells also show a gene dosage-dependent alteration in the Golgi morphology and reduced intraluminal pH that may account for the altered glycosylation. In summary, we identify a recurrent mutation in SLC37A4 that causes a dominantly inherited congenital disorder of glycosylation characterized by coagulopathy and liver dysfunction with abnormal serum N-glycans.

  • Journal article
    Matthews-Palmer T, Gonzalez-Rodriguez N, Calcraft T, Lagercrantz S, Zachs T, Yu X, Grabe G, Holden D, Nans A, Rosenthal P, Rouse S, Beeby Met al., 2021,

    Structure of the cytoplasmic domain of SctV (SsaV) from the Salmonella SPI-2 injectisome and implications for a pH sensing mechanism

    , Journal of Structural Biology, Vol: 213, ISSN: 1047-8477

    Bacterial type III secretion systems assemble the axial structures of both injectisomes and flagella. Injectisome type III secretion systems subsequently secrete effector proteins through their hollow needle into a host, requiring co-ordination. In the Salmonella enterica serovar Typhimurium SPI-2 injectisome, this switch is triggered by sensing the neutral pH of the host cytoplasm. Central to specificity switching is a nonameric SctV protein with an N-terminal transmembrane domain and a toroidal C-terminal cytoplasmic domain. A ‘gatekeeper’ complex interacts with the SctV cytoplasmic domain in a pH dependent manner, facilitating translocon secretion while repressing effector secretion through a poorly understood mechanism. To better understand the role of SctV in SPI-2 translocon-effector specificity switching, we purified full-length SctV and determined its toroidal cytoplasmic region’s structure using cryo-EM. Structural comparisons and molecular dynamics simulations revealed that the cytoplasmic torus is stabilized by its core subdomain 3, about which subdomains 2 and 4 hinge, varying the flexible outside cleft implicated in gatekeeper and substrate binding. In light of patterns of surface conservation, deprotonation, and structural motion, the location of previously identified critical residues suggest that gatekeeper binds a cleft buried between neighboring subdomain 4s. Simulations suggest that a local pH change from 5 to 7.2 stabilizes the subdomain 3 hinge and narrows the central aperture of the nonameric torus. Our results are consistent with a model of local pH sensing at SctV, where pH-dependent dynamics of SctV cytoplasmic domain affect binding of gatekeeper complex.

  • Journal article
    Garrood WT, Kranjc N, Petri K, Kim DY, Guo JA, Hammond AM, Morianou I, Pattanayak V, Joung JK, Crisanti A, Simoni Aet al., 2021,

    Analysis of off-target effects in CRISPR-based gene drives in the human malaria mosquito.

    , Proc Natl Acad Sci U S A, Vol: 118

    CRISPR-Cas9 nuclease-based gene drives have been developed toward the aim of control of the human malaria vector Anopheles gambiae Gene drives are based on an active source of Cas9 nuclease in the germline that promotes super-Mendelian inheritance of the transgene by homology-directed repair ("homing"). Understanding whether CRISPR-induced off-target mutations are generated in Anopheles mosquitoes is an important aspect of risk assessment before any potential field release of this technology. We compared the frequencies and the propensity of off-target events to occur in four different gene-drive strains, including a deliberately promiscuous set-up, using a nongermline restricted promoter for SpCas9 and a guide RNA with many closely related sites (two or more mismatches) across the mosquito genome. Under this scenario we observed off-target mutations at frequencies no greater than 1.42%. We witnessed no evidence that CRISPR-induced off-target mutations were able to accumulate (or drive) in a mosquito population, despite multiple generations' exposure to the CRISPR-Cas9 nuclease construct. Furthermore, judicious design of the guide RNA used for homing of the CRISPR construct, combined with tight temporal constriction of Cas9 expression to the germline, rendered off-target mutations undetectable. The findings of this study represent an important milestone for the understanding and managing of CRISPR-Cas9 specificity in mosquitoes, and demonstrates that CRISPR off-target editing in the context of a mosquito gene drive can be reduced to minimal levels.

  • Journal article
    Netsvetov M, Prokopuk Y, Ivanko I, Kotovych O, Romenskyy Met al., 2021,

    Quercus robur survival at the rear edge in steppe: Dendrochronological evidence

    , Dendrochronologia, Vol: 67, ISSN: 1125-7865

    Climate change is altering forest ecosystems worldwide, particularly in steppe landscapes, where the rare tree communities are challenged with steadily increasing droughts. In the steppe of Eastern Europe, amid dry conditions, Quercus robur occupies mostly riverine habitats and ravines. Here we study the climate sensitivity and drought vulnerability of a Q. robur population located at the rear edge of the species range, in the steppe of Ukraine. The population occupies two adjacent but clearly contrasting in their microclimatic conditions sites: a river floodplain and a steep-sloping river bank. We develop tree earlywood, latewood, and total ring width site-level chronologies and evaluate their relationship with regional climate variables and the local river's water level using response function analysis. We find that trees growing in the floodplain and at the steep river bank have exhibited slightly different growth patterns. The trees at the flooded site have benefited from water proximity, which facilitated their earlywood growth. These trees have responded positively to the current May and previous September precipitation and previous and current May temperatures. At the non-flooded site, the trees have experienced warm temperatures and the lack of precipitation in June. The extreme drought episodes have triggered a decrease in the latewood and total ring width in trees from both sites. We established that oak growth in the floodplain had been depressed by an unknown stressor around 1900, therefore limiting our ability to identify the more beneficial steppe habitat for Q. robur conclusively. Nevertheless, our results provide a dendrochronological evidence of Q. robur survival in a dry steppe environment and lend new insights into local microclimatic factors contributing to it.

  • Journal article
    Roberts LB, Jowett GM, Read E, Zabinski T, Berkachy R, Selkirk ME, Jackson I, Niazi U, Anandagoda N, Araki M, Araki K, Kasturiarachchi J, James C, Enver T, Nimmo R, Reis R, Howard JK, Neves JF, Lord GMet al., 2021,

    MicroRNA-142 Critically Regulates Group 2 Innate Lymphoid Cell Homeostasis and Function

    , JOURNAL OF IMMUNOLOGY, Vol: 206, Pages: 2725-2739, ISSN: 0022-1767
  • Journal article
    Howard SA, Furniss RCD, Bonini D, Amin H, Paracuellos P, Zlotkin D, R D Costa T, Levy A, A I Mavridou D, Filloux Aet al., 2021,

    The breadth and molecular basis of Hcp-driven type six secretion system (T6SS) effector delivery

    , mBio, Vol: 12, Pages: 1-19, ISSN: 2150-7511

    The type VI secretion system (T6SS) is a bacterial nanoscale weapon that delivers toxins into prey ranging from bacteria and fungi to animal hosts. The cytosolic contractile sheath of the system wraps around stacked hexameric rings of Hcp proteins, which form an inner tube. At the tip of this tube is a puncturing device comprising a trimeric VgrG topped by a monomeric PAAR protein. The number of toxins a single system delivers per firing event remains unknown, since effectors can be loaded on diverse sites of the T6SS apparatus, notably the inner tube and the puncturing device. Each VgrG or PAAR can bind one effector, and additional effector cargoes can be carried in the Hcp ring lumen. While many VgrG- and PAAR-bound toxins have been characterized, to date, very few Hcp-bound effectors are known. Here, we used 3 known Pseudomonas aeruginosa Hcp proteins (Hcp1 to -3), each of which associates with one of the three T6SSs in this organism (H1-T6SS, H2-T6SS, and H3-T6SS), to perform in vivo pulldown assays. We confirmed the known interactions of Hcp1 with Tse1 to -4, further copurified a Hcp1-Tse4 complex, and identified potential novel Hcp1-bound effectors. Moreover, we demonstrated that Hcp2 and Hcp3 can shuttle T6SS cargoes toxic to Escherichia coli. Finally, we used a Tse1-Bla chimera to probe the loading strategy for Hcp passengers and found that while large effectors can be loaded onto Hcp, the formed complex jams the system, abrogating T6SS function.

  • Journal article
    Oliver T, Sanchez-Baracaldo P, Larkum AW, Rutherford AW, Cardona Londono Tet al., 2021,

    Time-resolved comparative molecular evolution of oxygenic photosynthesis

    , BBA: Bioenergetics, Vol: 1862, Pages: 1-20, ISSN: 0005-2728

    Oxygenic photosynthesis starts with the oxidation of water to O2, a light-driven reaction catalysed by photosystem II. Cyanobacteria are the only prokaryotes capable of water oxidation and therefore, it is assumed that the origin of oxygenic photosynthesis is a late innovation relative to the origin of life and bioenergetics. However, when exactly water oxidation originated remains an unanswered question. Here we use phylogenetic analysis to study a gene duplication event that is unique to photosystem II: the duplication that led to the evolution of the core antenna subunits CP43 and CP47. We compare the changes in the rates of evolution of this duplication with those of some of the oldest well-described events in the history of life: namely, the duplication leading to the Alpha and Beta subunits of the catalytic head of ATP synthase, and the divergence of archaeal and bacterial RNA polymerases and ribosomes. We also compare it with more recent events such as the duplication of Cyanobacteria-specific FtsH metalloprotease subunits and the radiation leading to Margulisbacteria, Sericytochromatia, Vampirovibrionia, and other clades containing anoxygenic phototrophs. We demonstrate that the ancestral core duplication of photosystem II exhibits patterns in the rates of protein evolution through geological time that are nearly identical to those of the ATP synthase, RNA polymerase, or the ribosome. Furthermore, we use ancestral sequence reconstruction in combination with comparative structural biology of photosystem subunits, to provide additional evidence supporting the premise that water oxidation had originated before the ancestral core duplications. Our work suggests that photosynthetic water oxidation originated closer to the origin of life and bioenergetics than can be documented based on phylogenetic or phylogenomic species trees alone.

  • Journal article
    Mullish BH, Ghani R, McDonald JAK, Davies F, Marchesi JRet al., 2021,

    Reply to Woodworth, et al

    , Clinical Infectious Diseases, Vol: 72, Pages: e924-e925, ISSN: 1058-4838
  • Journal article
    Cadoux M, Caruso S, Pham S, Gougelet A, Pophillat C, Riou R, Loesch R, Colnot S, Nguyen CT, Calderaro J, Celton-Morizur S, Guerra N, Zucman-Rossi J, Desdouets C, Couty J-Pet al., 2021,

    Expression of NKG2D ligands is downregulated by beta-catenin signalling and associates with HCC aggressiveness

    , JOURNAL OF HEPATOLOGY, Vol: 74, Pages: 1386-1397, ISSN: 0168-8278
  • Journal article
    Gonzalo X, Broda A, Drobniewski F, Larrouy-Maumus Get al., 2021,

    Performance of lipid fingerprint-based MALDI-ToF for the diagnosis of mycobacterial infections

    , Clinical Microbiology and Infection, Vol: 27, Pages: 912.e1-912.e5, ISSN: 1198-743X

    ObjectivesBacterial diagnosis of mycobacteria is often challenging because of the variability of the sensitivity and specificity of the assay used, and it can be expensive to perform accurately. Although matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) has become the workhorse of clinical laboratories, the current MALDI methodology (which is based on cytosolic protein profiling) for mycobacteria is still challenging due to the number of steps involved (up to seven) and potential biosafety concerns. Knowing that mycobacteria produce surface-exposed species-specific lipids, we here hypothesized that the detection of those molecules could offer a rapid, reproducible and robust method for mycobacterial identification.MethodsWe evaluated the performance of an alternative methodology based on characterized species-specific lipid profiling of intact bacteria, without any sample preparation, by MALDI MS; it uses MALDI-time-of-flight (ToF) MS combined with a specific matrix (super-2,5-dihydroxybenzoic acid solubilized in an apolar solvent system) to analyse lipids of intact heat-inactivated mycobacteria. Cultured mycobacteria are heat-inactivated and loaded directly onto the MALDI target followed by addition of the matrix. Acquisition of the data is done in both positive and negative ion modes. Blinded studies were performed using 273 mycobacterial strains comprising both the Mycobacterium tuberculosis (Mtb) complex and non-tuberculous mycobacteria (NTMs) subcultured in Middlebrook 7H9 media supplemented with 10% OADC (oleic acid/dextrose/catalase) growth supplement and incubated for up to 2 weeks at 37°C.ResultsThe method we have developed is fast (<10 mins) and highly sensitive (<1000 bacteria required); 96.7% of the Mtb complex strains (204/211) were correctly assigned as MTB complex and 91.7% (22/24) NTM species were correctly assigned based only on intact bacteria species-specific lipid profiling by MALDI-ToF MS.ConclusionsIntact bacter

  • Journal article
    Koneru SL, Hintze M, Katsanos D, Barkoulas Met al., 2021,

    Cryptic genetic variation in a heat shock protein modifies the outcome of a mutation affecting epidermal stem cell development in C. elegans.

    , Nat Commun, Vol: 12

    A fundamental question in medical genetics is how the genetic background modifies the phenotypic outcome of mutations. We address this question by focusing on the seam cells, which display stem cell properties in the epidermis of Caenorhabditis elegans. We demonstrate that a putative null mutation in the GATA transcription factor egl-18, which is involved in seam cell fate maintenance, is more tolerated in the CB4856 isolate from Hawaii than the lab reference strain N2 from Bristol. We identify multiple quantitative trait loci (QTLs) underlying the difference in phenotype expressivity between the two isolates. These QTLs reveal cryptic genetic variation that reinforces seam cell fate through potentiating Wnt signalling. Within one QTL region, a single amino acid deletion in the heat shock protein HSP-110 in CB4856 is sufficient to modify Wnt signalling and seam cell development, highlighting that natural variation in conserved heat shock proteins can shape phenotype expressivity.

  • Journal article
    Casadio R, Lenhard B, Sternberg MJE, 2021,

    Computational Resources for Molecular Biology 2021

    , JOURNAL OF MOLECULAR BIOLOGY, Vol: 433, ISSN: 0022-2836
  • Journal article
    Real E, Howick VM, Dahalan FA, Witmer K, Cudini J, Andradi-Brown C, Blight J, Davidson MS, Dogga SK, Reid AJ, Baum J, Lawniczak MKNet al., 2021,

    A single-cell atlas of Plasmodium falciparum transmission through the mosquito.

    , Nat Commun, Vol: 12

    Malaria parasites have a complex life cycle featuring diverse developmental strategies, each uniquely adapted to navigate specific host environments. Here we use single-cell transcriptomics to illuminate gene usage across the transmission cycle of the most virulent agent of human malaria - Plasmodium falciparum. We reveal developmental trajectories associated with the colonization of the mosquito midgut and salivary glands and elucidate the transcriptional signatures of each transmissible stage. Additionally, we identify both conserved and non-conserved gene usage between human and rodent parasites, which point to both essential mechanisms in malaria transmission and species-specific adaptations potentially linked to host tropism. Together, the data presented here, which are made freely available via an interactive website, provide a fine-grained atlas that enables intensive investigation of the P. falciparum transcriptional journey. As well as providing insights into gene function across the transmission cycle, the atlas opens the door for identification of drug and vaccine targets to stop malaria transmission and thereby prevent disease.

  • Journal article
    Chatterjee S, Lekmeechai S, Constantinou N, Grzybowska EA, Kozik Z, Choudhary JS, Berger CN, Frankel G, Clements Aet al., 2021,

    The type III secretion system effector EspO of enterohaemorrhagic Escherichia coli inhibits apoptosis through an interaction with HAX-1.

    , Cell Microbiol

    Many enteric pathogens employ a type III secretion system (T3SS) to translocate effector proteins directly into the host cell cytoplasm, where they subvert signaling pathways of the intestinal epithelium. Here, we report that the anti-apoptotic regulator HS1-associated protein X1 (HAX-1) is an interaction partner of the T3SS effectors EspO of enterohaemorrhagic Escherichia coli (EHEC) and Citrobacter rodentium, OspE of Shigella flexneri and Osp1STYM of Salmonella enterica serovar Typhimurium. EspO, OspE and Osp1STYM have previously been reported to interact with the focal adhesions protein integrin linked kinase (ILK). We found that EspO localizes both to the focal adhesions (ILK localization) and mitochondria (HAX-1 localization), and that increased expression of HAX-1 leads to enhanced mitochondrial localization of EspO. Ectopic expression of EspO, OspE and Osp1STYM protects cells from apoptosis induced by staurosporine and tunicamycin. Depleting cells of HAX-1 indicates that the anti-apoptotic activity of EspO is HAX-1 dependent. Both HAX-1 and ILK were further confirmed as EspO1 interacting proteins during infection using T3SS-delivered EspO1. Using cell detachment as a proxy for cell death we confirmed that T3SS-delivered EspO1 could inhibit cell death induced during EPEC infection, to a similar extent as the anti-apoptotic effector NleH, or treatment with the pan caspase inhibitor z-VAD. In contrast, in cells lacking HAX-1, EspO1 was no longer able to protect against cell detachment, while NleH1 and z-VAD maintained their protective activity. Therefore during both infection and ectopic expression EspO protects cells from cell death by interacting with HAX-1. These results suggest that despite the differences between EHEC, C. rodentium, Shigella and S. Typhimurium infections, hijacking HAX-1 anti-apoptotic signaling is a common strategy to maintain the viability of infected cells. This article is protected by copyright. All rights reserved.

  • Journal article
    McKee KK, Hohenester E, Aleksandrova M, Yurchenco PDet al., 2021,

    Organization of the laminin polymer node

    , Matrix Biology, Vol: 98, Pages: 49-63, ISSN: 0174-173X

    Laminin polymerization is a key step of basement membrane assembly that depends on the binding of α, β and γ N-terminal LN domains to form a polymer node. Nodal assembly can be divided into two steps consisting of β- and γ-LN dimerization followed by calcium-dependent addition of the α-LN domain. The assembly and structural organization of laminin-111 LN-LEa segments was examined by size-exclusion chromatography (SEC) and electron microscopy. Triskelion-like structures were observed in negatively-stained images of purified α1/β1/γ1 LN-LEa trimers. Image averaging of these revealed a heel-to-toe organization of the LN domains with angled outward projections of the LEa stem-like domains. A series of single-amino acid substitutions was introduced into the polymerization faces of the α1, β1 and γ1 LN domains followed by SEC analysis to distinguish between loss of β-γ mediated dimerization and loss of α-dependent trimerization (with intact β-γ dimers). Dimer-blocking mutations were confined to the γ1-toe and the β1-heel, whereas the trimer-only-blocking mutations mapped to the γ1-heel, β1-toe and the α1-toe and heel. Thus, in the polymer node the γ1-toe pairs with the β1-heel, the β1-toe pairs with the α1-heel, and the α1-toe pairs with the γ1-heel.

  • Journal article
    Wang S-S, Solar VD, Yu X, Antonopoulos A, Friedman AE, Agarwal K, Garg M, Ahmed SM, Addhya A, Nasirikenari M, Lau JT, Dell A, Haslam SM, Sampathkumar S-G, Neelamegham Set al., 2021,

    Efficient inhibition of O-glycan biosynthesis using the hexosamine analog Ac(5)GalNTGc

    , CELL CHEMICAL BIOLOGY, Vol: 28, Pages: 699-+, ISSN: 2451-9448
  • Journal article
    Hintze M, Katsanos D, Shahrezaei V, Barkoulas Met al., 2021,

    Phenotypic Robustness of Epidermal Stem Cell Number in C. elegans Is Modulated by the Activity of the Conserved N-acetyltransferase nath-10/NAT10

    , FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY, Vol: 9, ISSN: 2296-634X
  • Journal article
    Yunus IS, Wang Z, Sattayawat P, Muller J, Zemichael FW, Hellgardt K, Jones PRet al., 2021,

    Improved Bioproduction of 1-Octanol Using Engineered Synechocystis sp. PCC 6803.

    , ACS Synth Biol

    1-Octanol has gained interest as a chemical precursor for both high and low value commodities including fuel, solvents, surfactants, and fragrances. By harnessing the power from sunlight and CO2 as carbon source, cyanobacteria has recently been engineered for renewable production of 1-octanol. The productivity, however, remained low. In the present work, we report efforts to further improve the 1-octanol productivity. Different N-terminal truncations were evaluated on three thioesterases from different plant species, resulting in several candidate thioesterases with improved activity and selectivity toward octanoyl-ACP. The structure/function trials suggest that current knowledge and/or state-of-the art computational tools are insufficient to determine the most appropriate cleavage site for thioesterases in Synechocystis. Additionally, by tuning the inducer concentration and light intensity, we further improved the 1-octanol productivity, reaching up to 35% (w/w) carbon partitioning and a titer of 526 ± 5 mg/L 1-octanol in 12 days. Long-term cultivation experiments demonstrated that the improved strain can be stably maintained for at least 30 days and/or over ten times serial dilution. Surprisingly, the improved strain was genetically stable in contrast to earlier strains having lower productivity (and hence a reduced chance of reaching toxic product concentrations). Altogether, improved enzymes and environmental conditions (e.g., inducer concentration and light intensity) substantially increased the 1-octanol productivity. When cultured under continuous conditions, the bioproduction system reached an accumulative titer of >3.5 g/L 1-octanol over close to 180 days.

  • Journal article
    Ferdous Z, Fuchs S, Behrends V, Trasanidis N, Vlachou D, Christophides GKet al., 2021,

    Anopheles coluzziistearoyl-CoA desaturase is essential for adult female survival and reproduction upon blood feeding

    , PLoS Pathogens, ISSN: 1553-7366

    <jats:title>Abstract</jats:title><jats:p>Vitellogenesis and oocyte maturation require anautogenous female<jats:italic>Anopheles</jats:italic>mosquitoes to obtain a bloodmeal from a vertebrate host. The bloodmeal is rich in proteins that are readily broken down into amino acids in the midgut lumen and absorbed by the midgut epithelial cells where they are converted into lipids and then transported to other tissues including ovaries. The stearoyl-CoA desaturase (SCD) plays a pivotal role in this process by converting saturated (SFAs) to unsaturated (UFAs) fatty acids; the latter being essential for maintaining cell membrane fluidity amongst other housekeeping functions. Here, we report the functional and phenotypic characterization of SCD1 in the malaria vector mosquito<jats:italic>Anopheles coluzzii</jats:italic>. We show that RNA interference (RNAi) silencing of<jats:italic>SCD1</jats:italic>and administration of sterculic acid (SA), a small molecule inhibitor of SCD1, significantly impact on the survival and reproduction of female mosquitoes following blood feeding. Microscopic observations reveal that the mosquito thorax is quickly filled with blood, a phenomenon likely caused by the collapse of midgut epithelial cell membranes, and that epithelial cells are depleted of lipid droplets and oocytes fail to mature. Transcriptional profiling shows that genes involved in protein, lipid and carbohydrate metabolism and immunity-related genes are the most affected by<jats:italic>SCD1</jats:italic>knock down (KD) in blood-fed mosquitoes. Metabolic profiling reveals that these mosquitoes exhibit increased amounts of saturated fatty acids and TCA cycle intermediates, highlighting the biochemical framework by which the<jats:italic>SCD1</jats:italic>KD phenotype manifests as a result of a detrimental metabolic syndrome. Accumulation of SFAs is also the likely cause of the potent immune response obser

  • Conference paper
    Miguens Blanco J, Liu Z, Mullish BH, Danckert NP, Alexander JL, Chrysostomou D, Sengupta R, McHugh N, McDonald JAK, Abraham SM, Marchesi JRet al., 2021,

    A Phenomic Characterization of the Gut Microbiota - Associations with Psoriatic Arthritis and Ankylosing Spondylitis

    , World Microbe Forum
  • Journal article
    Viola S, Sellés J, Bailleul B, Joliot P, Wollman F-Aet al., 2021,

    In vivo electron donation from plastocyanin and cytochrome c6 to PSI in Synechocystis sp. PCC6803.

    , Biochim Biophys Acta Bioenerg, Vol: 1862

    Many cyanobacteria species can use both plastocyanin and cytochrome c6 as lumenal electron carriers to shuttle electrons from the cytochrome b6f to either photosystem I or the respiratory cytochrome c oxidase. In Synechocystis sp. PCC6803 placed in darkness, about 60% of the active PSI centres are bound to a reduced electron donor which is responsible for the fast re-reduction of P700in vivo after a single charge separation. Here, we show that both cytochrome c6 and plastocyanin can bind to PSI in the dark and participate to the fast phase of P700 reduction, but the fraction of pre-bound PSI is smaller in the case of cytochrome c6 than with plastocyanin. Because of the inter-connection of respiration and photosynthesis in cyanobacteria, the inhibition of the cytochrome c oxidase results in the over-reduction of the photosynthetic electron transfer chain in the dark that translates into a lag in the kinetics of P700 oxidation at the onset of light. We show that this is true both with plastocyanin and cytochrome c6, indicating that the partitioning of electron transport between respiration and photosynthesis is regulated in the same way independently of which of the two lumenal electron carriers is present, although the mechanisms of such regulation are yet to be understood.

  • Journal article
    Da DF, McCabe R, Somé BM, Esperança PM, Sala KA, Blight J, Blagborough AM, Dowell F, Yerbanga SR, Lefèvre T, Mouline K, Dabiré RK, Churcher TSet al., 2021,

    Detection of Plasmodium falciparum in laboratory-reared and naturally infected wild mosquitoes using near-infrared spectroscopy.

    , Scientific Reports, Vol: 11, Pages: 10289-10289, ISSN: 2045-2322

    There is an urgent need for high throughput, affordable methods of detecting pathogens inside insect vectors to facilitate surveillance. Near-infrared spectroscopy (NIRS) has shown promise to detect arbovirus and malaria in the laboratory but has not been evaluated in field conditions. Here we investigate the ability of NIRS to identify Plasmodium falciparum in Anopheles coluzzii mosquitoes. NIRS models trained on laboratory-reared mosquitoes infected with wild malaria parasites can detect the parasite in comparable mosquitoes with moderate accuracy though fails to detect oocysts or sporozoites in naturally infected field caught mosquitoes. Models trained on field mosquitoes were unable to predict the infection status of other field mosquitoes. Restricting analyses to mosquitoes of uninfectious and highly-infectious status did improve predictions suggesting sensitivity and specificity may be better in mosquitoes with higher numbers of parasites. Detection of infection appears restricted to homogenous groups of mosquitoes diminishing NIRS utility for detecting malaria within mosquitoes.

  • Journal article
    Weber DK, Reddy UV, Wang S, Larsen EK, Gopinath T, Gustavsson MB, Cornea RL, Thomas DD, De Simone A, Veglia Get al., 2021,

    Structural basis for allosteric control of the SERCA-Phospholamban membrane complex by Ca2+ and phosphorylation.

    , Elife, Vol: 10

    Phospholamban (PLN) is a mini-membrane protein that directly controls the cardiac Ca2+-transport response to β-adrenergic stimulation, thus modulating cardiac output during the fight-or-flight response. In the sarcoplasmic reticulum membrane, PLN binds to the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), keeping this enzyme's function within a narrow physiological window. PLN phosphorylation by cAMP-dependent protein kinase A or increase in Ca2+ concentration reverses the inhibitory effects through an unknown mechanism. Using oriented-sample solid-state NMR spectroscopy and replica-averaged NMR-restrained structural refinement, we reveal that phosphorylation of PLN's cytoplasmic regulatory domain signals the disruption of several inhibitory contacts at the transmembrane binding interface of the SERCA-PLN complex that are propagated to the enzyme's active site, augmenting Ca2+ transport. Our findings address long-standing questions about SERCA regulation, epitomizing a signal transduction mechanism operated by posttranslationally modified bitopic membrane proteins.

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