Biology subjects

Padalino, G.

Publications and source records attributed to Padalino, G..

7 recordsLinked to original sources

bioRxiv2026

The histone code of love: epigenetics of maturation of gonads in the human blood fluke Schistosoma mansoni

Schistosoma mansoni is a parasitic flatworm that has two, genetically determined, sexes. We used aggregated data of 8 posttranslational histone modifications (ChIP-Seq), chromatin accessibility (ATAC-Seq), transcription (RNA-Seq) and genome feature annotations to decipher the histone code of genes involved in the differentiation of schistosome gonads (i.e. female ovaries and male testes). We show schistosome gonads express at least two classes of protein coding genes: H3K4me3-positive genes that display canonical features of eukaryotic protein-coding genes such as peaks of H3K4me3 at the transcription start sites (TSS) and increases in histone acetylation marks towards the transcription end site (TES), but also a non-canonical H3K9/27me3 plateau just upstream of the TSS. H3K4me3 enrichment at the TSS is highly predictive for transcription strength in these genes compared to a second class of protein coding genes (H3K4me3-negative genes) that do not display this pattern and is characterised by absence of the investigated histone marks at TSS and TES. This is indicative of the existence of hitherto unknown, potentially schistosome-specific histone marks in these genes. The absence of H3K4me3 at the TSS is not associated with inducible or stable gene expression in the gonads. Instead, gene ontology analysis indicates that H3K4me3-positive genes are related to functions which typically govern processes such as metabolism or signal transduction while H3K4me3-negative genes are dedicated to cell communication or immune responses. Second, individual histone modifications and their combinations are associated with functional features of the schistosome genome, known as "chromatin colours". In H3K4me3-positive genes, there is clear co-linearity of 3 colours, which strongly suggests a functional role for histone modifications in the control of transcription pre-initiation, promotor release, and transcription termination. Third, there are striking chromatin structure changes during maturation of the gonads in all genomic features including protein-coding and non-protein coding genes as well as repetitive sequences. The nature of these changes is different in both sexes. H3K36me3 and H3K9me3, as well as H3K23ac and H3K9ac show the strongest variations. Last, we show that pharmacological inhibition of histone demethylation activity by IOX1 leads to a concentration-dependent separation ("divorce") of schistosome couples confirming the importance of H3K36/H3K9 methylation for pairing maintenance and indicating histone demethylases as a potential drug target family. Collectively, our findings offer unprecedented insights into histone codes and chromatin dynamics governing the reproductive development of S. mansoni gonads.

developmental biology↗

bioRxiv2024

2,3-bis (phenylamino) quinoxaline - containing compounds display potent activity against Methicillin-resistant Staphylococcus aureus, Enterococcus faecalis and their biofilms

Antimicrobial resistance remains a global challenge threatening the ability to control diseases caused by bacterial infections. Here, we explored the antimicrobial activity of 2,3-N,N-diphenyl quinoxaline derivatives against representative Gram-positive, Gram-negative and Mycobacterium species. Two quinoxaline derivatives (compounds 25 and 31) demonstrated particularly potent activity against most Staphylococcus aureus, Enterococcus faecium and Enterococcus faecalis strains tested (MIC values between 0.25 to 1 mg/l). These compounds also demonstrated potent antibacterial activity against methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecium/E. faecalis (VRE) strains. Against an extensive panel of clinically relevant isolates, they further showed comparable or better activity to four currently used antibiotics (vancomycin, teicoplanin, daptomycin and linezolid). Finally, they performed better in preventing S. aureus and E. faecalis biofilm formation when compared to several other antibiotics. In conclusion, these two quinoxaline derivatives have promising activities that could be further explored as part of efforts to identify urgently needed new antibacterial agents.

microbiology↗

bioRxiv2024

Quinoxaline-Based Anti-Schistosomal Compounds Have Potent Anti-Malarial Activity

The human pathogens Plasmodium and Schistosoma are each responsible for over 200 million infections annually, being particularly problematic in low- and middle-income countries. There is a pressing need for new drug targets for these diseases, driven by emergence of drug-resistance in Plasmodium and the overall dearth of new drug targets for Schistosoma. Here, we explored the opportunity for pathogen-hopping by evaluating a series of quinoxaline-based anti-schistosomal compounds for activity against P. falciparum. We identified compounds with low nanomolar potency against 3D7 and multidrug-resistant strains. Evolution of resistance using a mutator P. falciparum line revealed a low propensity for resistance. Only one of the series, compound 22, yielded resistance mutations, including point mutations in a non-essential putative hydrolase pfqrp1, as well as copy-number amplification of a phospholipid-translocating ATPase, pfatp2, a potential target. Notably, independently generated CRISPR-edited mutants in pfqrp1 also showed resistance to compound 22 and a related analogue. Moreover, previous lines with pfatp2 copy-number variations were similarly less susceptible to challenge with the new compounds. Finally, we examined whether the predicted hydrolase activity of PfQRP1 underlies its mechanism of resistance, showing that both mutation of the putative catalytic triad and a more severe loss of function mutation elicited resistance. Collectively, we describe a compound series with potent activity against two important pathogens and their potential target in P. falciparum.

microbiology↗

bioRxiv2021

Schistosoma mansoni α-N-acetylgalactosaminidase (SmNAGAL) regulates coordinated parasite movement and egg production

-galactosidase (-GAL) and -N-acetylgalactosaminidase (-NAGAL) are two glycosyl hydrolases responsible for maintaining cellular homeostasis by regulating glycan substrates on proteins and lipids. Mutations in the human genes encoding either enzyme lead to neurological and neuromuscular impairments seen in both Fabry- and Schindler/Kanzaki-diseases. Here, we investigate whether the parasitic blood fluke Schistosoma mansoni, responsible for the neglected tropical disease schistosomiasis, also contains functionally important -GAL and -NAGAL proteins. As infection, parasite maturation and host interactions are all governed by carefully-regulated glycosylation processes, inhibiting S. mansonis -GAL and -NAGAL activities could lead to the development of novel chemotherapeutics. Sequence and phylogenetic analyses of putative -GAL/-NAGAL protein types showed Smp_089290 to be the only S. mansoni protein to contain the functional amino acid residues necessary for -GAL/-NAGAL substrate cleavage. Both -GAL and -NAGAL enzymatic activities were higher in females compared to males (p<0.05; -NAGAL > -GAL), which was consistent with smp_089290s female biased expression. Spatial localisation of smp_089290 revealed accumulation in parenchymal cells, neuronal cells, and the vitellaria and mature vitellocytes of the adult schistosome. siRNA-mediated knockdown (>90%) of smp_089290 in adult worms significantly inhibited -NAGAL activity when compared to control worms (siLuc treated males, p<0.01; siLuc treated females, p<0.05). No significant reductions in -GAL activities were observed in the same extracts. Despite this, decreases in -NAGAL activities correlated with a significant inhibition in adult worm motility as well as in egg production. Programmed CRISPR/Cas9 editing of smp_089290 in adult worms confirmed the egg reduction phenotype. Based on these results, Smp_089290 was determined to act predominantly as an -NAGAL (hereafter termed SmNAGAL) in schistosome parasites where it participates in coordinating movement and oviposition processes. Pharmacological inhibition of SmNAGAL may lead to the development of a novel anthelmintic class of compounds. Author summarySchistosomiasis is a parasitic disease caused by infection with blood flukes, which leads to acute and chronic pathology in millions of infected individuals located in deprived tropical and subtropical regions. Elucidating the function of schistosome genes has provided a clearer view on their roles in various molecular pathways, which are critical to successful parasitism. This information is invaluable when progressing novel drug and vaccine candidates. Here, we add to the existing knowledge of the Schistosoma mansoni parasitic glycan processing and modification machinery by functionally characterising a glycosyl hydrolase (S. mansoni -N-acetylgalactosaminidase, SmNAGAL). We demonstrate that this protein is enzymatically active and important in coordinating parasite movement in adult male and female schistosomes. Additionally, we provide evidence that this protein regulates pathways associated with egg production in female schistosomes, which is responsible for inducing pathological reactions. Developing drugs that inhibit SmNAGAL enzymatic activity could provide a novel approach for controlling schistosomiasis.

molecular biology↗

bioRxiv2020

Schistosoma mansoni lysine specific demethylase 1 (SmLSD1) is a druggable target involved in parasite survival, oviposition and stem cell proliferation.

Schistosomiasis is a chronically-debilitating neglected tropical disease (NTD) that predominantly affects people living in resource-poor communities of tropical and subtropical countries. Schistosoma mansoni, one of three species responsible for most human infections, undergoes strict developmental regulation of gene expression that is carefully controlled by both genetic- and epigenetic- processes. As inhibition of S. mansoni epigenetic machinery components has been shown to impair key transitions throughout the parasites digenetic lifecycle, this knowledge is currently fuelling the search for new epi-drug - based anthelmintics. In this study, the anti-schistosomal activity of 39 re-purposed Homo sapiens Lysine Specific Demethylase 1 (HsLSD1) inhibitors was investigated on key life cycle stages associated with both definitive (schistosomula, juvenile worms, sexually-mature adults) and intermediate host (miracidia) infection. The most active compound (compound 33; e.g. schistosomula phenotype EC50 = 4.370 {micro}M; adult worm motility EC50 = 2.137 {micro}M) was subsequently used to provide further insight into the critical role of S. mansoni lysine specific demethylase 1 (SmLSD1) in adult worm oviposition and stem cell proliferation. Here, compound 33 treatment of adult schistosomes led to significant defects in egg production, intra-egg vitellocyte/ovum packaging and gonadal/neoblast stem cell proliferation. A greater abundance of H3K4me2 marks accompanied these phenotypes and supported specific inhibition of SmLSD1 in adult schistosomes by compound 33. In silico screening indicated that compound 33 likely inhibits SmLSD1 activity by covalently reacting with the FAD cofactor. This work suggests that evaluation of HsLSD1 - targeting epi-drugs could have utility in the search for next-generation anti-schistosomals. The ability of compound 33 to inhibit parasite survival, oviposition, H3K4me2 demethylation and stem cell proliferation warrants further investigations of this compound and its epigenetic target. This data further highlights the importance of histone methylation in S. mansoni lifecycle transitions. Author summaryAffecting over 200 million people, schistosomiasis is a chronic disease caused by the parasitic worm Schistosoma mansoni. The frontline drug for schistosomiasis treatment is praziquantel. Owing to the concern surrounding praziquantel insensitivity or resistance developing, current research is directed towards the identification of novel drugs. We have focused our search for compounds that affect essential aspects of schistosome biology including parasite movement, fertility, cell proliferation and survival. Since all of these functions are potentially influenced by epigenetic regulation of gene expression, we investigated the activity of compounds that alter histone methylation status. In this report, we show that S. mansoni Lysine Specific Demethylase 1 (SmLSD1), a histone demethylase, is critical to miracidia-to-sporocyst transitioning, adult worm motility, egg production and parasite survival. Inhibition of SmLSD1 with compounds developed to inhibit the human paralog show promising potential as novel anti-schistosomal agents.

microbiology↗

bioRxiv2020

Large-scale RNAi screening uncovers new therapeutic targets in the human parasite Schistosoma mansoni

Schistosomes kill 250,000 people every year and are responsible for serious morbidity in 240 million of the world's poorest people. Despite their profound global impact, only a single drug (praziquantel) is available to treat schistosomiasis, highlighting the need to better understand schistosome biology to drive the development of a new generation of therapeutics. A major barrier to this goal is the paucity of large-scale datasets exploring schistosome gene function. Here, we describe the first large-scale RNA interference screen in adult Schistosoma mansoni examining the function of over 2000 genes representing approximately 20 percent of the protein coding genome. More than 250 genes were found to have phenotypes affecting neuromuscular function, tissue integrity, stem cell maintenance, and parasite survival. Leveraging these data, we bioinformatically prioritized several compounds with in vitro activity against parasites and validated p97, a component of the ubiquitin proteasome system, as a drug target in the worm. We further reveal a potentially druggable protein kinase-signaling module involving the TAO and STK25 kinases that are essential for maintaining the transcription of muscle-specific mRNAs. Importantly, loss of either of these kinases results in paralysis and death of schistosomes following surgical transplantation into a mammalian host. We anticipate this work will invigorate studies into the biology of these poorly studied organisms and expedite the development of new therapeutics to treat an important neglected tropical disease.

microbiology↗

bioRxiv2019

The repositioning of epigenetic probes/inhibitors identifies new anti-schistosomal lead compounds and chemotherapeutic targets

BackgroundPraziquantel represents the frontline chemotherapy used to treat schistosomiasis, a neglected tropical disease (NTD) caused by infection with macro-parasitic blood fluke schistosomes. While this drug is safe, its inability to kill all schistosome lifecycle stages within the human host often requires repeat treatments. This limitation, amongst others, has led to the search for novel anti-schistosome replacement or combinatorial chemotherapies. Here, we describe a repositioning strategy to assess the anthelmintic activity of epigenetic probes/inhibitors obtained from the Structural Genomics Consortium.\n\nMethodology/Principle findingsThirty-seven epigenetic probes/inhibitors targeting histone readers, writers and erasers were initially screened against Schistosoma mansoni schistosomula using the high-throughput Roboworm platform. At 10 {micro}M, 14 of these 37 compounds (38%) negatively affected schistosomula motility and phenotype after 72 hours of continuous co-incubation. Subsequent dose-response titrations against schistosomula and adult worms revealed epigenetic probes targeting one reader (NVS-CECR2-1), one writer (LLY-507 and BAY-598) and one eraser (GSK-J4) to be particularly active. As LLY-507/BAY-598 (SMYD2 histone methyltransferase inhibitors) and GSK-J4 (a JMJD3 histone demethylase inhibitor) regulate an epigenetic process (protein methylation) known to be critical for schistosome development, further characterisation of these compounds/putative targets was performed. RNA interference (RNAi) of one putative LLY-507/BAY-598 S. mansoni target (Smp_000700) in adult worms replicated the compound-mediated motility and egg production defects. Furthermore, H3K36me2, a known product catalysed by SMYD2 activity, was also reduced by LLY-507 (25%), BAY-598 (23%) and siSmp_000700 (15%) treatment of adult worms. Oviposition and packaging of vitelline cells into in vitro laid eggs was also significantly affected by GSK-J4 (putative cell permeable prodrug inhibitor of Smp_034000), but not by the related structural analogue GSK-J1 (non-permeable inhibitor).\n\nConclusion/SignificanceCollectively, these results provide further support for the development of next-generation drugs targeting schistosome epigenetic pathway components. In particular, the progression of histone methylation/demethylation modulators presents a tractable strategy for anti-schistosomal control.\n\nAuthor SummaryHuman schistosomiasis is caused by infection with parasitic blood fluke worms. Global control of this NTD is currently facilitated by administration of a single drug, praziquantel (PZQ). This mono-chemotherapeutic strategy of schistosomiasis control presents challenges as PZQ is not active against all human-dwelling schistosome lifecycle stages and the evolution of PZQ resistant parasites remains a theat. Therefore, new drugs to be used in combination with or in replacement of PZQ are urgently needed. Here, continuing our studies on Schistosoma mansoni epigenetic processes, we performed anthelmintic screening of 37 epigenetic probes/epigenetic inhibitors obtained from the Structural Genomics Consortium (SGC). The results of these studies highlighted that schistosome methylation/demethylation processes are acutely vulnerable. In particular, compounds affecting schistosome SMYD (LLY-507, BAY-598) or JMJD (GSK-J4) homologues are especially active on schistosomula and adult worms during in vitro phenotypic drug screens. The active epigenetic probes identified here as well as their corresponding S. mansoni protein targets offers new starting points for the development of next-generation anti-schistosomals.

zoology↗