Biology subjects

Martorelli Di Genova, B.

Publications and source records attributed to Martorelli Di Genova, B..

5 recordsLinked to original sources

bioRxiv2026

Beyond presence and absence: a survey of TOR and AMPK across 210 eukaryotes reveals unusually long coccidian AMPKγ

Energy and nutrient regulation in eukaryotes depends on AMP-activated protein kinase (AMPK) and target of rapamycin (TOR), yet their evolution is usually described by which components are present or absent. We surveyed 10 TOR- and AMPK-related targets across 210 eukaryotic species and paired that inventory with sequence- and structure-level measurements of the proteins that remain. Across 2100 species-by-target cells, 1255 were coded present, 683 absent and 162 not assessable, with repertoires differing among lineages. Phylogenetically corrected analyses recovered associations among a subset of component absences, especially within canonical complexes, although correlated non-detection in incomplete proteomes remains an alternative explanation. Sequence features varied substantially. In TOR, N-terminal HEAT-region divergence was associated with absence of several TOR-complex partners in discrete-state analyses. In AMPK, the activation-loop threonine was retained in every species with a readable site, while other features varied strongly: {beta} carbohydrate-binding-module conservation covaried with C-terminal length under the primary phylogenetic model, and AMPK{gamma} showed pronounced lineage-associated length variation. In the focal coccidian-group cohort, {gamma} sequences had a median length of 909 residues versus 396 in other sampled copies; the median was 924 among the nine sequences with a tree-based orthology determination, while two Eimeria sequences remain provisional candidates. Low primary nucleotide-contact-site identity was shared more broadly across sampled Apicomplexa. Together, these analyses reveal lineage-structured variation in TOR and AMPK at three levels: component repertoires, sequence features of retained proteins, and associations among components. Signaling architecture cannot be inferred from presence and absence alone.

evolutionary biology↗

bioRxiv2026

Identification of UCB-9721 as a potent inhibitor of MyoA, the essential class XIV myosin motor of apicomplexan parasites

The virulence of Toxoplasma gondii and other apicomplexan parasites relies on a unique form of cellular motility driven by MyoA, an unconventional class XIV myosin motor protein. To identify new chemical probes for investigating the molecular mechanisms of parasite motility, we screened over 50,000 small molecules for inhibitors of T. gondii MyoA (TgMyoA). The top hit from the screen, UCB-9721, is almost 40-fold more potent as an inhibitor of TgMyoA actin-activated ATPase activity than the previously described TgMyoA inhibitor, KNX-002, and 45-fold more potent at inhibiting parasite motility, with no detectable toxicity towards mammalian cells. UCB-9721 also inhibited the motility and/or growth of the related apicomplexan parasites Plasmodium falciparum, Cryptosporidium parvum, and Babesia duncani, suggesting that this compound will be a useful new chemical probe for studying motility and MyoA function in apicomplexan parasites more broadly. While UCB-9721 and KNX-002 were identified independently, they share a similar chemical scaffold. To determine why UCB-9721 is so much more potent than KNX-002 and to inform future development of this inhibitor class, we undertook comparative molecular docking analyses, targeted TgMyoA mutagenesis, and a directed structure-activity relationship analysis. The results identified the sulfonamide group of UCB-9721 and its hydrogen bond interactions with R249, E275 and a stabilized water network within the TgMyoA binding pocket as key to the compounds increased potency. Further development of UCB-9721, informed by the results presented here, may transform this promising new chemical class into actionable drug development leads against this important group of human and animal pathogens.

microbiology↗

bioRxiv2024

Impact of Equilibrative Nucleoside Transporters TgAT1, TgENT1, and TgENT3 on Toxoplasma gondii Infection and Differentiation

Toxoplasma gondii cannot synthesize purines de novo and must import them; yet, the functional interplay among its four equilibrative nucleoside transporters (ENTs) homologs remains unclear. We systematically deconstructed this network by combining CRISPR-Cas9 knockouts with an auxin-inducible degron. Across all phenotypic assays, tachyzoite replication, nucleoside-analogue sensitivity, alkaline-stress-induced differentiation, and murine cyst formation, the {Delta}TgENT2 strain was indistinguishable from the parental line, indicating that TgENT2 is dispensable under the conditions tested. In contrast, the double mutant {Delta}TgAT1{Delta}TgENT3 exhibited delayed bradyzoite differentiation in vitro and produced smaller brain cysts in vivo. This double deletion triggered a [~]3-fold transcriptional up-regulation of TgENT1, whose product we partially localized to the plant-like vacuolar compartment (PLVAC). Conditional depletion of TgENT1 caused complete intracellular growth arrest, PLVAC swelling, and a purine-starvation-like transcriptomic program enriched for nucleoside phosphatases and cyclic-nucleotide phosphodiesterases. These findings reveal a compensatory salvage pathway in which the parasite reroutes purine acquisition through a vacuolar route when plasma-membrane import is compromised. Although this adaptation sustains tachyzoite proliferation, it fails during the energetically demanding transition to bradyzoites, creating a metabolic bottleneck that impairs chronic infection. Our work reveals an adaptable yet ultimately limited purine-import network and identifies TgENT1, along with the vacuolar salvage axis it mediates, as a promising therapeutic target for blocking lifelong toxoplasmosis.

microbiology↗

bioRxiv2024

Variant mutation in SARS-CoV-2 nucleocapsid enhances viral infection via altered genomic encapsidation

The evolution of SARS-CoV-2 variants and their respective phenotypes represents an important set of tools to understand basic coronavirus biology as well as the public health implications of individual mutations in variants of concern. While mutations outside of Spike are not well studied, the entire viral genome is undergoing evolutionary selection, particularly the central disordered linker region of the nucleocapsid (N) protein. Here, we identify a mutation (G215C), characteristic of the Delta variant, that introduces a novel cysteine into this linker domain, which results in the formation of a disulfide bond and a stable N-N dimer. Using reverse genetics, we determined that this cysteine residue is necessary and sufficient for stable dimer formation in a WA1 SARS-CoV-2 background, where it results in significantly increased viral growth both in vitro and in vivo. Finally, we demonstrate that the N:G215C virus packages more nucleocapsid per virion and that individual virions are larger, with elongated morphologies.

microbiology↗

bioRxiv2019

Intestinal delta-6-desaturase activity determines host range for Toxoplasma sexual reproduction

Many eukaryotic microbes have complex lifecycles that include both sexual and asexual phases with strict species-specificity. While the asexual cycle of the protistan parasite Toxoplasma gondii can occur in any warm-blooded mammal, the sexual cycle is restricted to the feline intestine1. The molecular determinants that identify cats as the definitive host for T. gondii are unknown. Here, we defined the mechanism of species specificity for T. gondii sexual development and break the species barrier to allow the sexual cycle to occur in mice. We determined that T. gondii sexual development occurs when cultured feline intestinal epithelial cells are supplemented with linoleic acid. Felines are the only mammals that lack delta-6-desaturase activity in their intestines, which is required for linoleic acid metabolism, resulting in systemic excess of linoleic acid2, 3. We found that inhibition of murine delta-6-desaturase and supplementation of their diet with linoleic acid allowed T. gondii sexual development in mice. This mechanism of species specificity is the first defined for a parasite sexual cycle. This work highlights how host diet and metabolism shape coevolution with microbes. The key to unlocking the species boundaries for other eukaryotic microbes may also rely on the lipid composition of their environments as we see increasing evidence for the importance of host lipid metabolism during parasitic lifecycles4, 5. Pregnant women are advised against handling cat litter as maternal infection with T. gondii can be transmitted to the fetus with potentially lethal outcomes. Knowing the molecular components that create a conducive environment for T. gondii sexual reproduction will allow for development of therapeutics that prevent shedding of T. gondii parasites. Finally, given the current reliance on companion animals to study T. gondii sexual development, this work will allow the T. gondii field to use of alternative models in future studies.

microbiology↗