Biology subjects

Hartl, D. L.

Publications and source records attributed to Hartl, D. L..

7 recordsLinked to original sources

bioRxiv2025

Cellular and molecular basis of PfCoronin function in artemisinin resistance in Plasmodium falciparum

Artemisinin-based therapies are central to malaria treatment, but their efficacy is threatened by the emergence of resistant Plasmodium falciparum strains. While the role of Pfkelch13 mutations in clinical resistance is well established, recent reports from Africa implicate Pfcoronin mutations in treatment failure, adding to the complexity of resistance mechanisms. Here, we show that PfCoronin, a non-essential actin regulator active during the ring-stage, facilitates efficient hemoglobin uptake. We demonstrate that resistance-associated Pfcoronin mutations disrupt PfCoronins interaction with PfActin and its ring-stage localization, leading to impaired endocytosis and reduced hemoglobin acquisition. PfCoronin and PfKelch13 function in distinct cellular regions; mutations in both converge to limit heme availability and artemisinin activation. Although PfCoronin is dispensable for parasite viability, our findings demonstrate that Pfcoronin mutations reduce endocytosis and modulate artemisinin susceptibility during the clinically relevant ring stage--highlighting how non-essential, temporally restricted proteins can shape antimalarial drug response and resistance.

cell biology↗

bioRxiv2024

MalKinID: A Likelihood-Based Model for Identifying Malaria Parasite Genealogical Relationships Using Identity-by-Descent

Pathogen genomics is a powerful tool for tracking infectious disease transmission. In malaria, identity-by-descent (IBD) is used to assess the genetic relatedness between parasites and has been used to study transmission and importation. In theory, IBD can be used to distinguish genealogical relationships to reconstruct transmission history or identify parasites for genotype- to-phenotype quantitative-trait-locus experiments. MalKinID (Malaria Kinship Identifier) is a new likelihood-based classification model designed to identify genealogical relationships among malaria parasites based on genome-wide IBD proportions and IBD segment distributions. MalKinID was calibrated to the genomic data from three laboratory-based genetic crosses (yielding 440 parent-child and 9060 full-sibling comparisons). MalKinID identified lab generated F1 progeny with >80% sensitivity and showed that 0.39 (95% CI 0.28, 0.49) of the second- generation progeny of a NF54 and NHP4026 cross were F1s and 0.56 (0.45, 0.67) were backcrosses of an F1 with the parental NF54 strain. In simulated outcrossed importations, MalKinID accurately reconstructs genealogy history with high precision and sensitivity, with F1- scores exceeding 0.84. However, when importation involves inbreeding, such as during serial co-transmission, the precision and sensitivity of MalKinID declined, with F1-scores of 0.76 (0.56, 0.92) and 0.23 (0.0, 0.4) for PC and FS and <0.05 for second-degree and third-degree relatives. Genealogical inference is most powered 1) when outcrossing is the norm or 2) when multi- sample comparisons based on a predefined pedigree are used. MalKinID lays the foundations for using IBD to track parasite transmission history and for separating progeny for quantitative- trait-locus experiments.

genetics↗

bioRxiv2023

Artemisinin resistance mutations in Pfcoronin impede hemoglobin uptake

Artemisinin (ART) combination therapies have been critical in reducing malaria morbidity and mortality, but these important drugs are threatened by growing resistance associated with mutations in Pfcoronin and Pfkelch13. Here, we describe the mechanism of Pfcoronin-mediated ART resistance. PfCoronin interacts with PfActin and localizes to the parasite plasma membrane (PPM), the digestive vacuole (DV) membrane, and membrane of a newly identified preDV compartment--all structures involved in the trafficking of hemoglobin from the RBC for degradation in the DV. Pfcoronin mutations alter PfActin homeostasis and impair the development and morphology of the preDV. Ultimately, these changes are associated with decreased uptake of red blood cell cytosolic contents by ring-stage Plasmodium falciparum. Previous work has identified decreased hemoglobin uptake as the mechanism of Pfkelch13-mediated ART resistance. This work demonstrates that PfCoronin appears to act via a parallel pathway. For both Pfkelch13-mediated and Pfcoronin-mediated ART resistance, we hypothesize that the decreased hemoglobin uptake in ring stage parasites results in less heme-based activation of the artemisinin endoperoxide ring and reduced cytocidal activity. This study deepens our understanding of ART resistance, as well as hemoglobin uptake and development of the DV in early-stage parasites.

cell biology↗

bioRxiv2021

Switching an active site helix in dihydrofolate reductase reveals limits to sub-domain modularity

To what degree are individual structural elements within proteins modular such that similar structures from unrelated proteins can be interchanged? We study sub-domain modularity by creating 20 chimeras of an enzyme, E. coli dihydrofolate reductase (DHFR), in which a catalytically important, 10-residue -helical sequence is replaced by -helical sequences from a diverse set of proteins. The chimeras stably fold but have a range of diminished thermal stabilities and catalytic activities. Evolutionary coupling analysis indicates that the residues of this -helix are under selection pressure to maintain catalytic activity in DHFR. We performed molecular dynamics simulations using replica exchange with solute-tempering. Chimeras with low catalytic activity exhibit non-helical conformations that block the binding site and disrupt the positioning of the catalytically essential residue D27. Simulation observables and in vitro measurements of thermal stability and substrate binding affinity are strongly correlated. Several E. coli strains with chromosomally integrated chimeric DHFRs can grow, with growth rates that follow predictions from a kinetic flux model that depends on the intracellular abundance and catalytic activity of DHFR. Our findings show that although -helices are not universally substitutable, the molecular and fitness effects of modular segments can be predicted by the biophysical compatibility of the replacement segment. Statement of Significance-helices are ubiquitous components of protein structure that exhibit a degree of independent folding behavior, making them plausible structural modules within proteins. Here, we assess the effects of switching the sequence of an -helix in an essential enzyme for -helical sequences from evolutionarily unrelated proteins. The resultant chimeric proteins can still fold but enzymatic activity, stability, and cellular growth rates are negatively affected. Computational investigations reveal how residues in an -helix have been shaped by selection pressure to maintain catalytic activity and a specific, helical conformation of the protein. More broadly, we illustrate how molecular and fitness effects of switching protein segments depend on the protein and cellular context.

biophysics↗

bioRxiv2019

Experimental evolution for niche breadth in bacteriophage T4 highlights the importance of structural genes

Ecologists have long studied the evolution of niche breadth, including how variability in environments can drive the evolution of specialism and generalism. This concept is of particular interest in viruses, where niche-breadth evolution may explain viral disease emergence, or underlie the potential for therapeutic measures like phage therapy. Despite the significance and potential applications of virus-host interactions, the genetic determinants of niche-breadth evolution remain unexplored in many bacteriophage. In this study, we present the results of an evolution experiment with a model bacteriophage system, Escherichia virus T4, in several host environments: exposure to E. coli C, exposure to E. coli K-12, and exposure to both E. coli C and E. coli K-12. This experimental framework allowed us to investigate the phenotypic and molecular manifestations of niche-breadth evolution. First, we show that selection on different hosts led to measurable changes in phage productivity in all experimental populations. Second, whole--genome sequencing of experimental populations revealed signatures of selection. Finally, clear and consistent patterns emerged across the host environments, especially the presence of new mutations in phage structural genes. A comparison of mutations found across functional gene categories revealed that structural genes acquired significantly more mutations than other categories. Our findings suggest that structural genes--those that provide morphological and biophysical integrity to a virus--are central determinants in bacteriophage niche breadth.

evolutionary biology↗

bioRxiv2018

Proteostasis environment shapes higher-order epistasis operating on antibiotic resistance

Recent studies have shown that higher-order epistasis is ubiquitous and can have large effects on complex traits. Yet, we lack frameworks for understanding how epistatic interactions are influenced by basic aspects of cell physiology. In this study, we assess how protein quality control machinery--a critical component of cell physiology--affects epistasis for different traits related to bacterial resistance to antibiotics. Specifically, we attempt to disentangle the interactions between different protein quality control genetic backgrounds and two sets of mutations: (i) SNPs associated with resistance to antibiotics in an essential bacterial enzyme (dihydrofolate reductase, or DHFR) and (ii) differing DHFR bacterial species-specific amino acid background sequences (Escherichia coli, Listeria grayi, and Chlamydia muridarum). In doing so, we add nuance to the generic observation that non-linear genetic interactions are widespread and capricious in nature, by proposing a mechanistically-grounded analysis of how proteostasis shapes epistasis. These findings simultaneously fortify and demystify the role of environmental context in modulating higher-order epistasis, with direct implications for evolutionary theory, genetic modification technology, and efforts to manage antimicrobial resistance.

genetics↗

bioRxiv2017

De Novo Mutations Resolve Disease Transmission Pathways in Clonal Malaria

Detecting de novo mutations in viral and bacterial pathogens enables researchers to reconstruct detailed networks of disease transmission and is a key technique in genomic epidemiology. However these techniques have not yet been applied to the malaria parasite, Plasmodium falciparum, in which a larger genome, slower generation times, and a complex life cycle make them difficult to implement. Here we demonstrate the viability of de novo mutation studies in P. falciparum for the first time. Using a set of clinical samples and novel methods of sequencing, library preparation, and genotyping, we have genotyped low-complexity regions of the genome with a high degree of accuracy. Despite its slower evolutionary rate compared to bacterial or viral species, de novo mutation can be detected in P. falciparum across timescales of just 1-2 years and evolutionary rates in low-complexity regions of the genome can be up to twice that detected in the rest of the genome. The increased mutation rate allows the identification of separate clade expansions that cannot be found using previous genomic epidemiology approaches and could be a crucial tool for mapping residual transmission patterns in disease elimination campaigns and reintroduction scenarios.

genomics↗