Radical cure of Plasmodium vivax malaria must include elimination of quiescent "hypnozoite" forms in the liver; however, the only FDA-approved treatments are contraindicated in many vulnerable populations. To identify new drugs and drug targets, we screened the Repurposing, Focused Rescue, and Accelerated Medchem library against P. vivax liver stages and identified the DNA methyltransferase inhibitors hydralazine and cadralazine as active against hypnozoites. We then used bisulfite sequencing and immunostaining to identify cytosine modifications in the infectious stage (sporozoites) and liver stages, respectively. A subsequent screen of epigenetic inhibitors revealed hypnozoites are broadly sensitive to histone acetyltransferase and methyltransferase inhibitors, indicating that several epigenetic mechanisms are likely modulating hypnozoite persistence. Our data present an avenue for the discovery and development of improved radical cure antimalarials.

DNA cytosine methylation is an epigenetic modification that has a critical role in gene regulation and genome stability. DNA methylation can be detected and measured using sequencing instruments after sodium bisulfite conversion, but experiments can be expensive for large eukaryotic genomes. Sequencing non-uniformity and mapping biases can leave parts of the genome with low or no coverage, thus hampering the ability of obtaining DNA methylation levels for all cytosines. To address these limitations, several computational methods have been proposed that can predict DNA methylation from the DNA sequence around the cytosine, or from the methylation level of nearby cytosines. Most of these methods are, however, entirely focused on CG methylation in humans and other mammals. In this work, we study for the first time the problem of predicting cytosine methylation for CG, CHG, and CHH contexts on five plant species, either from the DNA primary sequence around the cytosine or the methylation levels of neighboring cytosines. In this framework, we also study (1) the cross-species prediction problem, i.e., the classification performance when training on one species and testing on another species, and the (2) the cross-context prediction problem, i.e., the classification performance when training on one context and testing on another context (within the same species). Finally, we show that providing the classifier with gene annotation information allows our classifier to outperform the prediction accuracy of state-of-the-art methods.

DNA methylation can be detected and measured using sequencing instruments after sodium bisulfite conversion, but experiments can be expensive for large eukaryotic genomes. Sequencing nonuniformity and mapping biases can leave parts of the genome with low or no coverage, thus hampering the ability of obtaining DNA methylation levels for all cytosines. To address these limitations, several computational methods have been proposed that can predict DNA methylation from the DNA sequence around the cytosine or from the methylation level of nearby cytosines. However, most of these methods are entirely focused on CG methylation in humans and other mammals. In this work, we study, for the first time, the problem of predicting cytosine methylation for CG, CHG and CHH contexts on six plant species, either from the DNA primary sequence around the cytosine or from the methylation levels of neighboring cytosines. In this framework, we also study the cross-species prediction problem and the cross-context prediction problem (within the same species). Finally, we show that providing gene and repeat annotations allows existing classifiers to significantly improve their prediction accuracy. We introduce a new classifier called AMPS (annotation-based methylation prediction from sequence) that takes advantage of genomic annotations to achieve higher accuracy.