Apicomplexan parasites are a heterogeneous group of more than 6000 intracellular protists [1], many of which have important clinical and economical relevance as causative agents of human and veterinary diseases. Apicomplexan parasites have evolved a wide variety of strategies to infect and thrive in different hosts often while maintaining complex life cycles that include alternation between different biological niches. Their ability to proliferate and sustain infections in such conditions requires specific adaptations that are controlled by tight regulation of gene expression. In recent years, noncoding RNAs (ncRNAs) have been identified as important epigenetic regulators of eukaryotic gene expression. In parallel, technical advances in whole-genome transcriptomics revealed that Apicomplexan parasites transcribe a large number of noncoding transcripts. Studies using these approaches were performed primarily on human pathogens such as Plasmodium falciparum, the pathogen responsible for the deadliest form of human malaria, Toxoplasma gondii, which poses a threat to immunocompromised individuals and pregnant women and Cryptosporidium parvum, one of the most common agents of cryptosporidiosis that often leads to chronic diarrhea in humans [2]. Studies using strand-specific RNA-seq have revealed the presence of thousands of novel noncoding transcripts in Plasmodium 3, 4, 5, 6•, Cryptosporidium 7, 8• and Toxoplasma [9]. Annotation of RNA-seq data from T. gondii led to the estimation of approximately 2700 putative noncoding transcripts [9]. Nascent strand-specific RNA-seq was recently used to identify 1768 putative noncoding RNAs in Plasmodium. The subcellular localization and stage-specific expression of several of these long noncoding RNA (lncRNAs) were further validated using RNA Fluorescence In Situ Hybridization and single-cell RNA sequencing to resolve their temporal expression profile across the parasite life cycle [6]. Merging the newly annotated putative noncoding RNA genes with the data of a genome-wide mutation screen 10, 11 suggested that 335 lncRNAs might have essential functions in P. falciparum [12]. Recent breakthroughs in the ability to culture C. parvum [13] paved the way to study the molecular and cellular biology of this pathogen using advanced genetics, including next-generation sequencing technologies. Transcriptomic analysis using strand-specific RNA-seq throughout the parasite’s developmental cycle revealed significant stage-specific antisense transcription, identifying approximately 400 noncoding RNAs, of which most are putative lncRNA antisense transcripts [7]. In a later study, Li et al. applied small RNA-seq and found additional 79 novel intermediate-size ncRNAs of unknown function as well as numerous small nucleolar RNAs and tRNA-derived small RNAs [8]. Yet, the function of the vast majority of individual transcripts identified in these large-scale screens thus far remains elusive.

Numerous studies have implicated eukaryotic lncRNAs as regulators of gene expression that could function through several mechanisms. They may interact directly with DNA and form R-loops that maintain the locus open and accessible for transcription [14]. In addition, they may incorporate into chromatin and activate or silence transcription, either by recruiting chromatin modifiers or by serving as their decoys and thus influence chromatin condensation [15]. lncRNAs appear to also be involved in nuclear architecture, and compartmentalization is shown to be associated with chromatin loops. In other examples, they serve as scaffold for specific subnuclear compartments by forming molecular condensates, such as nuclear speckles [16]. In addition, lncRNAs interact with RNA-binding proteins (RBPs) and were implicated in several post-transcriptional regulatory mechanisms, such as splicing, RNA stability and even translation, as some were found to be associated with ribosomes 17, 18.

One of the challenges in studying the biological functions of lncRNAs is the relatively low sequence conservation observed among species and that they may function in different ways by forming RNA–RNA, RNA–DNA or RNA–protein interactions. In addition, different domains along the transcript may create different interactions with RNA, DNA, or protein so that a single lncRNA transcript may have different functions [15]. One of the marked examples is the XIST lncRNA that regulates X chromosome dosage compensation in mammals, which is involved in several mechanisms, including removal of euchromatin markers and replacing them with heterochromatin factors as well as reorganizing the 3D structure of the X chromosome 19, 20.

While it becomes clear that Apicomplexan parasites transcribe a variety of lncRNAs 2, 17, they are still understudied, and their biological functions and mechanisms of action remain mostly unknown. Here, we update the current knowledge on the role of noncoding RNAs in epigenetic gene regulation in Apicomplexan parasites, highlight recent advances, and focus on possible regulatory mechanisms that may lead to a better understanding of the parasites’ biology.