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Epithemiaspp. diatoms contain obligate, nitrogen-fixing endosymbionts, or “diazoplasts”, derived from cyanobacteria. These algae are a rare example of photosynthetic eukaryotes that have successfully coupled oxygenic photosynthesis with oxygen-sensitive nitrogenase activity. Here, we report a newly-isolated species,E. clementina, as a model to investigate endosymbiotic acquisition of nitrogen fixation. To detect the metabolic changes associated with endosymbiotic specialization, we compared nitrogen fixation, associated carbon and nitrogen metabolism, and their regulatory pathways in theEpithemiadiazoplast with its close, free-living cyanobacterial relative,Crocosphaera subtropica. UnlikeC. subtropica, we show that nitrogenase activity in the diazoplast is concurrent with, and even dependent on, host photosynthesis and no longer associated with cyanobacterial glycogen storage suggesting carbohydrates are imported from the host diatom. Carbohydrate catabolism in the diazoplast indicates that the oxidative pentose pathway and oxidative phosphorylation, in concert, generates reducing equivalents and ATP and consumes oxygen to support nitrogenase activity. In contrast to expanded nitrogenase activity, the diazoplast has diminished ability to utilize alternative nitrogen sources. Upon ammonium repletion, negative feedback regulation of nitrogen fixation was conserved, however ammonia assimilation showed paradoxical responses in the diazoplast compared withC. subtropica. The altered nitrogen regulation likely favors nitrogen transfer to the host. Our results suggest that the diazoplast is specialized for endosymbiotic nitrogen fixation. Altogether, we establish a new model for studying endosymbiosis, perform the first functional characterization of this diazotroph endosymbiosis, and identify metabolic adaptations for endosymbiotic acquisition of a critical biological function.

This study focuses on the social presence framework (Rourke et al., 2001), in order to examine the ways that university-level international students develop social interaction and support in a virtual asynchronous learning community in an online class during the COVID-19 pandemic. English language learners (ELLs) participated in weekly online exchanges on a video discussion platform called Flipgrid in the form of oral dialogue journals for reflection on their academic learning and experiences during these disruptive times. These ELLs’ video journals and peer responses (N = 198) were collected for content analysis, in order to investigate how the use of video-based asynchronous computer-mediated communication (ACMC) can establish positive social and emotional support and a sense of community. The findings of the study indicate that ACMC was successful in establishing interconnectedness in terms of high levels of selfdisclosure, positive facial expressions, and other indicators of social and emotional support, demonstrating social presence. Implications of the findings are discussed in terms of how social presence is expressed and fostered in videobased ACMC communities during emergency remote teaching.

Changes in the copy number of large genomic regions, termed copy number variations or CNVs, are an important adaptive strategy for malaria parasites. Numerous CNVs across thePlasmodium falciparumgenome contribute directly to drug resistance or impact fitness of this protozoan parasite. CNVs that encompass the dihydroorotate dehydrogenase (DHODH) gene confer resistance to antimalarials that target this enzyme in the pyrimidine biosynthesis pathway (i.e. DSM1). During the characterization of DSM1 resistant parasite lines with DHODH CNVs, we detected selection of an additional CNV that encompasses 3 genes (~5 kb) including GTP cyclohydrolase I (GCH1 amplicon). While this locus has been implicated in increased fitness of antifolate resistant parasites, GCH1 CNVs had not previously been reported to contribute to resistance to other antimalarials. Here, we further explored the association between GCH1 and DHODH copy number. We visualized single long reads and directly quantified the number of tandem GCH1 amplicons in a parental line versus a DSM1-selected line. We found that the GCH1 amplicons share a consistent structure in all lines. However, we detected more reads that encompassed a higher number of amplicons in the resistant (up to 7 amplicons) compared to the parental line (3 amplicons). To better understand the implications of this result, we evaluated variation at this locus across multiple short- and long-read data sets collected from various parasite lines. Based on our analysis of parasites resistant to other DHODH inhibitors (DSM265, DSM267, and DSM705), GCH1 is not likely contributing directly to resistance; however, higher numbers of the GCH1 amplicon are associated with increased DHODH copies and may compensate for changes in metabolism of parasites. This is supported by the direct connection between folate and pyrimidine metabolism, which together contribute to nucleic acid biosynthesis. This study highlights the importance of studying clonal variation and potential biochemical connections as novel antimalarials move closer to clinical approval.

To stem the text-borrowing practices of language learners—considered plagiarism in the U.S. academic community—educators can use the strategies presented in this article to teach writing processes ...

This study applied pedagogically-focused project design by using Instagram as a platform to investigate how the use of social media such as Instagram in a multimodal digital storytelling model could bridge the skills English language learners (ELLs) learn in the classroom to out-of-school literacy practices. The study applied the five learning objectives of the bridging-activities framework to investigate to what extent ELLs achieve these objectives. There were forty-two participants (female: n= 22; male: n= 17), international arts and media students. The study collected their main Instagram posts, questionnaires were given with a five-point Likert scale and open-ended questions, and individual follow-up interviews for content analysis. A cross-tabulation was conducted to investigate the relationships between ELLs' Instagram use and how they perceive Instagram as a meaningful way of communication for professional purposes. The findings revealed pedagogical practices of using Instagram as a tool for professional purposes and how ELLs achieved the bridging-activities learning outcomes.

Online participatory cultures (e.g. social media) present a number of opportunities and challenges for language learners. Learners can collaborate with like-minded peers in online affinity spaces o...

With the rapid growth of Asian immigrants in American educational settings, this mixed method study examined whether a professional development program on cultural awareness and pedagogy for faculty members would impact their knowledge and skills in working with Asian immigrants to ensure these students’ retention and success. This study, therefore, assessed the effectiveness and the impact of the program designed to address the needs of Asian immigrants. Guided and supported by the constructivism learning theory (Vygotsky, 1980), the professional development program applied theories of intercultural communication training (Bennett & Salonen, 2004). A mixed method of inquiry involved surveys with ten faculty members and 16 students, in-depth interviews, classroom observations, and document analysis. Faculty members attended the workshops during the first year of students’ enrollment in their classes. The data stem from a longitudinal study conducted throughout the three years of students’ enrollment in the institution. The results established a probable relationship between the workshops and faculty members’ success in working with Asian immigrants.

This study applied pedagogically-focused project design by using Instagram as a platform to investigate how the use of social media such as Instagram in a multimodal digital storytelling model could bridge the skills English language learners (ELLs) learn in the classroom to out-of-school literacy practices. The study applied the five learning objectives of the bridging-activities framework to investigate to what extent ELLs achieve these objectives. There were forty-two participants (female: n= 22; male: n= 17), international arts and media students. The study collected their main Instagram posts, questionnaires were given with a five-point Likert scale and open-ended questions, and individual follow-up interviews for content analysis. A cross-tabulation was conducted to investigate the relationships between ELLs' Instagram use and how they perceive Instagram as a meaningful way of communication for professional purposes. The findings revealed pedagogical practices of using Instagram as a tool for professional purposes and how ELLs achieved the bridging-activities learning outcomes.

This study examines the extent to which parental involvement factors predict the likelihood of Hispanic non-native English speakers’ (NNESs) postsecondary education (PES) enrollment after controlli...

Many learners, even those studying at universities in regions where the target language is spoken, lack opportunities for meaningful language use outside of the classroom. One avenue for learners to increase authentic target-language communication is online affinity spaces within social media platforms, where interactions with other users are formed around shared interests rather than personal connections. International students at an arts and media college in the Midwestern United States were asked to read a discussion thread within a social media platform, summarize what they found useful, and respond to pre- and posttask questionnaires. The platform used, Reddit, features anonymous user-generated content in a wide range of discussion forums based around specific interests and geographic locations. This study used qualitative data to investigate the extent to which international students participate in online communities like these and the factors or barriers that keep them from achieving full participation. The findings are then used to present learner training strategies that can help reduce or remove those barriers, enabling language learners to increase their participation in target-language online communities. Plusieurs apprenantes et apprenants, même parmi celles et ceux qui étudient dans une université située dans une région où la langue cible est parlée, n’ont pas suffisamment d’occasions de pratiquer avantageusement leur nouvelle langue en dehors de la salle de classe. Une avenue qui s’ouvre à elles et à eux pour augmenter leurs chances de s’adonner à des communications authentiques dans leur langue cible est l’espace d’affinité en ligne sur les réseaux sociaux, endroit où les interactions sont davantage basées sur le partage d’intérêts communs que sur des relations personnelles. Des étudiantes et étudiants internationaux d’un collège des arts et des médias du Midwest des États-Unis ont été invités à lire un fil de discussion sur une plateforme de réseau social, à en résumer les éléments jugés utiles et à répondre à un questionnaire avant et après l’exercice. La plateforme utilisée, Reddit, présente des contenus qui sont générés anonymement par les utilisateurs dans un large éventail de forums de discussion et qui sont regroupés autour d’intérêts et de secteurs géographiques particuliers. Cette étude utilise des données qualitatives permettant d’évaluer dans quelle mesure les étudiantes et étudiants internationaux participent à la vie de communautés en ligne de ce genre et de déterminer quels sont les facteurs ou obstacles qui les empêchent de le faire pleinement. Les constatations sont ensuite utilisées pour présenter des stratégies de formation des apprenantes et apprenants qui sont susceptibles d’aider à réduire ou aplanir ces obstacles et à aider par le fait même les participants à s’impliquer davantage dans la vie de communautés en ligne s’exprimant dans leur langue cible.

This qualitative study demonstrates how a culturally responsive teaching (CRT) approach builds the capacity of faculty and staff to create a social and emotional learning (SEL) environment that is conducive to success for international students. The researchers propose a CRT-SEL model to address the increasing needs of mid-sized universities with emerging international students. The findings from 14 faculty and staff participants point to professional development strategies that can help reduce or remove barriers for working with international students. This study highlights the need to build upon the CRT and SEL tenets and align them with the existing support networks within the mid-size higher education institutions for classroom learnings and off-campus activities to bolster international students’ overall success. The results also indicate the faculty and staff’s varying priorities and diverging trajectories.

While the number of the Asian non-native English speakers (NNESs) is increasing, their postsecondary education (PSE) enrollment rate has remained low in comparison to enrollment rates of Asian native English speakers (NESs). The achievement gap in postsecondary enrollment between NNESs and NESs has widened, due not only to content areas such as reading and language arts that require higher English proficiency but also sociocultural factors, such as parental involvement. The current study aims to investigate the extent to which parental involvement factors predict the likelihood of Asian NNESs PSE enrollment after controlling for socioeconomic and linguistic factors. This study, being an expansion of previous work, which explored parental involvement and NNESs' PSE in the U.S., uses the national representative data from the Education Longitudinal Study dataset in 2002 (ELS: 2002) and a binary multilevel logistic regression model analysis. The results indicate that parental involvement is related to a greater likelihood of attending PSE institutions. Among the various forms of parental involvement, parent-student involvement and parent-school involvement have the greatest impact on Asian NNESs' PSE enrollment. The results also showed that Asian NNESs' enrollment is higher if their parents participate in school volunteer work. Limitations, future studies, and implications for educators, parents and school policy makers will also be discussed.

Atg8 family proteins are highly-conserved eukaryotic proteins with diverse autophagy and non-autophagic functions in eukaryotes. While the structural features required for conserved autophagy functions of Atg8 are well-established, little is known about the molecular changes that facilitated acquisition of divergent, non-autophagic functions of Atg8. The malaria parasite Plasmodium falciparum offers a unique opportunity to study non-autophagic functions of Atg8 family proteins because it encodes a single Atg8 homolog whose only essential function is in the inheritance of an unusual secondary plastid called the apicoplast. Here we used functional complementation to investigate the structure-function relationship for this divergent Atg8 protein. We showed that the LC3-interacting region (LIR) docking site (LDS), the major interaction interface of Atg8 protein family, is not sufficient for PfAtg8 apicoplast function. Other regions previously implicated in canonical Atg8 interactions, the ubiquitin-interacting motif (UIM) docking site (UDS) and the N-terminal helix are not required for PfAtg8 function. Finally, the unique Apicomplexan-specific loop previously implicated in interaction with membrane conjugation machinery in vitro, is not required in vivo neither for membrane conjugation nor for the effector function of PfAtg8. These results suggest that the effector function of PfAtg8 is mediated by structural features distinct from those previously identified for macroautophagy and selective autophagy functions.ImportanceThe most extensively studied role of Atg8 proteins is in autophagy. However, it is clear that they have other non-autophagic functions critical to cell function and disease pathogenesis yet understudied compared to their canonical role in autophagy. Mammalian cells contain multiple Atg8 paralogs that have diverse, specialized functions. Gaining molecular insight into their non-autophagic functions is difficult because of redundancy between the homologs and their role in both autophagy and non-autophagic pathways. Malaria parasites such as Plasmodium falciparum are a unique system to study a novel, non-autophagic function of Atg8 separate from its role in autophagy: They have only one Atg8 protein whose only essential function is in the inheritance of the apicoplast, a unique secondary plastid organelle. Insights into the molecular basis of PfAtg8’s function in apicoplast biogenesis will have important implications for the evolution of diverse non-autophagic functions of the Atg8 protein family.

A series of novel thiazole-containing amides were synthesized. A structure-activity relationship study of these compounds led to the identification of potent and selective PfFPPS/GGPPS inhibitors with good in vitro ADME profiles. The most promising candidate molecules were progressed to mouse in vivo PK studies and demonstrated adequate free drug exposure to warrant further investigation.

Plasmodium parasites, which cause malaria, and related apicomplexans are important human and veterinary pathogens. These parasites represent a highly divergent and understudied branch of eukaryotes, and as such often defy the expectations set by model organisms. One striking example of unique apicomplexan biology is the apicoplast, an essential but nonphotosynthetic plastid derived from an unusual secondary (eukaryote-eukaryote) endosymbiosis. Endosymbioses are a major driver of cellular innovation, and apicoplast biogenesis pathways represent a hot spot for molecular evolution. We previously conducted an unbiased screen for apicoplast biogenesis genes in P. falciparum to uncover these essential and innovative pathways. Here, we validate a novel gene candidate from our screen and show that its role in apicoplast biogenesis does not match its functional annotation predicted by model eukaryotes. Our findings suggest that an uncharacterized chloroplast maintenance pathway has been reused for complex plastid biogenesis in this divergent branch of pathogens., Plasmodium parasites and related apicomplexans contain an essential “complex plastid” organelle of secondary endosymbiotic origin, the apicoplast. Biogenesis of this complex plastid poses a unique challenge requiring evolution of new cellular machinery. We previously conducted a mutagenesis screen for essential apicoplast biogenesis genes to discover organellar pathways with evolutionary and biomedical significance. Here we validate and characterize a gene candidate from our screen, Pf3D7_0913500. Using a conditional knockdown strain, we show that Pf3D7_0913500 depletion causes growth inhibition that is rescued by the sole essential product of the apicoplast, isopentenyl pyrophosphate (IPP), and results in apicoplast loss. Because Pf3D7_0913500 had no previous functional annotation, we name it apicoplast-minus IPP-rescued 4 (AMR4). AMR4 has an annotated CaaX protease and bacteriocin processing (CPBP) domain, which in eukaryotes typically indicates a role in CaaX postprenylation processing. Indeed, AMR4 is the only putative CaaX-like protease in Plasmodium parasites which are known to require protein prenylation, and we confirm that the conserved catalytic residue of AMR4 (E352) is required for its apicoplast function. However, we unexpectedly find that AMR4 does not act in a CaaX postprenylation processing pathway in Plasmodium falciparum. Instead, we find that AMR4 is imported into the apicoplast and is derived from a cyanobacterial CPBP gene which was retained through both primary and secondary endosymbiosis. Our findings suggest that AMR4 is not a true CaaX protease, but instead it performs a conserved, uncharacterized chloroplast function that has been retained for complex plastid biogenesis.

This study employed qualitative evidence synthesis (Saldaña, 2012) to critically examine and systematically analyze 63 studies published between 2000-2018 reporting positive educational impacts on English language learners (ELLs). Drawing on Scarcella’s (2003) academic English literacy framework and culturally responsive teaching (Gay, 2001), this study demonstrates three mechanisms to improve ELL outcomes. First, teachers should use both culturally responsive practices and knowledge of language acquisition. Second, fostering family and peer supports creates positive learning environments. Finally, long-term solutions require policies addressing the socio-politico-economic disparities affecting ELLs. These results show, in a synthesized fashion, an approach to equitable quality education for ELLs. Article visualizations

This study presents an innovative approach to developing interlanguage pragmatics (ILP) by training students to engage in online participatory cultures and analyzing their participation through the lens of communities of practice. Participants were university-level English language learners studying in the United States who were trained on the basic layout and practices of a social news and discussion site (Reddit), then assigned weekly postings within forums based on their individual interests. Social media engagement metrics (upvotes and responses) were used to measure and observe the quantity and quality of online interaction, and post-task questionnaires and follow-up interviews investigated learner perceptions about the language and cyberpragmatics in the online communities they encountered. Quantitative results show that participants struggled to achieve high levels of interaction with other users, but qualitative results indicate a wide range of potential benefits for ILP exploration and development. The findings and implications of this study contribute towards best practices in developing strategies for ILP in online spaces and cyberpragmatic awareness among language learners, enabling them to reach higher levels of participation in online communities.

Plasmodium parasites contain several unique membrane compartments in which prenylated proteins may play important roles in pathogenesis. Protein prenylation has also been proposed as an antimalarial drug target because farnesyltransferase inhibitors cause potent growth inhibition of blood-stage Plasmodium. However, the specific prenylated proteins that mediate antimalarial activity have yet to be identified. Given the potential for new parasite biology and elucidating drug mechanism-of-action, we performed a large-scale identification of the prenylated proteome in blood-stage P. falciparum parasites using an alkyne-labeled prenyl analog to specifically enrich parasite prenylated proteins. Twenty high-confidence candidates were identified, including several examples of pathogen-specific prenylation activity. One unique parasite prenylated protein was FYVE-containing coiled-coil protein (FCP), which is only conserved in Plasmodium and related Apicomplexan parasites and localizes to the parasite food vacuole. Targeting of FCP to this parasite-specific compartment was dependent on prenylation of its CaaX motif, as mutation of the prenylation site caused cytosolic mislocalization. We also showed that PfRab5b, which lacks C-terminal cysteines that are the only known site of Rab GTPase modification, is prenylated. Finally, we show that the THQ class of farnesyltransferase inhibitors abolishes FCP prenylation and causes its mislocalization, providing the first demonstration of a specific prenylated protein disrupted by antimalarial farnesyl transferase inhibitors. Altogether, these findings identify prenylated proteins that reveal unique parasite biology and are useful for evaluating prenyltransferase inhibitors for antimalarial drug development.

Parasites such as Plasmodium and Toxoplasma possess a vestigial plastid homologous to the chloroplasts of algae and plants. The plastid (known as the apicoplast; for apicomplexan plastid) is non-photosynthetic and very much reduced, but has clear endosymbiotic ancestry including a circular genome that encodes RNAs and proteins and a suite of bacterial biosynthetic pathways. Here we review the initial discovery of the apicoplast, and recount the major new insights into apicoplast origin, biogenesis and function. We conclude by examining how the apicoplast can be removed from malaria parasites in vitro, ultimately completing its reduction by chemical supplementation.

Endosymbiosis has driven major molecular and cellular innovations. Plasmodium spp. parasites that cause malaria contain an essential, non-photosynthetic plastid—the apicoplast—which originated from a secondary (eukaryote–eukaryote) endosymbiosis. To discover organellar pathways with evolutionary and biomedical significance, we performed a mutagenesis screen for essential genes required for apicoplast biogenesis in Plasmodium falciparum. Apicoplast(−) mutants were isolated using a chemical rescue that permits conditional disruption of the apicoplast and a new fluorescent reporter for organelle loss. Five candidate genes were validated (out of 12 identified), including a triosephosphate isomerase (TIM)-barrel protein that likely derived from a core metabolic enzyme but evolved a new activity. Our results demonstrate, to our knowledge, the first forward genetic screen to assign essential cellular functions to unannotated P. falciparum genes. A putative TIM-barrel enzyme and other newly identified apicoplast biogenesis proteins open opportunities to discover new mechanisms of organelle biogenesis, molecular evolution underlying eukaryotic diversity, and drug targets against multiple parasitic diseases., The apicoplast of the malaria parasite Plasmodium falciparum provides critical metabolites during host cell infection. An unbiased forward genetic screen identifies previously unknown genes required for biogenesis of this organelle, potentially yielding novel antimalarial drug targets., Author summary Plasmodium parasites, which cause malaria, and related apicomplexan parasites evolved from photosynthetic algae that acquired their chloroplast through two successive endosymbioses. Although no longer photosynthetic, the apicomplexan plastid—or apicoplast—was retained in these pathogens and provides critical metabolites during host cell infection. The apicoplast is of major interest for its unique biology and potential to yield new antimalarial drug targets. Here, we focused on the critical genes required to grow, divide, and inherit new apicoplasts during parasite replication. Given the apicoplast’s divergent evolution, most of these cannot be recognized by their homology to genes with known functions. Instead, we overcame significant technical challenges in the Plasmodium experimental system to perform an unbiased screen to search for these critical genes. Our screen has uncovered new genes with intriguing evolution and function that open up opportunities to understand and ultimately exploit apicoplast biology. Finally, assigning new, essential gene functions in Plasmodium parasites remains a daunting task. The successful identification of essential gene functions using an unbiased approach in this study provides a viable route for expansion of this screen or developing screens for other novel Plasmodium pathways in the future.

This book chapter presents, a review of the literature from 2004-2014 regarding the various models of virtual worlds used in foreign language teaching and learning, the impact of virtual world learning environments and the implications of language teaching. The study being reported aims to address the following questions: (1) What are the models of virtual worlds used in language learning instruction in K-12 and higher education; (2) How do VWLEs impact language learning in terms of motivation, communicative competency, intercultural competency, collaborative competency, constructivist learning, and sociocultural competency; and (3) What are the implications of using VWLEs in foreign language teaching and learning? Results indicate that social context and task-based learning enhanced language learners' participation and motivations. Findings also indicate that consistent use of interactional strategies encouraged learners to engage in the tasks and stay motivated. The study suggested that a VWLE offers a motivating, engaging, and multi-dynamic environment for language learners.

Malaria is a major cause of global childhood mortality. To sustain progress in disease control made in the last decade, new antimalarial therapies are needed to combat emerging drug resistance. Malaria parasites contain a relict chloroplast called the apicoplast, which harbors new targets for drug discovery. Unfortunately, some drugs targeting apicoplast pathways exhibit a delayed-death phenotype, which results in a slow onset-of-action that precludes their use as fast-acting, frontline therapies. Identification of druggable apicoplast biogenesis factors that will avoid the delayed-death phenotype is an important priority. Here, we find that chemical stabilization of an apicoplast-targeted mDHFR domain disrupts apicoplast biogenesis and inhibits parasite growth after a single lytic cycle, suggesting a non-delayed-death target. Our finding indicates that further interrogation of the mechanism-of-action of this exogenous fusion protein may reveal novel therapeutic avenues., Malaria parasites (Plasmodium spp.) contain a nonphotosynthetic plastid organelle called the apicoplast, which houses essential metabolic pathways and is required throughout the parasite life cycle. The biogenesis pathways responsible for apicoplast growth, division, and inheritance are of key interest as potential drug targets. Unfortunately, several known apicoplast biogenesis inhibitors are of limited clinical utility because they cause a peculiar “delayed-death” phenotype in which parasites do not stop replicating until the second lytic cycle posttreatment. Identifying apicoplast biogenesis pathways that avoid the delayed-death phenomenon is a priority. Here, we generated parasites targeting a murine dihydrofolate reductase (mDHFR) domain, which can be conditionally stabilized with the compound WR99210, to the apicoplast. Surprisingly, chemical stabilization of this exogenous fusion protein disrupted parasite growth in an apicoplast-specific manner after a single lytic cycle. WR99210-treated parasites exhibited an apicoplast biogenesis defect beginning within the same lytic cycle as drug treatment, indicating that stabilized mDHFR perturbs a non-delayed-death biogenesis pathway. While the precise mechanism-of-action of the stabilized fusion is still unclear, we hypothesize that it inhibits apicoplast protein import by stalling within and blocking translocons in the apicoplast membranes. IMPORTANCE Malaria is a major cause of global childhood mortality. To sustain progress in disease control made in the last decade, new antimalarial therapies are needed to combat emerging drug resistance. Malaria parasites contain a relict chloroplast called the apicoplast, which harbors new targets for drug discovery. Unfortunately, some drugs targeting apicoplast pathways exhibit a delayed-death phenotype, which results in a slow onset-of-action that precludes their use as fast-acting, frontline therapies. Identification of druggable apicoplast biogenesis factors that will avoid the delayed-death phenotype is an important priority. Here, we find that chemical stabilization of an apicoplast-targeted mDHFR domain disrupts apicoplast biogenesis and inhibits parasite growth after a single lytic cycle, suggesting a non-delayed-death target. Our finding indicates that further interrogation of the mechanism-of-action of this exogenous fusion protein may reveal novel therapeutic avenues.

Hydrolase are enzymes that regulate diverse biological processes, including posttranslational protein modifications. Recent work identified four active serine hydrolases (ASHs) in Toxoplasma gondii as candidate depalmitoylases. However, only TgPPT1 (ASH1) has been confirmed to remove palmitate from proteins. ASH4 (TgME49_264290) was reported to be refractory to genetic disruption. We demonstrate that recombinant ASH4 is an esterase that processes short acyl esters but not palmitoyl thioesters. Genetic disruption of ASH4 causes defects in cell division and premature scission of parasites from residual bodies. These defects lead to the presence of vacuoles with a disordered intravacuolar architecture, with parasites arranged in pairs around multiple residual bodies. Importantly, we found that the deletion of ASH4 correlates with a defect in radial dispersion from host cells after egress. This defect in dispersion of parasites is a general phenomenon that is observed for disordered vacuoles that occur at low frequency in wild-type parasites, suggesting a possible general link between intravacuolar organization and dispersion after egress. IMPORTANCE This work defines the function of an enzyme in the obligate intracellular parasite Toxoplasma gondii. We show that this previously uncharacterized enzyme is critical for aspects of cellular division by the parasite and that loss of this enzyme leads to parasites with cell division defects and which also are disorganized inside their vacuoles. This leads to defects in the ability of the parasite to disseminate from the site of an infection and may have a significant impact on the parasite's overall infectivity of a host organism.

Volume 3, no. 5, e00393-18, 2018, [https://doi.org/10.1128/mSphere.00393-18][1]. The ToxoDB identifier for ASH4 should be TgME49\_262490 instead of TgME49\_264290. The incorrect identifier appears twice in the PDF version of the article: line 5 of the abstract and line 26 of page 2. [1]: /lookup/

Hydrolase are enzymes that regulate diverse biological processes, including posttranslational protein modifications. Recent work identified four active serine hydrolases (ASHs) in

Malaria parasites (Plasmodium spp.) contain a nonphotosynthetic plastid organelle called the apicoplast, which houses essential metabolic pathways and is required throughout the parasite life cycle. Hundreds of proteins are imported across 4 membranes into the apicoplast to support its function and biogenesis. The machinery that mediates this import process is distinct from proteins in the human host and may serve as ideal drug targets. However, a significant concern is whether inhibition of apicoplast protein import will result in a “delayed-death” phenotype that limits clinical use, as observed for inhibitors of apicoplast housekeeping pathways. To assess the growth inhibition kinetics of disrupting apicoplast protein import, we targeted a murine dihydrofolate reductase (mDHFR) domain, which is stabilized by the compound WR99210, to the apicoplast to enable inducible blocking of apicoplast-localized protein translocons. We show that stabilization of this apicoplast-targeted mDHFR disrupts parasite growth within a single lytic cycle in an apicoplast-specific manner. Consistent with inhibition of apicoplast protein import, stabilization of this fusion protein disrupted transit peptide processing of endogenous apicoplast proteins and caused defects in apicoplast biogenesis. These results indicate that disruption of apicoplast protein import avoids delayed-death growth inhibition and that target-based approaches to develop inhibitors of import machinery may yield viable next-generation antimalarials.ImportanceMalaria is a major cause of global childhood mortality. To sustain progress in disease control made in the last decade, new antimalarial therapies are needed to combat emerging drug resistance. Malaria parasites contain a relict chloroplast called the apicoplast, which harbors new targets for drug discovery, including import machinery that transports hundreds of critical proteins into the apicoplast. Unfortunately, some drugs targeting apicoplast pathways show delayed growth inhibition, which results in a slow onset-of-action that precludes their use as fast-acting, frontline therapies. We used a chemical biology approach to disrupt apicoplast protein import and showed that chemical disruption of this pathway avoids delayed growth inhibition. Our finding indicates that prioritization of proteins involved in apicoplast protein import for target-based drug discovery efforts may aid in the development of novel fast-acting antimalarials.

Social media, with its ability to create opportunities for interaction, presents a platform for applying technology into social constructivist learning contexts in innovative and meaningful ways. This chapter proposes a model for integrating mobile social media in a content-based ESL curriculum. Newly-arrived international art students were introduced to popular social media platforms and were trained on how to use these tools to conduct research and document their experiences in the field during a summer intensive program. Results are discussed in terms of effects on students' academic English knowledge and abilities, knowledge of local academic and creative cultures, and technology literacy. Finally, limitations and implications for future studies are explored.

Language educators encounter challenges of integrating meaningful cultural content and assessment into ELL curricula, mainly due to inadequate professional support and training. Regardless of the growing number of young language learners and institutes around the globe, very limited studies have been conducted and published regarding young ELLs' intercultural competence (IC) development. The American Council on the Teaching of Foreign Languages (ACTFL) Global Competence Position Statement (2014) reinforces the importance of teachers helping learners deal with these challenges. This chapter conducted qualitative meta-analysis reviewing the existing literature on the various models of IC teacher preparation programs for young ELLs aiming to improve the IC components in teacher preparation curricula. The main components of IC to be infused into curricula include: (1) the educational needs and state standards of IC in teacher education programs; (2) IC theoretical frameworks and assessment models; and (3) IC practices in teacher education for young ELLs.

The apicoplast is an essential plastid organelle found in Plasmodium parasites which contains several clinically validated antimalarial-drug targets. A chemical rescue screen identified MMV-08138 from the “Malaria Box” library of growth-inhibitory antimalarial compounds as having specific activity against the apicoplast. MMV-08138 inhibition of blood-stage Plasmodium falciparum growth is stereospecific and potent, with the most active diastereomer demonstrating a 50% effective concentration (EC 50 ) of 110 nM. Whole-genome sequencing of 3 drug-resistant parasite populations from two independent selections revealed E688Q and L244I mutations in P. falciparum IspD, an enzyme in the MEP (methyl- d -erythritol-4-phosphate) isoprenoid precursor biosynthesis pathway in the apicoplast. The active diastereomer of MMV-08138 directly inhibited PfIspD activity in vitro with a 50% inhibitory concentration (IC 50 ) of 7.0 nM. MMV-08138 is the first PfIspD inhibitor to be identified and, together with heterologously expressed PfIspD, provides the foundation for further development of this promising antimalarial drug candidate lead. Furthermore, this report validates the use of the apicoplast chemical rescue screen coupled with target elucidation as a discovery tool to identify specific apicoplast-targeting compounds with new mechanisms of action.