Malaria-associated pathogenesis such as parasite invasion, egress, host cell remodelling and antigenic variation requires concerted action by many proteins, but the molecular regulation is poorly understood. Here we have characterized an essential Plasmodium-specific Apicomplexan AP2 transcription factor in Plasmodium falciparum (PfAP2-P; pathogenesis) during the blood-stage development with two peaks of expression. An inducible knockout of gene function showed that PfAP2-P is essential for trophozoite development, and critical for var gene regulation, merozoite development and parasite egress. Chromatin immunoprecipitation sequencing data collected at timepoints matching the two peaks of pfap2-p expression demonstrate PfAP2-P binding to promoters of genes controlling trophozoite development, host cell remodelling, antigenic variation and pathogenicity. Single-cell RNA sequencing and fluorescence-activated cell sorting revealed de-repression of most var genes in Δpfap2-p parasites. Δpfap2-p parasites also overexpress early gametocyte marker genes, indicating a regulatory role in sexual stage conversion. We conclude that PfAP2-P is an essential upstream transcriptional regulator at two distinct stages of the intra-erythrocytic development cycle.

Plants continuously adapt to their environments by responding to various intrinsic and extrinsic signals. They face numerous biotic and abiotic stresses such as extreme temperatures, drought, or pathogens, requiring complex regulatory mechanisms to control gene activity and adapt their proteome for survival. Epigenetic regulation plays a crucial role in these adaptations, potentially leading to both heritable and non-heritable changes across generations. This process enables plants to adjust their gene expression profiles and acclimate effectively. It is also vital for plant development and productivity, affecting growth, yield, and seed quality, and enabling plants to “remember” environmental stimuli and adapt accordingly. Key epigenetic mechanisms that play significant roles include DNA methylation, histone modification, and ubiquitin ligase complex activity. These processes, which have been extensively studied in the last two decades, have led to a better understanding of the underlying mechanisms and expanded the potential for improving agriculturally and economically important plant traits. DNA methylation is a fundamental process that regulates gene expression by altering chromatin structure. The addition of methyl groups to cytosines by DNA methylases leads to gene suppression, whereas DNA demethylases reverse this effect. Histone modifications, on the other hand, collectively referred to as the “histone code”, influence chromatin structure and gene activity by promoting either gene transcription or gene silencing. These modifications are either recognized, added, or removed by a variety of enzymes that act practically as an environmental memory, having a significant impact on plant development and the responses of plants to environmental stimuli. Finally, ubiquitin ligase complexes, which tag specific histones or regulatory proteins with ubiquitin, are also crucial in plant epigenetic regulation. These complexes are involved in protein degradation and play important roles in regulating various cellular activities. The intricate interplay between DNA methylation, histone modifications, and ubiquitin ligases adds complexity to our understanding of epigenetic regulation. These mechanisms collectively control gene expression, generating a complex and branching network of interdependent regulatory pathways. A deeper understanding of this complex network that helps plants adapt to environmental changes and stressful conditions will provide valuable insights into the regulatory mechanisms involved. This knowledge could pave the way for new biotechnological approaches and plant breeding strategies aimed at enhancing crop resilience, productivity, and sustainable agriculture.

In contrast to the short-term (ST) CD34+ stem cells, studies have suggested that long-term (LT) hematopoietic stem cells (HSCs) found in the CD34− stem cell pool have trouble migrating and engrafting when introduced through IV. To understand why these deficiencies exist, we set out to fully elucidate the adhesion mechanisms used by ST and LT-HSCs to migrate to the bone marrow(BM). Specifically focusing on murine ST-HSCs (Flk2−CD34+) and LT-HSCs (Flk2−CD34−), we observed a distinctive expression pattern of BM homing effectors necessary for the first step, namely sialyl Lewis-X (sLex) (ligand for E-selectin), and the second step, namely CXCR4 chemokine receptor (receptor for SDF-1). sLex expression was higher on Flk2−CD34+ ST-HSCs (>60%) compared with Flk2−CD34− LT-HSCs (<10%), which correlated to binding to E-selectin. Higher concentrations of CXCR4 were observed on Flk2−CD34+ ST-HSCs compared with Flk2−CD34− LT-HSCs. Interestingly, the expression of CD26, a peptidase known to deactivate chemokines (ie, SDF-1), was higher on Flk2−CD34− LT-HSCs. Given that both E-selectin–binding and CXCR4-mediated migration are compromised in Flk2−CD34− LT-HSCs, we aimed to enhance their ability to migrate using recombinant human fucosyltransferase 6 (rhFTVI) and the CD26 inhibitor, Dip A (diprotin A). To this end, we observed that although LT-HSCs expressed low concentrations of sLex, they were able to engraft when transplanted into recipient mice. Moreover, although both CD26 inhibition and fucosylation enhanced migration of both HSC populations in vitro, only pretreatment of LT-HSCs with Dip A enhanced engraftment in vivo after transplantation into recipient mice. Remarkably, fucosylation of Flk2−CD34+ ST-HSCs consistently led to their ability to transplant secondary recipients. These data suggest that using fucosylation and Dip A to overcome the molecular disparity in adhesion mechanisms among ST-HSCs and LT-HSCs differentially influences their abilities to migrate and engraft in vivo and promotes the ability of ST-HSCs to engraft secondary recipient mice, the gold standard for testing functionality of LT-HSCs.

The polyhistidine tag (His-tag) is one of the most popular protein tags used in the life sciences. Traditionally, the detection of His-tagged proteins relies on immunoblotting with anti-His antibodies. This approach is laborious for certain applications, such as protein purification, where time and simplicity are critical. The His-tag can also be directly detected by metal ion–loaded nickel-nitrilotriacetic acid–based chelator heads conjugated to fluorophores, which is a convenient alternative method to immunoblotting. Typically, such chelator heads are conjugated to either green or red fluorophores, the detection of which requires specialized excitation sources and detection systems. Here, we demonstrate that post-run staining is ideal for His-tag detection by metal ion–loaded and fluorescently labeled chelator heads in PAGE and blot membranes. Additionally, by comparing the performances of different chelator heads, we show how differences in microscopic affinity constants translate to macroscopic differences in the detection limits in environments with limited diffusion, such as PAGE. On the basis of these results, we devise a simple approach, called UVHis-PAGE, that uses metal ion–loaded and fluorescently labeled chelator heads to detect His-tagged proteins in PAGE and blot membranes. Our method uses a UV transilluminator as an excitation source, and the results can be visually inspected by the naked eye.

Circuitries of signaling pathways integrate distinct hormonal and environmental signals, and influence development in plants. While a crosstalk between brassinosteroid (BR) and gibberellin (GA) signaling pathways has recently been established, little is known about other components engaged in the integration of the two pathways. Here, we provide supporting evidence for the role of HSP90 (HEAT SHOCK PROTEIN 90) in regulating the interplay of the GA and BR signaling pathways to control hypocotyl elongation of etiolated seedlings in Arabidopsis. Both pharmacological and genetic depletion of HSP90 alter the expression of GA biosynthesis and catabolism genes. Major components of the GA pathway, like RGA (REPRESSOR of ga1–3) and GAI (GA-INSENSITIVE) DELLA proteins, have been identified as physically interacting with HSP90. Interestingly, GA-promoted DELLA degradation depends on the ATPase activity of HSP90, and inhibition of HSP90 function stabilizes the DELLA/BZR1 (BRASSINAZOLE-RESISTANT 1) complex, modifying the expression of downstream transcriptional targets. Our results collectively reveal that HSP90, through physical interactions with DELLA proteins and BZR1, modulates DELLA abundance and regulates the expression of BZR1-dependent transcriptional targets to promote plant growth.

WD40 repeat proteins (WDRs) are present in all eukaryotes and include members that are implicated in numerous cellular activities. They act as scaffold proteins and thus as molecular “hubs” for protein–protein interactions, which mediate the assembly of multifunctional complexes that regulate key developmental processes in Arabidopsis thaliana, such as flowering time, hormonal signaling, and stress responses. Despite their importance, many aspects of their putative functions have not been elucidated yet. Here, we show that the late-flowering phenotype of the anthesis promoting factor 1 (aprf1) mutants is temperature-dependent and can be suppressed when plants are grown under mild heat stress conditions. To gain further insight into the mechanism of APRF1 function, we employed a co-immunoprecipitation (Co-IP) approach to identify its interaction partners. We provide the first interactome of APRF1, which includes proteins that are localized in several subcellular compartments and are implicated in diverse cellular functions. The dual nucleocytoplasmic localization of ARRF1, which was validated through the interaction of APRF1 with HEAT SHOCK PROTEIN 1 (HSP90.1) in the nucleus and with HSP90.2 in the cytoplasm, indicates a dynamic and versatile involvement of APRF1 in multiple biological processes. The specific interaction of APRF1 with the chaperon HSP90.1 in the nucleus expands our knowledge regarding the epigenetic regulation of flowering time in A. thaliana and further suggests the existence of a delicate thermoregulated mechanism during anthesis.