Stage-specific Expression Research Articles

Expression patterns of MIH, EcR2, and RXR3 in the moult cycle of the red swamp crayfish, Procambarus clarkii (Girard, 1852) (Decapoda, Cambaridae)

Abstract In arthropods, the moult-inhibiting hormone (MIH), the ecdysone receptor (EcR), and the retinoid X receptor (RXR) are key regulators in moulting. In the present study, the full-length cDNAs of the MIH, EcR2, and RXR3 genes from the red swamp crayfish, Procambarus clarkii (Girard, 1852) (denoted as PcMIH, PcEcR2, and PcRXR3) were cloned. Tissue-specific and moult stage-specific mRNA expression patterns of these genes were detected by real-time quantitative polymerase chain reaction. PcMIH was detected only in the eyestalk, whereas PcEcR2 and PcRXR3 mRNA were expressed in all tissues tested. The highest levels of PcEcR2 and PcRXR3 were detected in the gill and hepatopancreas. Expression of PcMIH mRNA in the eyestalk increased from postmoult to peak in intermoult and then decreased in premoult. Expression of PcEcR2 mRNA in the eyestalk, hepatopancreas, and muscle increased from postmoult to peak in early premoult and then decreased. However, expression of PcEcR2 mRNA in the gill increased from postmoult to reach a maximum in intermoult and then decreased in premoult. Expression of PcRXR3 mRNA also fluctuated in the eyestalk, hepatopancreas, muscle, and gill, with a decrease from postmoult to late premoult. Expression of PcEcR2 and PcRXR3 mRNA increased relative to the control in the hepatopancreas and gill after unilateral and bilateral eyestalk ablation, which suggested that PcMIH can inhibit their mRNA expression. Double-stranded RNA-mediated RNA interference of PcRXR3 caused different changes in mRNA expression of these genes in different tissues and resulted in decreased expression of PcEcR2 mRNA, which suggested a collaborative relationship between PcEcR2 and PcRXR3.

A low molecular-weight cyclophilin localizes in different cell compartments of Pyrus communis pollen and is released in vitro under Ca2+ depletion

Cyclophilins (CyPs) are ubiquitous proteins involved in a wide variety of processes including protein maturation and trafficking, receptor complex stabilization, apoptosis, receptor signaling, RNA processing, and spliceosome assembly. The ubiquitous presence is justified by their peptidyl-prolyl cis-trans isomerase (PPIase) activity, catalyzing the rotation of X-Pro peptide bonds from a cis to a trans conformation, a critical rate-limiting step in protein folding, as over 90% of proteins contain trans prolyl imide bonds. In Arabidopsis 35 CyPs involved in plant development have been reported, showing different subcellular localizations and tissue- and stage-specific expression. In the present work, we focused on the localization of CyPs in pear (Pyrus communis) pollen, a model system for studies on pollen tube elongation and on pollen-pistil self-incompatibility response. Fluorescent, confocal and immuno-electron microscopy showed that this protein is present in the cytoplasm, organelles and cell wall, as confirmed by protein fractionation. Moreover, an 18-kDa CyP isoform was specifically released extracellularly when pear pollen was incubated with the Ca2+ chelator EGTA.

Transcriptome analysis of Plasmodium berghei during exo-erythrocytic development

BackgroundThe complex life cycle of malaria parasites requires well-orchestrated stage specific gene expression. In the vertebrate host the parasites grow and multiply by schizogony in two different environments: within erythrocytes and within hepatocytes. Whereas erythrocytic parasites are well-studied in this respect, relatively little is known about the exo-erythrocytic stages.MethodsIn an attempt to fill this gap, genome wide RNA-seq analyses of various exo-erythrocytic stages of Plasmodium berghei including sporozoites, samples from a time-course of liver stage development and detached cells were performed. These latter contain infectious merozoites and represent the final step in exo-erythrocytic development.ResultsThe analysis represents the complete transcriptome of the entire life cycle of P. berghei parasites with temporal detailed analysis of the liver stage allowing comparison of gene expression across the progression of the life cycle. These RNA-seq data from different developmental stages were used to cluster genes with similar expression profiles, in order to infer their functions. A comparison with published data from other parasite stages confirmed stage-specific gene expression and revealed numerous genes that are expressed differentially in blood and exo-erythrocytic stages. One of the most exo-erythrocytic stage-specific genes was PBANKA_1003900, which has previously been annotated as a “gametocyte specific protein”. The promoter of this gene drove high GFP expression in exo-erythrocytic stages, confirming its expression profile seen by RNA-seq.ConclusionsThe comparative analysis of the genome wide mRNA expression profiles of erythrocytic and different exo-erythrocytic stages could be used to improve the understanding of gene regulation in Plasmodium parasites and can be used to model exo-erythrocytic stage metabolic networks toward the identification of differences in metabolic processes during schizogony in erythrocytes and hepatocytes.

Molecular Analysis of Iron Deficiency Response in Hexaploid Wheat

Iron deficiency leads to severe chlorosis in crop plants, including wheat, thereby reducing total yield and quality. Furthermore, grains of most bread wheat varieties are poor source of iron, which is vital for human nutrition. Despite the significance, iron uptake and translocation mechanisms in bread wheat have not been studied in detail, particularly under iron limited growth conditions. In this study, bread wheat plants were grown under iron deficiency stress until maturity. Data were collected at three distinct developmental time points during grain-filling. The plants experiencing low iron availability exhibited significantly lower chlorophyll content as well as low iron concentration in leaves and grains. The expression levels of bread wheat genes homologous to iron deficiency responsive genes of rice, barley, and Arabidopsis were significantly changed under iron deficiency stress. The wheat homologs of genes involved in phytosiderophore (PS) synthesis and transport were significantly up-regulated in the iron-deficient roots through all development stages, confirming an important role of deoxymugineic acid (DMA) in iron acquisition. The up-regulation of NICOTIANAMINE SYNTHASE (NAS) and DEOXYMUGINEIC ACID SYNTHASE (DMAS) in flag leaves and grains suggested the involvement of nicotianamine (NA) and DMA in iron chelation and translocation in wheat, particularly at the commencement of grain-filling. In line with this, the homolog of gene encoding TRANSPORTER OF MUGINEIC ACID (TOM) was up-regulated in the wheat roots under iron deficiency. Additionally, genes encoding long-distance iron transporter YELLOW STRIPE-LIKE (YSL), the vacuolar transporter NATURAL RESISTANCE ASSOCIATED MACROPHAGE PROTEIN (NRAMP), and the transcription factor BASIC HELIX-LOOP-HELIX (bHLH), were also up-regulated upon iron starvation. A tissue specific and growth stage specific gene expression differences in response to iron deficiency stress were observed, providing new insights into iron translocation, storage and regulation in bread wheat.

Suppression of Gene Juvenile Hormone Diol Kinase Delays Pupation in Heortia vitessoides Moore.

Juvenile hormone diol kinase (JHDK) is a critical enzyme involved in juvenile hormone degradation in insects. In this study, HvJHDK in the Heortia vitessoides Moore (Lepidoptera: Crambidae) transcriptional library was cloned. Stage-specific expression patterns of HvJHDK, HvJHEH, and HvJHE as well as juvenile hormone titers were determined. The three tested enzymes participated in juvenile hormone degradation. Moreover, juvenile hormone titers peaked after larval–larval molts, consistent with a role for juvenile hormone in inhibition of metamorphosis. HvJHDK was subsequently suppressed using RNA interference (RNAi) to reveal its functions. Different concentrations of dsJHDK elicited the optimal interference efficiency at different life stages of H. vitessoides. Suppression of HvJHDK decreased HvJHDK content and increased the juvenile hormone titer, thereby resulting in reduced triglyceride content, sharply declined survival rate, clearly lethal phenotypes, and extended larval growth. Moreover, suppression of HvJHDK upregulated HvJHEH and HvJHE expression levels, suggesting that there is feedback regulation in the juvenile hormone metabolic pathway. Taken together, our findings provide molecular references for the selection of novel insecticidal targets.

Vitellogenins - Yolk Gene Function and Regulation in Caenorhabditis elegans.

Vitellogenins are a family of yolk proteins that are by far the most abundant among oviparous animals. In the model nematode Caenorhabditis elegans, the 6 vitellogenins are among the most highly expressed genes in the adult hermaphrodite intestine, which produces copious yolk to provision eggs. In this article we review what is known about the vitellogenin genes and proteins in C. elegans, in comparison with vitellogenins in other taxa. We argue that the primary purpose of abundant vitellogenesis in C. elegans is to support post-embryonic development and fertility, rather than embryogenesis, especially in harsh environments. Increasing vitellogenin provisioning underlies several post-embryonic phenotypic alterations associated with advancing maternal age, demonstrating that vitellogenins can act as an intergenerational signal mediating the influence of parental physiology on progeny. We also review what is known about vitellogenin regulation – how tissue-, sex- and stage-specificity of expression is achieved, how vitellogenins are regulated by major signaling pathways, how vitellogenin expression is affected by extra-intestinal tissues and how environmental experience affects vitellogenesis. Lastly, we speculate whether C. elegans vitellogenins may play other roles in worm physiology.

Gene expression during genital morphogenesis in the ground beetle Carabus maiyasanus.

To investigate the developmental genetics of genital formation in the carabid beetle Carabus maiyasanus, we compared gene expression patterns among five stages using transcriptomic RNA sequencing data from abdominal segments and genitalia in the third (last) larval instar (including prepupa) and pupal stages. We identified 18 839 genes, of which 10 796 were differentially expressed among stages or between sexes. There were relatively few differentially expressed genes (DEGs) between the sexes (3%). The DEGs were clustered into six groups, mainly according to stage-specific expression patterns. Genes in clusters 1-3 showed high expression levels before pupation and low expression levels during the pupal period, whereas genes in clusters 4-6 showed high expression levels from the prepupal to the pupal stages. Genes related to the initial pupation process and differentiation of genital discs in Drosophila were involved in clusters 4 and 6 and showed low expression levels at early third instar and elevated expression levels from the early prepupal stage, suggesting that the pupation process and genital differentiation started in the prepupal stage. Clusters 4 and 5 included developmental genes related to organ size control, which may be important in the formation of internal genital structures during the pupal stage.

Expression of Kisspeptin and its receptor in the hypothalamus of cyclic and acyclic buffalo (Bubalus bubalis)

Kisspeptin (Kiss1), neurokinin-B (NKB) and dynorphin (Dyn) neurons regulate the surge and pulsatile centres of gonadotropin releasing hormone (GnRH) in the hypothalamus and are modulated by the ovarian steroids. Accordingly, we studied the temporospatial expression of Kiss1, its receptor and other genes that regulate GnRH in the preoptic area (POA) and arcuate (ARC) regions of hypothalamus at different phases of bubaline estrous cycle. Brain of buffalo (n = 32) was collected immediately after exsanguination and categorized into early luteal (EL), mid luteal (ML), follicular (FL) stages and acyclic (n = 8/group). Total RNA was extracted from the POA and ARC of each stage and real time PCR amplification of Kiss1, Kiss1r, NKB, NKB receptor (NKBR), Dyn, Dyn receptor (OPRK1), GnRH1, ERα, PR, LEPR and GHSR was done using GAPDH as endogenous control and acyclic stage as calibrator group. Further, immunolocalization of Kiss1 and Kiss1r was done on the hypothalamus. In the POA, significant up-regulation of Kiss1 and NKB with a concomitant down-regulation of Dyn transcripts was recorded at FL stage. There was, however, down-regulation of Kiss1 and Kiss1r during the EL perhaps due to the loss of estradiol as a consequence of ovulation. On the other hand, in the ARC, there was a significant up-regulation of Kiss1 and Dyn at FL and ML, while NKB transcript was consistently down-regulated at any stage of estrous cycle. In the POA, expression of ERα was not modulated; however, PR was down-regulated in the EL. In the ARC, the ERα expression was significantly up-regulated in the EL, whereas, PR was moderately expressed irrespective of the stage of estrous cycle. The immunolocalization study revealed the presence of Kiss1 and Kiss1r in the POA and ARC in the cyclic buffalo with relative abundance at FL. The transcriptional profile of the genes suggests that there is estrous cycle stage specific expression of Kiss1, Kiss1r and other GnRH regulating genes in the POA and ARC regions of hypothalamus in the buffalo. Up-regulation of Kiss1r in the POA during ML and ARC during EL indicates the involvement of kisspeptinergic system in the regulation of low LH pulse frequencies during the early and mid luteal phases in the cyclic buffalo.

Malaria.tools—comparative genomic and transcriptomic database for Plasmodium species

Malaria is a tropical parasitic disease caused by the Plasmodium genus, which resulted in an estimated 219 million cases of malaria and 435 000 malaria-related deaths in 2017. Despite the availability of the Plasmodium falciparum genome since 2002, 74% of the genes remain uncharacterized. To remedy this paucity of functional information, we used transcriptomic data to build gene co-expression networks for two Plasmodium species (P. falciparum and P. berghei), and included genomic data of four other Plasmodium species, P. yoelii, P. knowlesi, P. vivax and P. cynomolgi, as well as two non-Plasmodium species from the Apicomplexa, Toxoplasma gondii and Theileria parva. The genomic and transcriptomic data were incorporated into the resulting database, malaria.tools, which is preloaded with tools that allow the identification and cross-species comparison of co-expressed gene neighbourhoods, clusters and life stage-specific expression, thus providing sophisticated tools to predict gene function. Moreover, we exemplify how the tools can be used to easily identify genes relevant for pathogenicity and various life stages of the malaria parasite. The database is freely available at www.malaria.tools.

Landscape of Enhancer-Enhancer Cooperative Regulation during Human Cardiac Commitment

Although accumulating evidence has demonstrated the key roles of enhancers in gene expression regulation, the contribution of genome-wide enhancer-enhancer interactions to developmental decisions remains unclear. Here we explored the cooperative regulation patterns among enhancers to understand their regulatory mechanism. We first filtered robust enhancers in embryonic stem cells (ESCs) through integrating bidirectional transcription, genomic location, and epigenetic modification. Genome-wide enhancer-enhancer interactions were then identified based on enhancer-promoter relationships that were derived from Hi-C data. We further explored the interacting principles of the identified enhancer-enhancer interactions. The results revealed that the observed cooperativity occurred mainly between enhancers distributed within a 1-kb to 10-Mb distance across the genome. In addition, enhancer-enhancer pairs had higher expression correlations than non-interacting pairs. Finally, we identified robust enhancers during human cardiac commitment, and we found that enhancers exhibited strong stage-specific expression patterns. We further inferred the enhancer-enhancer interactions based on RNA sequencing (RNA-seq) data in heart development, according to the regulatory principles characterized from Hi-C data. The identified enhancer-enhancer interaction networks (EEINs) presented highly dynamic linkages. Moreover, enhancers cooperatively targeted many marker genes in each developmental stage to regulate stage-specific functions, which contribute to the organization of cell identity in heart development. Our work will increase the understanding of enhancer regulation in human heart development.

Implication for DNA methylation involved in the host transfer of diamondback moth, Plutella xylostella (L.).

DNA methylation exerts extensive impacts on gene expression of various living organisms exposed to environmental variation. However, little is known whether DNA methylation is involved in the host transfer of diamondback moth, Plutella xylostella (L.), a worldwide destructive pest of crucifers. In this study, we found that P. xylostella genome exhibited a relatively low level of DNA methylation on the basis of the CpG O/E prediction and experimental validation. A significant positive linear correlation was observed between the stage-specific expressions of PxDNMT1 and DNA methylation levels (5mC content). Particularly, high levels of DNA methylation and gene expression of PxDNMT1 were observed in eggs and mature females of P. xylostella. After host transfer of P. xylostella from Raphanus sativus to Arabidopsis thaliana, we identified some potential genomic loci that might have changed methylation levels. Using the method of fluorescence-labeled methylation-sensitive amplified polymorphism (F-MSAP), we also found the corresponding genes primarily involved in neural system and signaling. The expressions of six candidate genes were verified by qRT-PCR. One of the genes, Px009600, might be regulated by a DNA methylation-mediated mechanism in response to host transfer. Our study provides evidence for a functional system of DNA methylation in P. xylostella and its possible role in adaptation during host transfer. Further studies should examine methylation as responsive factors to different host plants and environmental cues in insect pests.

Endometrial luminal epithelial cells sense embryo elongation in the roe deer independent of interferon-tau†.

Numerous intrauterine changes take place across species during embryo development. Following fertilization in July/August, the European roe deer (Capreolus capreolus) embryo undergoes diapause until embryonic elongation in December/January. Embryonic elongation prior to implantation is a common feature among ungulates. Unlike many other ruminants, the roe deer embryo does not secrete interferon-tau (IFNτ). This provides the unique opportunity to unravel IFNτ-independent signaling pathways associated with maternal recognition of pregnancy (MRP). This study aimed at identifying the cell-type-specific endometrial gene expression changes associated with the MRP at the time of embryo elongation that are independent of IFNτ in roe deer. The messenger RNA (mRNA) expression of genes known to be involved in embryo-maternal communication in cattle, pig, sheep, and mice was analyzed in laser capture microdissected (LMD) endometrial luminal, glandular epithelial, as well as stromal cells. The mRNA transcript abundances of the estrogen (ESR1), progesterone receptor (PGR), and IFNτ-stimulated genes were lower in the luminal epithelium in the presence of an elongated embryo compared to diapause. Retinol Binding Protein-4 (RBP4), a key factor involved in placentation, was more abundant in the luminal epithelium in the presence of an elongated embryo. The progesterone receptor localization was visualized by immunohistochemistry, showing an absence in the luminal epithelium and an overall lower abundance with time and thus prolonged progesterone exposure. Our data show a developmental stage-specific mRNA expression pattern in the luminal epithelium, indicating that these cells sense the presence of an elongated embryo in an IFNτ-independent manner.

Accelerated photoreceptor differentiation of hiPSC-derived retinal organoids by contact co-culture with retinal pigment epithelium

Retinal organoids (ROs) derived from human-induced pluripotent stem cells recapitulate the three-dimensional structure of retina, mimic human retinal development, and provide cell sources for pre-clinical retinal transplantation. Retinal pigment epithelium (RPE) is crucial for normal outer retinal physiology, including phagocytosis of shed photoreceptor outer segments and secretion of neurotrophic and vasculotrophic growth factors. However, whether ROs-RPE co-culture can improve the differentiation of photoreceptors in ROs in vitro remains unknown. Herein, primary mouse RPE cells were contact co-cultured with ROs at different time points. Our results revealed that the RPE cells accelerated photoreceptor differentiation in ROs, as the cross talk between the RPE and ROs promoted the stage specific expression of photoreceptor markers at different differentiation stages. Thus, we established an improved co-culture system based on modeling of human retina-RPE dynamics during retinogenesis for the evaluation of ocular therapies.

Dephosphorylation of HDAC4 by PP2A-B\u03b4 unravels a new role for the HDAC4/MEF2 axis in myoblast fusion

Muscle formation is controlled by a number of key myogenic transcriptional regulators that govern stage-specific gene expression programs and act as terminal effectors of intracellular signaling pathways. To date, the role of phosphatases in the signaling cascades instructing muscle development remains poorly understood. Here, we show that a specific PP2A-B55δ holoenzyme is necessary for skeletal myogenesis. The primary role of PP2A-B55δ is to dephosphorylate histone deacetylase 4 (HDAC4) following myocyte differentiation and ensure repression of Myocyte enhancer factor 2D (MEF2D)-dependent gene expression programs during myogenic fusion. As a crucial HDAC4/MEF2D target gene that governs myocyte fusion, we identify ArgBP2, an upstream inhibitor of Abl, which itself is a repressor of CrkII signaling. Consequently, cells lacking PP2A-B55δ show upregulation of ArgBP2 and hyperactivation of CrkII downstream effectors, including Rac1 and FAK, precluding cytoskeletal and membrane rearrangements associated with myoblast fusion. Both in vitro and in zebrafish, loss-of-function of PP2A-B55δ severely impairs fusion of myocytes and formation of multinucleated muscle fibers, without affecting myoblast differentiation. Taken together, our results establish PP2A-B55δ as the first protein phosphatase to be involved in myoblast fusion and suggest that reversible phosphorylation of HDAC4 may coordinate differentiation and fusion events during myogenesis.

Cloudy with a Chance of Insights: Context Dependent Gene Regulation and Implications for Evolutionary Studies.

Research in various fields of evolutionary biology has shown that divergence in gene expression is a key driver for phenotypic evolution. An exceptional contribution of cis-regulatory divergence has been found to contribute to morphological diversification. In the light of these findings, the analysis of genome-wide expression data has become one of the central tools to link genotype and phenotype information on a more mechanistic level. However, in many studies, especially if general conclusions are drawn from such data, a key feature of gene regulation is often neglected. With our article, we want to raise awareness that gene regulation and thus gene expression is highly context dependent. Genes show tissue- and stage-specific expression. We argue that the regulatory context must be considered in comparative expression studies.

Temporal expression of floral proteins interacting with CArG1 region of CsAP3 gene in Crocus sativus L.

Crocus sativus L. (Saffron), a monocot triploid species belonging to the Iridaceae family, is also called golden crop owing to its precious stigmas enriched with flavanoids Crocin, Picocrocin and Safranal. Unlike majority of plants which follow typical ABC model of floral development, deviations from ABC model have been reported in Saffron flowers which could probably be implicated in growth and development of distinct Saffron stigmas. CsAP3 is a critical gene regulating stigma development in Saffron. Its promoter consists of three CArG regions which play pivotal role in the expression of AP3 gene, of which CArG1 is the binding site for activator proteins thus regulating floral growth. Keeping in view the growth regulating characteristics of CArG1, we choose to identify the nuclear factors binding to this region in Saffron. Purified nuclear proteins were allowed to bind synthetic CArG1 sequence and the complex was subjected to protein identification using LCMS. Out of the 5 major protein hits in LCMS, NAC-like protein (NAP) was identified as a conspicuous homeotic protein interacting with CArG1 region of AP3 promoter. Gene expression analysis of CsAP3 and CsNAP at different stages of flower development showed stage-specific expression of both the genes however, no direct correlation between CsAP3 and CsNAP gene expression could be observed, even though both these genes are present in the same pathway and expressed at same time. It is possible that CsNAP may be regulating CsAP3 expression via some other mediators, indirectly which needs further molecular profiling.

Induction of Apoptosis by Pierisin-6 in HPV Positive HeLa and HepG2 Cancer Cells is Mediated by the Caspase-3 Dependent Mitochondrial Pathway.

To explore the cytotoxic and apoptotic activity of the pierisin-6 protein in HPV HeLa and HepG2 cell lines. In this study, isolation, and purification of cytotoxic Prierisin-6 from the larvae of Pieris napi by affinity column chromatography techniques. Characterization of full-length mRNA of pierisin-6 gene was performed using 3'/5' RACE PCR. The quantitative RT-PCR used to study the developmental stage-specific expression of pierisin-6 mRNA. The most effective concentration of Pierisin-6 protein was determined by measuring cell proliferation. Apoptosis was assessed using AO/Et-Br, Propidium Iodide, and Rhodamine 123 assays, whereas protein levels of caspase 3, cytochrome C were evaluated by ELISA method. Pierisin-6 induced cell cycle arrest was determined using Propidium iodide by FACS. In this study, Pierisin-6, a novel apoptotic protein was found to have cytotoxicity against HeLa, HepG2 human cancer cell lines and L-132 human lung epithelial cell line. Among the target cells, HeLa was the most sensitive to Pierisin-6. Flow cytometry analysis confirms an increased percentage of apoptotic cells in sub G1 phase and cell cycle arrest at S phase. Alteration in the transmembrane potential of mitochondria, Cytochrome c released from the mitochondrial membrane, and caspase substrate assay demonstrated the cleavage of Ac- DEVD-pNA signifying the activation of Caspase-3. These findings suggested that Pierisin-6 significantly induce apoptosis in HeLa and HepG2 cells and is attributed mainly through a mitochondrial pathway by activation of caspases. The developmental and stage-specific expression of pierisin-6 mRNA was one thousand-fold increased from second to third instar larvae and gradually declined before pupation. Pierisin-6 represents a promising therapeutic approach for liver cancer patients.

Role of the Ubiquitin Proteasome System (UPS) in the HIV-1 Life Cycle.

Given that the ubiquitin proteasome system (UPS) is the major protein degradation process in the regulation of a wide variety of cellular processes in eukaryotic cells, including alteration of cellular location, modulation of protein activity, and regulation of protein interaction, it is reasonable to suggest that the infecting HIV-1 and the invaded hosts exploit the UPS in a contest for survival and proliferation. However, to date, regulation of the HIV-1 life cycle has been mainly explained by the stage-specific expression of HIV-1 viral genes, not by elimination processes of the synthesized proteins after completion of their duties in the infected cells, which is also quintessential for understanding the molecular processes of the virus life cycle and thereby HIV-1 pathogenesis. In fact, several previous publications have indicated that the UPS plays a critical role in the regulation of the proteasomal degradation of viral and cellular counterparts at every step of the HIV-1 life cycle, from the virus entry to release of the assembled virus particles, which is integral for the regulation of survival and proliferation of the infecting HIV-1 and to replication restriction of the invading virus in the host. However, it is unknown whether and how these individual events taking place at different stages of the HIV-1 life cycle are orchestrated as an overall strategy to overcome the restrictions conferred by the host cells. Thus, in this review, we overview the interplay between HIV-1 viral and cellular proteins for restrictions/competitions for proliferation of the virus in the infected cell, which could open a new avenue for the development of therapeutics against HIV-1 via targeting a specific step of the proteasome degradation pathway during the HIV-1 life cycle.

Nuclear Factor Y (NF-Y) Modulates Encystation in Entamoeba via Stage-Specific Expression of the NF-YB and NF-YC Subunits.

Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor composed of three subunits, namely, NF-YA, NF-YB, and NF-YC, which are conserved throughout evolution. In higher eukaryotes, NF-Y plays important roles in several cellular processes (development, cell cycle regulation, apoptosis, and response to growth, stress, and DNA damage) by controlling gene expression through binding to a CCAAT promoter motif. We demonstrated that NF-Y subunits in the protist Entamoeba, while significantly divergent from those of higher eukaryotes, have well-conserved domains important for subunit interactions and DNA binding and that NF-YB and NF-YC are developmentally expressed during encystation. Electrophoretic mobility shift assays confirmed that the NF-Y protein(s) from Entamoeba cysts binds to a CCAAT motif. Consistent with a role as a transcription factor, the NF-Y proteins show nuclear localization during development. Additionally, we demonstrated that NF-YC localizes to the chromatoid body (an RNA processing center) during development, indicating that it may have a role in RNA processing. Finally, silencing of the NF-YC subunit resulted in reduced stability of the NF-Y complex and decreased encystation efficiency. We demonstrated that the NF-Y complex functions at a time point subsequent to the NAD+ flux and expression of the transcription factor encystation regulatory motif-binding protein, both of which are early regulators of Entamoeba development. Taken together, our results demonstrate that the NF-Y complex plays an important role in regulating encystation in Entamoeba and add to our understanding of the transcriptional networks and signals that control this essential developmental pathway in an important human pathogen.IMPORTANCE The human parasite Entamoeba histolytica is an important pathogen with significant global impact and is a leading cause of parasitic death in humans. Since only the cyst form can be transmitted, blocking encystation would prevent new infections, making the encystation pathway an attractive target for the development of new drugs. Identification of the genetic signals and transcriptional regulatory networks that control encystation would be an important advance in understanding the developmental cascade. We show that the Entamoeba NF-Y complex plays a crucial role in regulating the encystation process in Entamoeba.

Expression dynamics of repetitive DNA in early human embryonic development

BackgroundThe last decade witnessed a number of genome-wide studies on human pre-implantation, which mostly focused on genes and provided only limited information on repeats, excluding the satellites. Considering the fact that repeats constitute a large portion of our genome with reported links to human physiology and disease, a thorough understanding of their spatiotemporal regulation during human embryogenesis will give invaluable clues on chromatin dynamics across time and space. Therefore, we performed a detailed expression analysis of all repetitive DNA elements including the satellites across stages of human pre-implantation and embryonic stem cells.ResultsWe uncovered stage-specific expressions of more than a thousand repeat elements whose expressions fluctuated with a mild global decrease at the blastocyst stage. Most satellites were highly expressed at the 4-cell level and expressions of ACRO1 and D20S16 specifically peaked at this point. Whereas all members of the SVA elements were highly upregulated at 8-cell and morula stages, other transposons and small RNA repeats exhibited a high level of variation among their specific subtypes. Our repeat enrichment analysis in gene promoters coupled with expression correlations highlighted potential links between repeat expressions and nearby genes, emphasising mostly 8-cell and morula specific genes together with SVA_D, LTR5_Hs and LTR70 transposons. The DNA methylation analysis further complemented the understanding on the mechanistic aspects of the repeatome’s regulation per se and revealed critical stages where DNA methylation levels are negatively correlating with repeat expression.ConclusionsTaken together, our study shows that specific expression patterns are not exclusive to genes and long non-coding RNAs but the repeatome also exhibits an intriguingly dynamic pattern at the global scale. Repeats identified in this study; particularly satellites, which were historically associated with heterochromatin, and those with potential links to nearby gene expression provide valuable insights into the understanding of key events in genomic regulation and warrant further research in epigenetics, genomics and developmental biology.