Single polysome analysis of mRNP.

Eukaryotic translation is a complex process that involves the interplay of various translation factors to convert genetic information into a specific amino acid chain. According to an elegant model of eukaryotic translation initiation, the 3' poly(A) tail of an mRNA, which is occupied by poly(A)-binding proteins (PABPs), communicates with the 5'-cap bound by eIF4E to enhance translation. Although the circularization of mRNA resulting from the communication is widely understood, it has yet to be directly observed. To explore mRNA circularization in translation, we analyzed the level of colocalization of eIF4E, eIF4G, and PABP on individual mRNAs in polysomal and subpolysomal fractions using single polysome analysis. Our results show that the three tested proteins barely coexist in mRNA in either polysomal or subpolysomal fractions, implying that the closed-loop structure generated by the communication between eIF4E, eIF4G, and PAPB may be transient during translation.

Prediction of inherited genomic susceptibility to 20 common cancer types by a supervised machine-learning method.

Prevention and early intervention are the most effective ways of avoiding or minimizing psychological, physical, and financial suffering from cancer. However, such proactive action requires the ability to predict the individual's susceptibility to cancer with a measure of probability. Of the triad of cancer-causing factors (inherited genomic susceptibility, environmental factors, and lifestyle factors), the inherited genomic component may be derivable from the recent public availability of a large body of whole-genome variation data. However, genome-wide association studies have so far showed limited success in predicting the inherited susceptibility to common cancers. We present here a multiple classification approach for predicting individuals' inherited genomic susceptibility to acquire the most likely phenotype among a panel of 20 major common cancer types plus 1 "healthy" type by application of a supervised machine-learning method under competing conditions among the cohorts of the 21 types. This approach suggests that, depending on the phenotypes of 5,919 individuals of "white" ethnic population in this study, (i) the portion of the cohort of a cancer type who acquired the observed type due to mostly inherited genomic susceptibility factors ranges from about 33 to 88% (or its corollary: the portion due to mostly environmental and lifestyle factors ranges from 12 to 67%), and (ii) on an individual level, the method also predicts individuals' inherited genomic susceptibility to acquire the other types ranked with associated probabilities. These probabilities may provide practical information for individuals, heath professionals, and health policymakers related to prevention and/or early intervention of cancer.

Seminal CD38 is a pivotal regulator for fetomaternal tolerance.

A successful pregnancy depends on a complex process that establishes fetomaternal tolerance. Seminal plasma is known to induce maternal immune tolerance to paternal alloantigens, but the seminal factors that regulate maternal immunity have yet to be characterized. Here, we show that a soluble form of CD38 (sCD38) released from seminal vesicles to the seminal plasma plays a crucial role in inducing tolerogenic dendritic cells and CD4(+) forkhead box P3(+) (Foxp3(+)) regulatory T cells (Tregs), thereby enhancing maternal immune tolerance and protecting the semiallogeneic fetus from resorption. The abortion rate in BALB/c females mated with C57BL/6 Cd38(-/-) males was high compared with that in females mated with Cd38(+/+) males, and this was associated with a reduced proportion of Tregs within the CD4(+) T-cell pool. Direct intravaginal injection of sCD38 to CBA/J pregnant mice at preimplantation increased Tregs and pregnancy rates in mice under abortive sonic stress from 48 h after mating until euthanasia. Thus, sCD38 released from seminal vesicles to the seminal plasma acts as an immunoregulatory factor to protect semiallogeneic fetuses from maternal immune responses.

The critical role of uterine CD31 as a post-progesterone signal in early pregnancy.

CD31 has been shown to play a role in endothelial cell migration and angiogenesis, which are critical to the formation and function of the endometrium and myometrium in uterine development during early pregnancy. However, the role of CD31 in uterine receptivity during blastocyst implantation is poorly understood. The pregnancy rate in CD31-/- female mice mated with CD31+/+ male mice was higher than that observed in CD31+/+ female mice mated with CD31+/+ male mice. During the receptive phase of implantation, uterine glands were more developed in CD31-/- mice than in CD31+/+ mice, and the uterine weights of CD31-/- mice were increased. Leukemia inhibitory factor (LIF) was highly expressed in the CD31-/- mice during implantation and the expression of LIF was up-regulated by estradiol-17ß (E2 ) + progesterone (P4 ) in ovariectomized CD31-/- mice, compared with CD31+/+ mice at 8 h after hormone treatment. E2 -induced protein synthesis was inhibited by P4 in the CD31+/+ uterus, but not in the uterus of CD31-/- mice. Also, STAT3, HAND2, LIF, and mTOR signals were enhanced in CD31-/- mice. Stromal DNA replication was highly activated in the uterus of CD31-/- mice, manifested by upregulated cyclin series signaling and PCNA expression after E2 + P4 treatment. Collectively, CD31 inhibits E2 -mediated epithelial proliferation via recruitment and phosphorylation of SHP-2 upon receiving P4 signal in early pregnancy.

Critical role for NAD glycohydrolase in regulation of erythropoiesis by hematopoietic stem cells through control of intracellular NAD content.

NAD glycohydrolases (NADases) catalyze the hydrolysis of NAD to ADP-ribose and nicotinamide. Although many members of the NADase family, including ADP-ribosyltransferases, have been cloned and characterized, the structure and function of NADases with pure hydrolytic activity remain to be elucidated. Here, we report the structural and functional characterization of a novel NADase from rabbit reticulocytes. The novel NADase is a glycosylated, glycosylphosphatidylinositol-anchored cell surface protein exclusively expressed in reticulocytes. shRNA-mediated knockdown of the NADase in bone marrow cells resulted in a reduction of erythroid colony formation and an increase in NAD level. Furthermore, treatment of bone marrow cells with NAD, nicotinamide, or nicotinamide riboside, which induce an increase in NAD content, resulted in a significant decrease in erythroid progenitors. These results indicate that the novel NADase may play a critical role in regulating erythropoiesis of hematopoietic stem cells by modulating intracellular NAD.

OCIAD2 activates γ-secretase to enhance amyloid ß production by interacting with nicastrin.

The gamma (γ)-secretase holoenzyme is composed of four core proteins and cleaves APP to generate amyloid beta (Aß), a key molecule that causes major neurotoxicity during the early stage of Alzheimer's disease (AD). However, despite its important role in Aß production, little is known about the regulation of γ-secretase. OCIAD2, a novel modulator of γ-secretase that stimulates Aß production, and which was isolated from a genome-wide functional screen using cell-based assays and a cDNA library comprising 6,178 genes. Ectopic expression of OCIAD2 enhanced Aß production, while reduction of OCIAD2 expression suppressed it. OCIAD2 expression facilitated the formation of an active γ-secretase complex and enhanced subcellular localization of the enzyme components to lipid rafts. OCIAD2 interacted with nicastrin to stimulate γ-secretase activity. OCIAD2 also increased the interaction of nicastrin with C99 and stimulated APP processing via γ-secretase activation, but did not affect Notch processing. In addition, a cell-permeable Tat-OCIAD2 peptide that interfered with the interaction of OCIAD2 with nicastrin interrupted the γ-secretase-mediated AICD production. Finally, OCIAD2 expression was significantly elevated in the brain of AD patients and PDAPP mice. This study identifies OCIAD2 as a selective activator of γ-secretase to increase Aß generation.

Autocrine/paracrine function of nicotinic acid adenine dinucleotide phosphate (NAADP) for glucose homeostasis in pancreatic ß-cells and adipocytes.

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger for mobilizing Ca(2+) from intracellular stores in various cell types. Extracellular application of NAADP has been shown to elicit intracellular Ca(2+) signals, indicating that it is readily transported into cells. However, little is known about the functional role of this NAADP uptake system. Here, we show that NAADP is effectively transported into selected cell types involved in glucose homeostasis, such as adipocytes and pancreatic ß-cells, but not the acinar cells, in a high glucose-dependent manner. NAADP uptake was inhibitable by Ned-19, a NAADP mimic; dipyridamole, a nucleoside inhibitor; or NaN3, a metabolic inhibitor or under Ca(2+)-free conditions. Furthermore, NAADP was found to be released from pancreatic islets upon stimulation by high glucose. Consistently, administration of NAADP to type 2 diabetic mice improved glucose tolerance. We propose that NAADP is functioning as an autocrine/paracrine hormone important in glucose homeostasis. NAADP is thus a potential antidiabetic agent with therapeutic relevance.

Whole-genome demography of COVID-19 virus during its pandemic period and on "panvalent" vaccine design.

With over 16 million submitted genomic sequences, the SARS-CoV-2 (SC2) virus, the cause of the most recent worldwide COVID-19 pandemic, has become the most sequenced genome of all known viruses, revealing, for example, a vast number of expanding viral lineages. Since the pandemic phase appears to be over, we performed a retrospective re-examination of the demographic grouping pattern and their genomic characteristics during the entire pandemic period up to the peak of the last pandemic wave. For our study, we extracted from the NCBI only unique viral sequences and converted each sequence data to a relational vector, indicating the presence/absence of each variational event compared to a "reference" sequence. Our study revealed several genomic features that are unexpected or different from those of previous studies. For example, approximately 44,000 variants with unique sequences emerged during the pandemic period; they group into only four major viral-genomic groups and each has a set of mostly unique highly-conserved variant-genotypes (HCVGs); and a small set from the first ("ancestral") group was inherited by the three ("descendant") groups, suggesting that HCVGs in the next group may be predictable from the current group(s). Such a concept may be potentially important in designing "panvalent" vaccines against the current and future waves of viral infections.

On whole-genome demography of world's ethnic groups and individual genomic identity.

All current categorizations of human population, such as ethnicity, ancestry and race, are based on various selections and combinations of complex and dynamic common characteristics, that are mostly societal and cultural in nature, perceived by the members within or from outside of the categorized group. During the last decade, a massive amount of a new type of characteristics, that are exclusively genomic in nature, became available that allows us to analyze the inherited whole-genome demographics of extant human, especially in the fields such as human genetics, health sciences and medical practices (e.g., 1,2,3), where such health-related characteristics can be related to whole-genome-based categorization. Here we show the feasibility of deriving such whole-genome-based categorization. We observe that, within the available genomic data at present, (a) the study populations form about 14 genomic groups, each consisting of multiple ethnic groups; and (b), at an individual level, approximately 99.8%, on average, of the whole autosomal-genome contents are identical between any two individuals regardless of their genomic or ethnic groups.

Single-molecule visualization of mRNA circularization during translation.

Translation is mediated by precisely orchestrated sequential interactions among translation initiation components, mRNA, and ribosomes. Biochemical, structural, and genetic techniques have revealed the fundamental mechanism that determines what occurs and when, where and in what order. Most mRNAs are circularized via the eIF4E-eIF4G-PABP interaction, which stabilizes mRNAs and enhances translation by recycling ribosomes. However, studies using single-molecule fluorescence imaging have allowed for the visualization of complex data that opposes the traditional "functional circularization" theory. Here, we briefly introduce single-molecule techniques applied to studies on mRNA circularization and describe the results of in vitro and live-cell imaging. Finally, we discuss relevant insights and questions gained from single-molecule research related to translation.

Monitoring Immune Modulation in Season Population: Identifying Effects and Markers Related to Apis mellifera ligustica Honey Bee Health.

Honey bees play a significant role in ecology, producing biologically active substances used to promote human health. However, unlike humans, the molecular markers indicating honey bee health remain unknown. Unfortunately, numerous reports of honey bee collapse have been documented. To identify health markers, we analyzed ten defense system genes in Apis mellifera ligustica honey bees from winter (Owb) and spring (Fb for foragers and Nb for newly emerged) populations sampled in February and late April 2023, respectively. We focused on colonies free from SBV and DWV viruses. Molecular profiling revealed five molecular markers of honey bee health. Of these, two seasonal molecular markers-domeless and spz genes-were significantly downregulated in Owb compared to Nb and Fb honey bees. One task-related marker gene, apid-1, was identified as being downregulated in Owb and Nb compared to Fb honey bees. Two recommended general health markers, SOD and defensin-2, were upregulated in honey bees. These markers require further testing across various honey bee subspecies in different climatic regions. They can diagnose bee health without colony intervention, especially during low-temperature months like winter. Beekeepers can use this information to make timely adjustments to nutrients or heating to prevent seasonal losses.

Association between obesity and the prevalence of allergic diseases, atopy, and bronchial hyperresponsiveness in Korean adolescents.

BACKGROUND: Although several mechanisms underlying the asthma-obesity connection have been proposed, debates still remain. This study was to determine whether overweight is associated with a higher prevalence of atopy, asthma symptoms, airway obstruction, bronchial hyperresponsiveness (BHR) or biomarkers of inflammation in a sample of Korean adolescents. METHODS: We conducted a cross-sectional survey involving questionnaires, skin tests, spirometry and methacholine challenge tests among 717 adolescents from Seoul (South Korea). Overweight status was defined as a BMI greater than the local age- and gender-specific 85th percentile. RESULTS: Overweight subjects more frequently reported ever having wheezing (24.6 vs. 14.0%, p = 0.001) and wheezing in the previous 12 months (11.5 vs. 6.3%, p = 0.02) than normal-weight subjects, especially in boys. Atopy was more common among overweight adolescents than among those of normal weight (61.5 vs. 49.2%, p = 0.002), especially in boys (65.0 vs. 52.8%, p = 0.005). Overweight subjects had higher total WBC counts and eosinophil counts, especially boys. The presence of BHR was more common only among overweight girls (32.8 vs. 18.0%, p = 0.028). Overweight status was a significant risk factor for the presence of atopy (odds ratio = 1.49; 95% CI 1.06-2.10), after adjusting for various confounders by logistic regression analysis. CONCLUSIONS: An association was found between overweight status and both atopy and an increased prevalence of wheezing in adolescent Korean boys. These findings suggest that being overweight in puberty may be one of several risk factors responsible for atopy, BHR, and asthma symptoms.

TRIM28 functions as a negative regulator of aggresome formation.

Selective recognition and elimination of misfolded polypeptides are crucial for protein homeostasis. When the ubiquitin-proteasome system is impaired, misfolded polypeptides tend to form small cytosolic aggregates and are transported to the aggresome and eventually eliminated by the autophagy pathway. Despite the importance of this process, the regulation of aggresome formation remains poorly understood. Here, we identify TRIM28/TIF1ß/KAP1 (tripartite motif containing 28) as a negative regulator of aggresome formation. Direct interaction between TRIM28 and CTIF (cap binding complex dependent translation initiation factor) leads to inefficient aggresomal targeting of misfolded polypeptides. We also find that either treatment of cells with poly I:C or infection of the cells by influenza A viruses triggers the phosphorylation of TRIM28 at S473 in a way that depends on double-stranded RNA-activated protein kinase. The phosphorylation promotes association of TRIM28 with CTIF, inhibits aggresome formation, and consequently suppresses viral proliferation. Collectively, our data provide compelling evidence that TRIM28 is a negative regulator of aggresome formation.Abbreviations: BAG3: BCL2-associated athanogene 3; CTIF: CBC-dependent translation initiation factor; CED: CTIF-EEF1A1-DCTN1; DCTN1: dynactin subunit 1; EEF1A1: eukaryotic translation elongation factor 1 alpha 1; EIF2AK2: eukaryotic translation initiation factor 2 alpha kinase 2; HDAC6: histone deacetylase 6; IAV: influenza A virus; IP: immunoprecipitation; PLA: proximity ligation assay; polypeptidyl-puro: polypeptidyl-puromycin; qRT-PCR: quantitative reverse-transcription PCR; siRNA: small interfering RNA.

Nonsense-mediated mRNA decay factor UPF1 promotes aggresome formation.

Nonsense-mediated mRNA decay (NMD) typifies an mRNA surveillance pathway. Because NMD necessitates a translation event to recognize a premature termination codon on mRNAs, truncated misfolded polypeptides (NMD-polypeptides) could potentially be generated from NMD substrates as byproducts. Here, we show that when the ubiquitin-proteasome system is overwhelmed, various misfolded polypeptides including NMD-polypeptides accumulate in the aggresome: a perinuclear nonmembranous compartment eventually cleared by autophagy. Hyperphosphorylation of the key NMD factor UPF1 is required for selective targeting of the misfolded polypeptide aggregates toward the aggresome via the CTIF-eEF1A1-DCTN1 complex: the aggresome-targeting cellular machinery. Visualization at a single-particle level reveals that UPF1 increases the frequency and fidelity of movement of CTIF aggregates toward the aggresome. Furthermore, the apoptosis induced by proteotoxic stresses is suppressed by UPF1 hyperphosphorylation. Altogether, our data provide evidence that UPF1 functions in the regulation of a protein surveillance as well as an mRNA quality control.

Trpm2 Ablation Accelerates Protein Aggregation by Impaired ADPR and Autophagic Clearance in the Brain.

TRPM2 a cation channel is also known to work as an enzyme that hydrolyzes highly reactive, neurotoxic ADP-ribose (ADPR). Although ADPR is hydrolyzed by NUT9 pyrophosphatase in major organs, the enzyme is defective in the brain. The present study questions the role of TRPM2 in the catabolism of ADPR in the brain. Genetic ablation of Trpm2 results in the disruption of ADPR catabolism that leads to the accumulation of ADPR and reduction in AMP. Trpm2-/- mice elicit the reduction in autophagosome formation in the hippocampus. Trpm2-/- mice also show aggregations of proteins in the hippocampus, aberrant structural changes and neuronal connections in synapses, and neuronal degeneration. Trpm2-/- mice exhibit learning and memory impairment, enhanced neuronal intrinsic excitability, and imbalanced synaptic transmission. These results respond to long-unanswered questions regarding the potential role of the enzymatic function of TRPM2 in the brain, whose dysfunction evokes protein aggregation. In addition, the present finding answers to the conflicting reports such as neuroprotective or neurodegenerative phenotypes observed in Trpm2-/- mice.

Chinese medicines reported to have effects on contact dermatitis in the last 20 years.

Contact dermatitis (CD) is one of the most common skin diseases in industrialized countries. Chinese medicines (CMs) have been investigated worldwide as complementary and alternative medicines for corticosteroids, which are the first choice for treatment of inflflammatory skin diseases owing to their favorable efficacy. This article describes the CMs that have been reported to have anti-dermatitis effects against CD in the last 20 years.

Exercise induces muscle fiber type switching via transient receptor potential melastatin 2-dependent Ca2+ signaling.

The aim of the present study was to examine whether transient receptor potential melastatin 2 (TRPM2) plays a role in muscle fiber-type transition during exercise. Mice were trained at a speed of 12 m/min at a slope of 0° for 60 min for 5 consecutive days/wk for 4 wk. Exhaustion tests were performed on the treadmill (the speed was set at 6 m/min at a slope of 0° and increased at a rate of 1 m/min every 6 min). Isolated primary skeletal muscle cells from TRPM2-knockout (KO) mice showed lower amplitudes of electrical stimuli (ES)-induced Ca2+ signals when compared with wild-type (WT) mice due to a defect in Ca2+ influx. Moreover, TRPM2-KO mice had a higher proportion of fast-twitch skeletal muscle fibers and a lower proportion of slow-twitch muscle fibers before exercise than WT mice. After exercise, the expression of slow-twitch skeletal muscle fibers was increased only in WT mice but not in TRPM2-KO mice. ES-induced nuclear translocation of the Ca2+-dependent transcription factor NFATc1 was significantly lower in TRPM2-KO mice than in WT mice. TRPM2-KO mice also showed decreased mitochondrial Ca2+ and membrane potential. Lactate levels were higher in the skeletal muscle cells of TRPM2-KO mice before and after ES compared with WT mice. Collectively, these data indicate that TRPM2-mediated Ca2+ signaling plays a critical role in the regulation of fiber-type switching and mitochondrial function in skeletal muscle. NEW & NOTEWORTHY TRPM2 has been shown to play an important role in a variety of cellular functions. However, the role of TRPM2 in skeletal muscle remains poorly understood. Here, we provide evidence that TRPM2-mediated Ca2+ signaling is required for training-induced improvement in skeletal muscle mitochondrial function and fiber type transition.

High levels of Cdc7 and Dbf4 proteins can arrest cell-cycle progression.

Cdc7-Dbf4 serine/threonine kinase is essential for initiation of DNA replication. It was previously found that overexpression of certain replication proteins such as Cdc6 and Cdt1 in fission yeast resulted in multiple rounds of DNA replication in the absence of mitosis. Since this phenomenon is dependent upon the presence of wild-type Cdc7/Hsk1, we hypothesized that high levels of Cdc7 and/or Dbf4 could also cause multiple rounds of DNA replication, or could facilitate entry into S phase. To test this hypothesis, we transiently overexpressed hamster Cdc7, Dbf4 or both in CHO cells. Direct observations of individual cells by fluorescence microscopy and flow cytometric analysis on cell populations suggest that overexpression of Cdc7 and/or Dbf4 does not result in multiple rounds of DNA replication or facilitating entry into S phase. In contrast, moderately increased levels of Dbf4, but not Cdc7, cause cell-cycle arrest in G2/M. This G2/M arrest coincides with hyperphosphorylation of Cdc2/Cdk1 at Tyr-15, raising the possibility that high levels of Dbf4 may activate a G2/M cell-cycle checkpoint. Further increase in Cdc7 and/or Dbf4 by 2-4 fold can arrest cells in G1 and significantly slow down S-phase progression for the cells already in S phase.

Overexpression of calsenilin enhances gamma-secretase activity.

Presenilin/gamma-secretase is a membrane-associated protease that cleaves within the transmembrane region of the amyloid precursor protein (APP) to generate amyloid-beta peptide (Abeta) whose deposition in the brain is a characteristic of Alzheimer's disease (AD). Calsenilin, a calcium binding protein that has been shown to interact with the C-termini of both presenilin 1 (PS1) and presenilin 2 (PS2), appears to play a role in transcriptional regulation and apoptosis and to bind to A-type voltage-gated potassium channels. Here, we report that overexpression of calsenilin enhanced gamma-secretase activity in cells. The effect of calsenilin on the gamma-cleavage of substrates was blocked by the selective gamma-secretase inhibitor L-685,458. We also employed a cellular gamma-cleavage GFP-reporter assay to demonstrate the effect of calsenilin on gamma-secretase activity. To establish a direct role for calsenilin in regulating gamma-secretase activity, we incubated purified calsenilin with isolated membrane fractions and found increased Abeta production in a cell free system. These data suggest that calsenilin may be one of the regulatory factors for gamma-secretase.

Seminal CD38 Enhances Human Sperm Capacitation through Its Interaction with CD31.

Human sperm have to undergo a maturational process called capacitation in the female reproductive tract. Capacitation confers upon the sperm an ability to gain hypermotility and undergo acrosome reaction. Previous studies have suggested that seminal plasma proteins induce the capacitation of sperm in the female reproductive tract for the successful fertilization of the oocyte. However, the function of seminal plasma proteins in capacitation remains largely unclear. To the end, we found that soluble CD38 (sCD38) in seminal plasma increases the capacitation of sperm via specific interactions between sCD38 and the CD31 on the sperm. Upon the association of sCD38 with CD31, tyrosine kinase Src phosphorylates CD31, a process blocked by Src inhibitors. Shc, SHP-2, Grb2, and SOS, as well as Src kinase were found to associate with the phosphorylated CD31. The sCD38-induced phosphorylation of CD31 initiates a cascade reaction through the phosphorylation of Erk1/2, which results in the acrosome reaction, and sperm hypermotility. These processes were prevented by Src, Ras and MEK inhibitors. Taken together, these data indicate that the sCD38 present in seminal plasma plays a critical role in the capacitation of sperm.