Dzip1 is dynamically expressed in the vertebrate germline and regulates the development of Xenopus primordial germ cells.

Primordial germ cells (PGCs) are the precursors of sperms and oocytes. Proper development of PGCs is crucial for the survival of the species. In many organisms, factors responsible for PGC development are synthesized during early oogenesis and assembled into the germ plasm. During early embryonic development, germ plasm is inherited by a few cells, leading to the formation of PGCs. While germline development has been extensively studied, how components of the germ plasm regulate PGC development is not fully understood. Here, we report that Dzip1 is dynamically expressed in vertebrate germline and is a novel component of the germ plasm in Xenopus and zebrafish. Knockdown of Dzip1 impairs PGC development in Xenopus embryos. At the molecular level, Dzip1 physically interacts with Dazl, an evolutionarily conserved RNA-binding protein that plays a multifaced role during germline development. We further showed that the sequence between amino acid residues 282 and 550 of Dzip1 is responsible for binding to Dazl. Disruption of the binding between Dzip1 and Dazl leads to defective PGC development. Taken together, our results presented here demonstrate that Dzip1 is dynamically expressed in the vertebrate germline and plays a novel function during Xenopus PGC development.

PTBP1 controls intestinal epithelial regeneration through post-transcriptional regulation of gene expression.

The intestinal epithelial regeneration is driven by intestinal stem cells under homeostatic conditions. Differentiated intestinal epithelial cells, such as Paneth cells, are capable of acquiring multipotency and contributing to regeneration upon the loss of intestinal stem cells. Paneth cells also support intestinal stem cell survival and regeneration. We report here that depletion of an RNA-binding protein named polypyrimidine tract binding protein 1 (PTBP1) in mouse intestinal epithelial cells causes intestinal stem cell death and epithelial regeneration failure. Mechanistically, we show that PTBP1 inhibits neuronal-like splicing programs in intestinal crypt cells, which is critical for maintaining intestinal stem cell stemness. This function is achieved at least in part through promoting the non-productive splicing of its paralog PTBP2. Moreover, PTBP1 inhibits the expression of an AKT inhibitor PHLDA3 in Paneth cells and permits AKT activation, which presumably maintains Paneth cell plasticity and function in supporting intestinal stem cell niche. We show that PTBP1 directly binds to a CU-rich region in the 3' UTR of Phlda3, which we demonstrate to be critical for downregulating the mRNA and protein levels of Phlda3. Our results thus reveal the multifaceted in vivo regulation of intestinal epithelial regeneration by PTBP1 at the post-transcriptional level.

Protective Effect of Red Light-Emitting Diode against UV-B Radiation-Induced Skin Damage in SKH:HR-2 Hairless Mice.

In this in vivo study on hairless mice, we examined the effects of light-emitting diode (LED) treatment applied prior to ultraviolet B (UVB) irradiation. We found that pre-treating with LED improved skin morphological and histopathological conditions compared to those only exposed to UVB irradiation. In our study, histological evaluation of collagen and elastic fibers after LED treatment prior to UVB irradiation showed that this pretreatment significantly enhanced the quality of fibers, which were otherwise poor in density and irregularly arranged due to UV exposure alone. This suggests that LED treatment promotes collagen and elastin production, leading to improved skin properties. Additionally, we observed an increase in Claudin-1 expression and a reduction in nuclear factor-erythroid 2-related factor 2 (Nrf-2) and heme-oxygenase 1 (HO-1) expression within the LED-treated skin tissues, suggesting that LED therapy may modulate key skin barrier proteins and oxidative stress markers. These results demonstrate that pretreatment with LED light can enhance the skin's resistance to UVB-induced damage by modulating gene regulation associated with skin protection. Further investigations are needed to explore the broader biological effects of LED therapy on other tissues such as blood vessels. This study underscores the potential of LED therapy as a non-invasive approach to enhance skin repair and counteract the effects of photoaging caused by UV exposure.

Precise modulation of embryonic development through optogenetics.

The past decade has witnessed enormous progress in optogenetics, which uses photo-sensitive proteins to control signal transduction in live cells and animals. The ever-increasing amount of optogenetic tools, however, could overwhelm the selection of appropriate optogenetic strategies. In this work, we summarize recent progress in this emerging field and highlight the application of opsin-free optogenetics in studying embryonic development, focusing on new insights gained into optical induction of morphogenesis, cell polarity, cell fate determination, tissue differentiation, neuronal regeneration, synaptic plasticity, and removal of cells during development.

Novel functions of the ubiquitin-independent proteasome system in regulating Xenopus germline development.

In most species, early germline development occurs in the absence of transcription with germline determinants subject to complex translational and post-translational regulations. Here, we report for the first time that early germline development is influenced by dynamic regulation of the proteasome system, previously thought to be ubiquitously expressed and to serve 'housekeeping' roles in controlling protein homeostasis. We show that proteasomes are present in a gradient with the highest levels in the animal hemisphere and extending into the vegetal hemisphere of Xenopus oocytes. This distribution changes dramatically during the oocyte-to-embryo transition, with proteasomes becoming enriched in and restricted to the animal hemisphere and therefore separated from vegetally localized germline determinants. We identify Dead-end1 (Dnd1), a master regulator of vertebrate germline development, as a novel substrate of the ubiquitin-independent proteasomes. In the oocyte, ubiquitin-independent proteasomal degradation acts together with translational repression to prevent premature accumulation of Dnd1 protein. In the embryo, artificially increasing ubiquitin-independent proteasomal degradation in the vegetal pole interferes with germline development. Our work thus reveals novel inhibitory functions and spatial regulation of the ubiquitin-independent proteasome during vertebrate germline development.

Combined Toxicological Effects of Di (2-Ethylhexyl) Phthalate and UV-B Irradiation through Endoplasmic Reticulum Stress-Tight Junction Disruption in Human HaCaT Keratinocytes.

Di (2-ethylhexyl) phthalate (DEHP) is widely used as a plasticizer, and human exposure to DEHP is widespread and frequent. However, information about the combined effect of DEHP and ultraviolet (UV)-B on the skin are still limited. We investigated the cytotoxic effects of DEHP and UV-B on HaCaT keratinocytes and evaluated the related underlying mechanisms involving endoplasmic reticulum (ER) stress signals and the disruption of junction complexes as an effective target for skin inflammation. Our results revealed that co-treatment with DEHP and UV-B irradiation alleviated the cell cytotoxicity and markedly decreased X-box binding protein 1 (XBP1), endoplasmic reticulum oxidoreductase 1 alpha (Ero1α), and C/EBP homologous protein (CHOP) whereas a single dose of 40 mJ/cm2 UV-B generated mild ER stress to slightly less or similar levels as that seen with DEHP. DEHP was also shown to inhibit tight junctions (TJs) after UV-B irradiation, increased apoptosis by altering apoptotic gene Bax and stress kinases, JNK, and p38 MAPK. Furthermore, exposure of HaCaT cells to DEHP and UV-B irradiation resulted in the marked suppression of the nuclear factor kappa B (NF-κB)/p65 signaling pathway. Taken together, our data suggest that nontoxic DEHP and UV-B irradiation regulated ER stress and epidermal TJ disruption with the induction of apoptosis activation and the secretion of proinflammatory cytokines such as interleukin 1 beta (IL-1ß) and IL-6 in human keratinocytes. Further investigation is needed to confirm the mechanisms implicated in its toxicity and determine the effects of exposure to DEHP and UV-B irradiation on markers involved in this study.

Structural basis of the specific interaction of SMRT corepressor with histone deacetylase 4.

Modification of chromatin and related transcription factors by histone deacetylases (HDACs) is one of the major strategies for controlling gene expression in eukaryotes. The HDAC domains of class IIa HDACs repress the respective target genes by interacting with the C-terminal region of the silencing mediator for retinoid and thyroid receptor (SMRT) repression domain 3 (SRD3c). However, latent catalytic activity suggests that their roles as deacetylases in gene regulation are unclear. Here, we found that two conserved GSI-containing motifs of SRD3c are critical for HDAC4 binding. Two SMRT peptides including these motifs commonly form a ß-hairpin structure in the cleft and block the catalytic entry site of HDAC4. They interact mainly with class IIa HDAC-specific residues of HDAC4 in a closed conformation. Structure-guided mutagenesis confirmed critical interactions between the SMRT peptides and HDAC4 and -5 as well as the contribution of the Arg1369 residue in the first motif for optimal binding to the two HDACs. These results indicate that SMRT binding does not activate the cryptic deacetylase activity of HDAC4 and explain how class IIa HDACs and the SMRT-HDAC3 complex are coordinated during gene regulation.

Single Cell Proteomics by Data-Independent Acquisition To Study Embryonic Asymmetry in Xenopus laevis.

Techniques that allow single cell analysis are gaining widespread attention, and most of these studies utilize genomics-based approaches. While nanofluidic technologies have enabled mass spectrometric analysis of single cells, these measurements have been limited to metabolomics and lipidomic studies. Single cell proteomics has the potential to improve our understanding of intercellular heterogeneity. However, this approach has faced challenges including limited sample availability, as well as a requirement of highly sensitive methods for sample collection, cleanup, and detection. We present a technique to overcome these limitations by combining a micropipette (pulled glass capillary) based sample collection strategy with offline sample preparation and nanoLC-MS/MS to analyze proteins through a bottom-up proteomic strategy. This study explores two types of proteomics data acquisition strategies namely data-dependent (DDA) and data-independent acquisition (DIA). Results from the study indicate DIA to be more sensitive enabling analysis of >1600 proteins from ∼130 µm Xenopus laevis embryonic cells containing <6 nL of cytoplasm. The method was found to be robust in obtaining reproducible protein quantifications from single cells spanning the 1-128-cell stages of development. Furthermore, we used micropipette sampling to study intercellular heterogeneity within cells in a single embryo and investigated embryonic asymmetry along both animal-vegetal and dorsal-ventral axes during early stages of development. Investigation of the animal-vegetal axis led to discovery of various asymmetrically distributed proteins along the animal-vegetal axis. We have further compared the hits found from our proteomic data sets with other studies and validated a few hits using an orthogonal imaging technique. This study forms the first report of vegetal enrichment of the germ plasm associated protein DDX4/VASA in Xenopus embyos. Overall, the method and data presented here holds promise to enable important leads in developmental biology.

Solution structure of the porcine sapovirus VPg core reveals a stable three-helical bundle with a conserved surface patch.

Viral protein genome-linked (VPg) proteins play a critical role in the life cycle of vertebrate and plant positive-sense RNA viruses by acting as a protein primer for genome replication and as a protein cap for translation initiation. Here we report the solution structure of the porcine sapovirus VPg core (VPg(C)) determined by multi-dimensional NMR spectroscopy. The structure of VPg(C) is composed of three α-helices stabilized by several conserved hydrophobic residues that form a helical bundle core similar to that of feline calicivirus VPg. The putative nucleotide acceptor Tyr956 within the first helix of the core is completely exposed to solvent accessible surface to facilitate nucleotidylation by viral RNA polymerase. Comparison of VPg structures suggests that the surface for nucleotidylation site is highly conserved among the Caliciviridae family, whereas the backbone core structures are different. These structural features suggest that caliciviruses share common mechanisms of VPg-dependent viral replication and translation.

Crystal structure of cbbF from Zymomonas mobilis and its functional implication.

A phosphate group at the C1-atom of inositol-monophosphate (IMP) and fructose-1,6-bisphosphate (FBP) is hydrolyzed by a phosphatase IMPase and FBPase in a metal-dependent way, respectively. The two enzymes are almost indiscernible from each other because of their highly similar sequences and structures. Metal ions are bound to residues on the ß1- and ß2-strands and one mobile loop. However, FBP has another phosphate and FBPases exist as a higher oligomeric state, which may discriminate FBPases from IMPases. There are three genes annotated as FBPases in Zymomonas mobilis, termed also cbbF (ZmcbbF). The revealed crystal structure of one ZmcbbF shows a globular structure formed by five stacked layers. Twenty-five residues in the middle of the sequence form an α-helix and a ß-strand, which occupy one side of the catalytic site. A non-polar Leu residue among them is protruded to the active site, pointing out unfavorable access of a bulky charged group to this side. In vitro assays have shown its dimeric form in solution. Interestingly, two ß-strands of ß1 and ß2 are disordered in the ZmcbbF structure. These data indicate that ZmcbbF might structurally belong to IMPase, and imply that its active site would be reorganized in a yet unreported way.

A fractionation-based protocol to investigate RNA solubility phase transition during Xenopus oocyte maturation.

While RNAs are soluble in vitro, their solubility may be altered when incorporated into some protein complexes inside the cell. The solubility phase transition of RNAs is thus indicative of changes in the function and activity of RNAs. Here, we present a protocol for the assessment of RNA solubility phase transition during Xenopus oocyte maturation. We describe steps for sample preparation, cell fractionation, RNA extraction, real-time PCR, and analysis of the obtained results. For complete details on the use and execution of this protocol, please refer to Hwang et al. (2023).1.

Investigating the Nutritional and Functional Properties of Protaetia brevitarsis Larvae and Isolated Soy Protein Mixtures as Alternative Protein Sources.

The growing demand for sustainable and alternative protein sources has spurred interest in insect-based and plant-based proteins. Protaetia brevitarsis (PB) larvae and isolated soy protein (ISP) are notable in this regard, offering potential health benefits and nutritional enhancements. We assessed the feasibility of PB larvae and ISP mixtures as alternative food ingredients. Methods included the optimized purification and freeze-drying of PB larvae, extraction and refinement of legume proteins, physicochemical and antioxidant capacity evaluations, DPPH radical scavenging activity measurement, total phenolic and flavonoids content quantification, general component analysis, amino acid profiling using HPLC, fatty acid profiling through gas chromatography, and mineral content analysis using inductively coupled plasma spectrometry. The study found that certain PB:ISP ratios, particularly a 7:3 ratio, significantly improved the blend's antioxidant capacity, as evidenced by DPPH scavenging activity. This ratio also impacted the nutritional profile by altering the mixture's general components, with a notable increase in moisture, crude protein, and fiber and a decrease in crude fat and ash. Amino acid analysis revealed a balanced presence of essential and non-essential amino acids. The fatty acid profile was rich in unsaturated fatty acids, especially in certain ratios. Mineral analysis showed a complex interplay between PB larvae and ISP, with some minerals decreasing and others increasing in the blend. PB larvae and ISP mixtures have significant potential as alternative protein sources, offering a diversified nutritional profile and enhanced antioxidant properties. The 7:3 ratio of PB larvae to ISP has been shown to be particularly effective, suggesting that this ratio may offer an optimal balance for enhancing the overall nutritional quality of the mixture. This study sets the stage for future research to further explore and optimize the potential of these mixtures for human consumption while considering the challenges of consumer acceptance and long-term safety.

Dzip1 is dynamically expressed in the vertebrate germline and regulates the development of Xenopus primordial germ cells.

Primordial germ cells (PGCs) are the precursors of sperms and oocytes. Proper development of PGCs is crucial for the survival of the species. In many organisms, factors responsible for PGC development are synthesized during early oogenesis and assembled into the germ plasm. During early embryonic development, germ plasm is inherited by a few cells, leading to the formation of PGCs. While germline development has been extensively studied, how components of the germ plasm regulate PGC development is not fully understood. Here, we report that Dzip1 is dynamically expressed in vertebrate germline and is a novel component of the germ plasm in Xenopus and zebrafish. Knockdown of Dzip1 impairs PGC development in Xenopus embryos. At the molecular level, Dzip1 physically interacts with Dazl, an evolutionarily conserved RNA-binding protein that plays a multifaced role during germline development. We further showed that the sequence between amino acid residues 282 and 550 of Dzip1 is responsible for binding to Dazl. Disruption of the binding between Dzip1 and Dazl leads to defective PGC development. Taken together, our results presented here demonstrate that Dzip1 is dynamically expressed in the vertebrate germline and plays a novel function during Xenopus PGC development.

Solubility phase transition of maternal RNAs during vertebrate oocyte-to-embryo transition.

The oocyte-to-embryo transition (OET) is regulated by maternal products stored in the oocyte cytoplasm, independent of transcription. How maternal products are precisely remodeled to dictate the OET remains largely unclear. In this work, we discover the dynamic solubility phase transition of maternal RNAs during Xenopus OET. We have identified 863 maternal transcripts that transition from a soluble state to a detergent-insoluble one after oocyte maturation. These RNAs are enriched in the animal hemisphere, and many of them encode key cell cycle regulators. In contrast, 165 transcripts, including nearly all Xenopus germline RNAs and some vegetally localized somatic RNAs, undergo an insoluble-to-soluble phase transition. This phenomenon is conserved in zebrafish. Our results demonstrate that the phase transition of germline RNAs influences their susceptibility to RNA degradation machinery and is mediated by the remodeling of germ plasm. This work thus identifies important remodeling mechanisms that act on RNAs to control vertebrate OET.

Phase transition of maternal RNAs during vertebrate oocyte-to-embryo transition.

The oocyte-to-embryo transition (OET) is regulated by maternal products stored in the oocyte cytoplasm, independent of transcription. How maternal products are precisely remodeled to dictate the OET remains an open question. In this work, we discover the dynamic phase transition of maternal RNAs during Xenopus OET. We have identified 863 maternal transcripts that transition from a soluble state to a detergent-insoluble one after oocyte maturation. These RNAs are enriched in the animal hemisphere and many of them encode key cell cycle regulators. In contrast, 165 transcripts, including nearly all Xenopus germline RNAs and some vegetally localized somatic RNAs, undergo an insoluble-to-soluble phase transition. This phenomenon is conserved in zebrafish. Our results demonstrate that the phase transition of germline RNAs influences their susceptibility to RNA degradation machinery and is mediated by the remodeling of germ plasm. This work thus uncovers novel remodeling mechanisms that act on RNAs to regulate vertebrate OET.

Structural characterization of a modification subunit of a putative type I restriction enzyme from Vibrio vulnificus YJ016.

In multifunctional type I restriction enzymes, active methyltransferases (MTases) are constituted of methylation (HsdM) and specificity (HsdS) subunits. In this study, the crystal structure of a putative HsdM subunit from Vibrio vulnificus YJ016 (vvHsdM) was elucidated at a resolution of 1.80 Å. A cofactor-binding site for S-adenosyl-L-methionine (SAM, a methyl-group donor) is formed within the C-terminal domain of an α/ß-fold, in which a number of residues are conserved, including the GxGG and (N/D)PP(F/Y) motifs, which are likely to interact with several functional moieties of the SAM methyl-group donor. Comparison with the N6 DNA MTase of Thermus aquaticus and other HsdM structures suggests that two aromatic rings (Phe199 and Phe312) in the motifs that are conserved among the HsdMs may sandwich both sides of the adenine ring of the recognition sequence so that a conserved Asn residue (Asn309) can interact with the N6 atom of the target adenine base (a methyl-group acceptor) and locate the target adenine base close to the transferred SAM methyl group.

Correlation of attention deficit hyperactivity disorder with gut microbiota according to the dietary intake of Korean elementary school students.

We investigated the impact of dietary patterns on the gut microbiota and concentration of short-chain fatty acids in the feces of Korean elementary school students. The dietary intake and ADHD assessment of 40 Korean elementary school students were analyzed using a dish-based semi-quantitative food frequency questionnaire. Analysis of gut microbiota and short-chain fatty acids composition were performed using the real-time polymerase chain reaction, metagenomics, and gas chromatography methods. The dietary patterns of participants were divided into four groups: healthy, processed food, fish and shellfish, and meat. The participants were also divided into two groups according to their ADHD scores: 0-30, control group; over 30, ADHD group. The ADHD score of the processed food group was significantly higher than that of the healthy group. The processed food and ADHD groups showed significantly higher abundance of harmful bacteria, such as the Enterobacter, Escherichia coli, and Clostridium strains, and markedly lower abundance of beneficial bacteria, such as the Bifidobacterium and Ruminococcus strains, than the control group. The heat maps of metagenomics indicated that each group was separated into distinct clusters, and the processed food and ADHD groups showed significantly lower α-diversity of gut microbiota than the control group. In these groups, the concentration of acetate or butyrate in the feces was significantly lower than that in the control group. These results may indicate that imbalanced diets can disturb the colonic microbial balance and are likely to become a potential risk factor for the prevalence of ADHD.

The Association between Self-Rated Health Status, Psychosocial Stress, Eating Behaviors, and Food Intake According to the Level of Sunlight Exposure in Korean Adults.

Sunlight exposure has been reported to have various beneficial effects on human health. This study investigated the relationship between self-rated health status, psychosocial stress, eating behaviors, and food intake according to sunlight exposure in 948 adults. Sunlight exposure was classified as less than one hour, less than three hours, and greater than three hours. Of the participants, 49.2% had fewer than three hours of daily exposure to sunlight. Regarding participants exposed to sunlight for less than one hour, the largest response was that they did not engage in outdoor activities on weekdays or weekends, and the rate of being outdoors in the shade on sunny days was the highest in this group at 42.7%. Furthermore, the participants exposed to sunlight for less than one hour had a lower health response than the other two groups, and there were significantly more participants classified in the stress risk group. Regarding eating habits, those with less than an hour of exposure to sunlight frequently ate fried foods, fatty foods, added salt, and snacks, and had significantly lower total dietary scores or three regular meals. Additionally, their frequency of consumption of cereals, milk and dairy products, orange juice, and pork was also significantly lower than the other groups. Thus, it is necessary to provide sufficient guidelines for adequate sunlight exposure and food intake because participants with low sunlight exposure may have low vitamin D synthesis and insufficient food intake.

Butyrate modulates mucin secretion and bacterial adherence in LoVo cells via MAPK signaling.

Short-chain fatty acids contribute to normal bowel function and prevent bacterial infections. In particular, butyrate is a promising candidate that plays an important role in regulating the functional integrity of the gastrointestinal tract by stimulating mucin secretion. We investigated whether butyrate treatment modulates mucin secretion and bacterial adherence in LoVo cells. In addition, the possible signaling pathways were also examined in connection with the upregulation of mucin secretion. The results showed that butyrate induced mucin secretion in LoVo cells, resulting in the inhibition of Escherichia coli adhesion by increasing the adherence of Lactobacillus acidophilus and Bifidobacterium longum. The gene expression analysis suggests that mitogen-activated protein kinase (MAPK) signaling pathways including Cdc42-PAK pathway appears to be involved in stimulating mucin secretion. More importantly, butyrate induced the increased actin expression and polymerization in LoVo cells, which could be attributable to the Cdc42-PAK signaling pathway, implicated in actin cytoskeleton and mucin secretion. Our results provide a molecular basis in modulating bacterial adherence and the MAPK signaling pathway for the improved homeostasis of colonic epithelial cells.

Quality Characteristics of Substitute Meat Patties Developed Using Aruncus dioicus var. kamtschaticus Hara.

We developed a vegetable alternative to meat patties using Aruncus dioicus var. kamtschaticus Hara (A. dioicus) and used it to generate basic data for the alternative meat market by comparing nutritional and microbiological components with commercially available vegetable and meat patties. Nutrient analysis, microbiological analysis, chromaticity, and texture analysis were performed on substitute meat patties (SMPs) with A. dioicus and commercially available vegetable and animal patties. Among sugars, the contents of fructose and maltose were respectively high in commercial meat patties (CMPs) and SMPs. SMPs were low in saturated and trans-fat, and high in ω-3 fatty acids. The contents (in descending order) of leucine > phenylalanine > threonine > isoleucine were high in SMPs and commercial vegetable patties (CVPs). Qualitative and quantitative findings of Escherichia coli (E. coli) and Staphylococcus aureus were all negative. Our SMPs had high lightness (L*), low redness (a*), and low yellowness (b*). The hardness, chewiness, and resilience of our SMPs were lower than those of other vegetable and animal patties. Considering our results, the method of manufacturing SMPs developed in the present study allows meat to be flavored without significant nutritional differences compared with commercially available CMPs. Our findings provide a base for studies on future meat alternatives.