Free Nucleotides Research Articles

Effect of co-fermentation system with isolated new yeasts on soymilk: microbiological, physicochemical, rheological, aromatic, and sensory characterizations.

The beany flavor adversely influences consumer acceptance of soymilk (SM) products. Thus, in this work, the co-fermentation of isolated new yeasts (Kluyveromyces marxianus SP-1, Candida ethanolica ATW-1, and Pichia amenthionina Y) and Kluyveromyces marxianus K (a commercial yeast) along with an XPL-1 starter (including five strains of lactic acid bacteria (LAB)) was utilized to mend the beany flavor of fermented SM (FSM) beverages. Probiotic count, pH, titratable acidity, syneresis, water holding capacity, rheological characteristics, and sensory attributes were investigated. Furthermore, the free amino acids, nucleotides, and volatile compounds (VCs) were analyzed, also presenting the collected VC data by exploiting a principal component analysis (PCA) and a heatmap with a hierarchical cluster analysis. The co-fermentation with Kluyveromyces marxianus SP-1 and K remarkably enhanced the LAB strain growth and acid production, improving the rheological attributes, whereas that of yeast along with XPL-1 as a mullite starter could reduce the beany odor. PCA chart displayed that higher amounts of alcohols, ketones, acids, and esters that significantly improved the flavor quality of FSM beverages were generated throughout the co-fermentation process. The co-fermentation with Pichia amenthionina Y generated the highest acetoin (36.19%) and diacetyl (2.02%), thus improving the overall acceptance of FSM, as well as the sensory characteristics of FSM beverages with the highest umami, sweet, odorless amino acids, and umami nucleotides, and the lowest content of alcohol and inosine. Taken together, the co-fermentation of Pichia amenthionina Y along with XPL-1 within SM provides novel insights regarding the development of FSM and fermented beverages.

Prozdrowotne właściwości larw barciaka większego Galleria mellonella L.

The larvae of the greater moth (Galleria mellonella L.), also known as the wax fluke, are the largest bee-keeping pest that feeds on beeswax. For supplementation purposes, mainly 10% and 20% ethanol extracts from fresh larvae are prepared, which contain free amino acids, nucleotides and nucleosides, carbohydrates, proteins, fatty acids, bioelements, vitamins, enzymes. The extracts show numerous biological properties: cardiotropic, neuroprotective, antioxidant, anti-inflammatory, antimicrobial, they increase the function of the immune system, preventing infections with pathogenic microorganisms (bacteria, viruses), frequent especially in spring and autumn. The ability to increase the body’s resistance to physical and mental stress factors, and to increase mental performance, can be used, especially in weakened and convalescent people, also in children. Greater moth larvae are widely used in immunological, biological and genetic research. They also digest hardly biodegradable polyethylene.

Natural feed additive containing Saccharomyces cerevisiae-originated free nucleotides improves innate immunity, gut histology and disease resistance in Nile tilapia

A 60-day feeding trial was conducted to assess the effects of dietary Saccharomyces cerevisiae-originated free nucleotides on growth, innate immunity, intestinal histomorphometry and disease resistance in Nile tilapia, Oreochromis niloticus (average initial weight of 7.85 g). Five diets (treatments) were prepared: negative control diet (NCD) - basal diet without additives; positive control diet (PCD) - basal diet supplemented with a commercial nucleotide-based additive at 170 ppm; and three experimental diets, N170, N320 and N470, supplemented with S. cerevisiae-originated free nucleotides at 170, 320 and 470 ppm, respectively. There were no significant differences in production parameters after the trial period. Nevertheless, the complement hemolytic activity was higher in fish subjected to the three experimental diets, while serum lysozyme concentration was higher in the N470 group compared with both controls. Intestinal villi height and density were greater in fish fed the N470 diet compared with the NCD group. In addition, fish receiving the three experimental diets showed higher absorption surface area magnification ratio, with the greatest values being observed in the N470 group. Lastly, the highest survival rate following Aeromonas hydrophila challenge was recorded in the N470 group. Taken together, the present findings indicate that supplementation of feed with S. cerevisiae-originated free nucleotides at 470 ppm improves non-specific immune responses and intestinal microstructure morphology as well as increases survival rate after bacterial infection in Nile tilapia. Therefore, this blend of natural nucleotides has the potential of being used as an alternative to antimicrobials when included as an additive in aquaculture feeds.

Too much and never enough: Synthetic excess and metabolic inefficiency of aneuploidy in tumorigenesis

Aneuploidy is a hallmark feature of cancer. Relative to normal diploid tissue, aneuploid tumors are often net copy number increased, in a range from hyper‐diploid to tetraploid. This results in widespread replication stress and proteotoxic stress that affects diverse cellular complexes. Utilizing multi‐omics approaches in novel models of cancer‐associated copy number alterations derived from diploid human mammary epithelial cells, we uncovered an aneuploidy stress response driven in part by Myc that transcriptionally upregulates biomass production capacity and stresses metabolic capacity. Basal and peak glycolytic and oxidative rates are increased in aneuploid cells compared to diploids, however free nucleotide pools and metabolic intermediates, particularly pyrimidines, are depleted. CRISPR screens revealed a universally increased dependence on de novo pyrimidine synthesis genes in aneuploid cells with diverse CNAs, suggesting a struggle to match pyrimidine synthesis capacity with need. The biosynthetic demand and inefficiency driven by aneuploidy may partially explain the switch to aerobic glycolysis in tumors and present vulnerabilities to be therapeutically exploited.

Effect of pH on Rhodomonas salina growth, biochemical composition, and taste, produced in semi-large scale under sunlight conditions

Rhodomonas salina is a microalgal species, belonging to the cryptophytes, and is widely used as aquaculture feed because of its high nutritional profile and phycoerythrin content. This study investigated the effect of pH on the growth, biochemical composition, and taste of R. salina when cultivated on a semi-large scale under natural light conditions. Two tubular photobioreactors (200 L) were used for the cultivation of R. salina with sunlight as the only illumination source. Two different pH setpoints were applied, 7 and 8.5. Optimal temperature and nutrient conditions were applied, according to previous research findings. The results demonstrated that the productivity of R. salina was higher at pH 7, 0.06–0.14 gdry weight L−1 day−1, compared to pH 8.5, 0.03–0.12 gdry weight L−1 day−1. It was found that protein and total fatty acid concentrations were higher in the biomass that was produced at pH 8.5, 33.7% and 12.3% of dry weight, respectively, while at pH 7, the protein content was 31.9% and the total fatty acids 8.8% of dry weight. The phycoerythrin concentration, like protein, was higher at pH 8.5, 2.7% of dry weight, compared to pH 7, 1% of dry weight. The free amino acid and nucleotide profile of R. salina was affected by the pH, resulting in increased equivalent umami concentration at pH 7. For the sensory evaluation, an expert panel on algae flavors evaluated the effect of pH on the taste of R. salina, reporting that the biomass that was produced at pH 7 had more umami flavor than the biomass that was produced at pH 8.5, which was evaluated as more bitter.

Effects of short-term low salinity stress on non-volatile flavor substances of muscle and hepatopancreas in Portunus trituberculatus

In order to determine the effects of different short - term low salinity stress on non - volatile flavor substances of Portunus trituberculatus, four salinity levels (24‰, 16‰, 12‰ and 8‰) were selected. The contents of free amino acids (FAA), nucleotides and lactic acid in both muscle and hepatopancreas of P. trituberculatus were determined by HPLC and spectrophotometry respectively. Results showed that muscle contained a relatively higher level of sweetish free amino acids (SFA), however short - term low salinity stress greatly increased the level of bitter free amino acid (BFA); hepatopancreas contained a higher level of umami free amino acids (UFA) and a higher equivalent umami concentration (EUC) value, which increased significantly with the decrease of salinity level and the degree of umami flavor reached the highest when salinity dropped to 8‰. The lactic acid content in muscle increased at 12‰ and then decreased significantly at 8‰ while no significant differences were observed in hepatopancreas. Results above revealed the flavor change in two edible parts of P.trituberculatus under short - term low salinity stress which provided a guidance for more systematic nutritional flavor evaluation in breeding process and consumption demand.

Oral administration of nucleotides in calves: Effects on oxidative status, immune response, and intestinal mucosa development

The present work aimed to investigate the effects of nucleotide oral administration on oxidative stress biomarkers, immune responses, gut morphology, serum biochemical parameters, and growth performance in calves from birth to 25 d of life. A total of 40 male Holstein Friesian calves were randomly divided in 2 groups. All the calves were born and reared on the same commercial dairy farm. They were fed the same colostrum, milk replacer, and calf starter. Five grams/head of an additive were orally administered with a syringe directly in the mouth to calves of the nucleotide group (NG). The additive contained 74.12 g/100 g of nucleic acids from hydrolyzed yeast, and 75.38% was free nucleotide sodium salt. The other group represented the negative control (CG). At 25 d of life all of the calves were slaughtered. Calves supplemented with nucleotides had a higher final live weight and improved average daily gain, which was associated with better efficiency of nutrient use. Oral nucleotide administration did not affect IgG absorption efficiency; however, NG calves showed greater duodenum villi length and higher crypt depth compared with CG. Oral nucleotide administration increased the activity of antioxidant enzymes (superoxide dismutase, catalase and glutathione peroxidase) and the antioxidant capacity [ferric reducing antioxidant power and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) scavenging activity] both in plasma and in liver. An enhanced ability of cells to counter reactive oxygen species- and reactive nitrogen species-mediated damage was also observed in peripheral blood mononuclear cells from NG. The findings highlight the effectiveness of oral nucleotide administration, and potentially dietary supplementation of nucleotides, in boosting oxidative and immune status in newborn calves.

The Effects of Exogenous Salicylic Acid on Endogenous Phytohormone Status in Hordeum vulgare L. under Salt Stress.

Acclimation to salt stress in plants is regulated by complex signaling pathways involving endogenous phytohormones. The signaling role of salicylic acid (SA) in regulating crosstalk between endogenous plant growth regulators’ levels was investigated in barley (Hordeum vulgare L. ‘Ince’; 2n = 14) leaves and roots under salt stress. Salinity (150 and 300 mM NaCl) markedly reduced leaf relative water content (RWC), growth parameters, and leaf water potential (LWP), but increased proline levels in both vegetative organs. Exogenous SA treatment did not significantly affect salt-induced negative effects on RWC, LWP, and growth parameters but increased the leaf proline content of plants under 150 mM salt stress by 23.1%, suggesting that SA enhances the accumulation of proline, which acts as a compatible solute that helps preserve the leaf’s water status under salt stress. Changes in endogenous phytohormone levels were also investigated to identify agents that may be involved in responses to increased salinity and exogenous SA. Salt stress strongly affected endogenous cytokinin (CK) levels in both vegetative organs, increasing the concentrations of CK free bases, ribosides, and nucleotides. Indole-3-acetic acid (IAA, auxin) levels were largely unaffected by salinity alone, especially in barley leaves, but SA strongly increased IAA levels in leaves at high salt concentration and suppressed salinity-induced reductions in IAA levels in roots. Salt stress also significantly increased abscisic acid (ABA) and ethylene levels; the magnitude of this increase was reduced by treatment with exogenous SA. Both salinity and SA treatment reduced jasmonic acid (JA) levels at 300 mM NaCl but had little effect at 150 mM NaCl, especially in leaves. These results indicate that under high salinity, SA has antagonistic effects on levels of ABA, JA, ethylene, and most CKs, as well as basic morphological and physiological parameters, but has a synergistic effect on IAA, which was well exhibited by principal component analysis (PCA).

Effects of temporary rearing time under salinity 7 on the non-volatile flavorings and fatty acids of Eriocheir sinensis

In this paper, the effects of temporary rearing time on the flavor substances of the edible parts of river crabs under the condition of 7 psu were determined based on high performance liquid chromatography (HPLC) and gas chromatograph (GC) techniques. The results showed that the content of free amino acids in the edible parts of river crabs could be increased under the condition of salinity of 7 psu. The content of organic acids in the edible part of Chinese mitten crabs was also affected by temporary culture under salinity 7 psu, and the change trend was the same as that of amino acids. The treatment of 7 psu can promote the accumulation of free nucleotides in the muscles, hepatopancreas and gonads of the crabs, and the equivalent umami concentration (EUC) values in the muscles and gonads of the crabs have been significantly increased. However, salinity 7 reduced the proportion of PUFA in muscles and gonads, resulting in a certain amount of nutrient loss. In summary, 15 days after treatment with salinity 7 can significantly improve the flavor of the edible parts of Chinese mitten crabs.

Effect of different thermal processing methods on water-soluble taste substances of tilapia fillets

The composition of free amino acids (FAAs) and nucleotides has an important effect on fish taste. In this study, the content of FAAs and nucleotides in raw, steamed, boiled and air-fried tilapia fillets were quantitatively analyzed. Thermal processing significantly influenced the composition of FAAs and nucleotides in tilapia. Moreover, there were substantial differences in the nutritional composition of tilapia processed by steaming, boiling and air frying. Compared with raw fillets, the total FAA (TFAA) increased after steaming, boiling, and air frying. The main umami and sweet amino acids in tilapia fillets were aspartic acid and glycine. The total nucleotide content of fresh tilapia was 299 mg/100 g, which increased to 305 and 366 mg/100 g after steaming and air frying, respectively, and decreased to 273 mg/100 g after boiling. The main taste nucleotide inosine monophosphate (IMP) with a taste activity value greater than 1, contributed greatly to the taste of tilapia. Equivalent umami concentration was used for taste evaluation: air frying > steaming > boiling. Therefore, our findings establish that the water-soluble taste substances of tilapia fillets, and the mutual regulatory effects of different compounds varied with different thermal processing methods. Hence, tilapia can be thermally processed according to different taste requirements.

Extraction and analysis of free amino acids and 5′-nucleotides, the key contributors to the umami taste of seaweed

Assessing the umami taste of seaweed on a chemical level can inform the use and selection of seaweed in European cuisine. Accordingly, we developed a method for the simultaneous extraction, separate clean-up and analysis of 21 free amino acids and 10 free nucleotides by reversed phase and mixed-mode HPLC respectively. Of multiple mouth emulating solvents, extracting in Milli-Q at 35 °C was found most suitable. This method showed good linearity (R2 > 0.9996), resolution (Rs ≥ 1.5) and picomole detection limits, and was successfully applied to determine the Equivalent Umami Concentration (EUC) and Taste Activity Values (TAV) of seven Dutch seaweed species. Phaeophyceae showed the highest EUC, followed by Chlorophyceae and Rhodophyceae (≈ 9.5, 3.7 and 1.1 g/100 g respectively). Glutamic acid always exceeded the TAV, while other umami compounds were species specific. Our method can accurately predict umami intensity and therefore contributes towards species selection for the European palette.

Structures of outer-arm dynein array on microtubule doublet reveal a motor coordination mechanism

Thousands of outer-arm dyneins (OADs) are arrayed in the axoneme to drive a rhythmic ciliary beat. Coordination among multiple OADs is essential for generating mechanical forces to bend microtubule doublets (MTDs). Using electron microscopy, we determined high-resolution structures of Tetrahymena thermophila OAD arrays bound to MTDs in two different states. OAD preferentially binds to MTD protofilaments with a pattern resembling the native tracks for its distinct microtubule-binding domains. Upon MTD binding, free OADs are induced to adopt a stable parallel conformation, primed for array formation. Extensive tail-to-head (TTH) interactions between OADs are observed, which need to be broken for ATP turnover by the dynein motor. We propose that OADs in an array sequentially hydrolyze ATP to slide the MTDs. ATP hydrolysis in turn relaxes the TTH interfaces to effect free nucleotide cycles of downstream OADs. These findings lead to a model explaining how conformational changes in the axoneme produce coordinated action of dyneins.

EVALUATION OF NON-VOLATILE KEY TASTE COMPOUNDS OF AIR-DRIED HERRING (Clupea pallasii) FILLET

An experiment was conducted to quantify the non-volatile key taste compounds, including free amino acids (FAAs), nucleotides, inorganic ions, organic acids, and bases of dried herring (Clupea pallassi) fillet (DHF). A taste recombinant was also prepared using those key taste compounds to compare the taste profiles with water soluble extracts (WSE) by sensory perception. The total FAA of DHF muscle was 12.25 mg/g on dry matter basis. The most dominant FAAs were alanine, glycine, lysine, and histidine. The nucleotides were identified as 5´-adenosine monophosphate (AMP) (1.74 mg/100 g) and 5´-inosine monophosphate (IMP) (5.28 mg/100 g). Potassium, phosphate, and chloride were the major inorganic ions while trimethylamine oxide, creatine, and lactic acid were the major organic bases. Sensory evaluation showed that WSE markedly increased the intensities of umami, mouthfulness, thickness, and taste continuity of Japanese noodle soup (JNS). However, addition of taste recombinant to the JNS did not bring any changes in the taste intensities of saltiness, umami, mouthfulness, taste continuity, and thickness of the soup. Results of this study indicated that the taste recombinant was lacking of some important key taste compounds that might be formed during drying of herring fillet.

Deep Sequencing Reveals the Comprehensive CRISPR-Cas9 Editing Spectrum in Bombyx mori.

Application of the clustered regularly interspaced short palindromic repeats associated 9 (CRISPR-Cas9) technology has revolutionized biology by greatly enhancing the ability to introduce mutations into DNA for research and prospective therapeutic purposes. However, the understanding of Cas9 editing outcomes is still limited. Previously, it was considered that Cas9 introduces stochastic insertions or deletions (indels) at the target site. In the current study, we performed in vivo multiplex editing, deep sequencing, and comprehensive analysis of its editing outcomes in Bombyx mori (B. mori). A total of 31161 editing events from 9 single-guide RNA (sgRNA) sites in 16 individuals were generated and analyzed, and we found that Cas9 introduces mutations with some regularity rather than via stochastic indels. The editing efficiency varies with sgRNA sequences, individuals, and orientation. Small deletions account for the vast majority of mutated sequences, followed by a small fraction of substitutions and insertions. The most likely mutations are deletions between two microhomologous sequences or single-base deletions at the cleavage site in the absence of microhomologous pairs. Insertions are formed by diverse mechanisms, including direct acquisition of free genomic fragments, duplication of broken ends, replication of adjacent sequences, or random addition of free nucleotides. The above results indicate that the Cas9 editing spectrum is reproducible and predictable. Thus, our findings enable a deeper understanding of Cas9-mediated mutagenesis and better design of genome editing experiments, as well as elucidate the DNA double-strand break repair processes in B. mori.

Change Regularity of Taste and the Performance of Endogenous Proteases in Shrimp (Penaens vannamei) Head during Autolysis.

This study evaluated the food safety and proximate composition of shrimp head (SH). Potentially toxic elements in SH were below European Union legislation limits. SH had a high content of tasting amino acids (sweet and umami amino acids was 57%) and a high content of functional amino acids (essential amino acids was 37%). Moreover, the changes of flavor and key umami molecules in SH were studied by sensory evaluation, electronic tongue, electronic nose, automated amino acid analyzer, and high performance liquid chromatography (HPLC). The results showed that the significant difference of flavor in SH happened during autolysis. SH with autolysis had the best umami taste at 6 h, which may result from the synergistic work of free amino acids and nucleotide related compounds. Additionally, the performance of endogenous proteases in SH was investigated to efficiently analyze autolysis. The optimum pH and temperature of endogenous proteases in SH were 7.5 and 50 °C, respectively. The autolysis of SH depends on two endogenous proteases (~50 kDa and ~75 kDa). These results suggest that the formation of flavor in SH during autolysis can be controlled, which could provide guidance for SH recycle. SH could consider as one of the food materials for producing condiments.

Cryo‐EM Structures of Outer‐arm Dynein Array Bound to Microtubule Doublet Reveal a Mechanism for Motor Coordination

Eukaryotic cilia are evolutionarily conserved organelles that are responsible for cellular motility, sensory reception, embryonic development and intercellular communication. A typical motile cilium is characterized by its ʹ9 + 2ʹ scaffold, composed of nine microtubule doublets (MTDs), and a central pair complex (CPC). Two rows of axonemal dyneins, the outer-arm dynein (OAD) and inner-arm dynein (IAD), power the ciliary beating by sliding two adjacent MTDs. OAD is the key motor protein that generates the majority of mechanical forces required for this fundamental cellular process. A complete OAD is ~1.5-2 megadalton in size and contains two (mammals) or three (in ciliates and algae) heavy chains (HCs), two intermediate chains (ICs) and a variety of light chains (LCs). Each OAD is divided into a head region that contains AAA+ rings for ATP hydrolysis and a tail that holds together the whole complex. The tail is permanently attached to MTD A-tubule, while the head region of each HC contains a microtubule-binding domain (MTBD) which binds and releases MTD B-tubule depending on the nucleotide-binding states. Thousands of OADs are arrayed in the axoneme to drive a rhythmic ciliary beat, which is required to OADs locally synchronize their conformations and coordinate with each other. Current understanding is mainly based on cryo-electron tomography (cryo-ET) which suggests that adjacent OADs are indirectly connected via a series of linker structures. Nevertheless, it is largely unclear how the OAD arrays are formed and why the array is important for ciliary beat. Furthermore, the mechanism underlying the motor coordination remains elusive. We set out to reveal the mechanism of motor coordination to shed lights on how the conformational changes of arrayed OAD are linked to force generation and beating propagation. We use the model system T. thermophila for biochemical and cryo-EM analysis. T. thermophila OAD is the first discovered dynein, which contains three HCs (α-, β- and γ-HC), two ICs (IC2 and IC3) and a set of indefinite LCs. We reconstitute the purified OADs onto MTD to mimic the native OAD arrays and determined the structures of free OAD and OAD arrays bound to MTDs (OAD-MTD) in two different microtubule-binding states (MTBS) by cryo-electron microscopy (cryo-EM). We show that microtubule binding induces free OADs to spontaneously adopt a parallel conformation, which is primed for array formation in a tail-to-head (TTH) manner. The conformations of arrayed OADs are synchronized in either microtubule-binding state. The array involves extensive network interactions and is coordinately remodeled when OADs take one step forward from MTBS-1 to MTBS-2. The TTH interactions remain nearly unchanged in both states but need to be broken to allow MTBS alteration during mechanochemical cycle. Nucleotide treatment on the OAD-MTD array reveals that it is the ATP hydrolysis that temporarily relaxes the TTH interfaces to allow free nucleotide cycle of downstream OADs. In combination with previously reported cryo-ET structures, we propose a most detailed mechanism of how OADs coordinate with each other to move one step on MTD and a possible model for ciliary beat propagation.

Flavor and Metabolite Profiles of Meat, Meat Substitutes, and Traditional Plant-Based High-Protein Food Products Available in Australia

Demand for plant-based proteins and plant-based food products is increasing globally. This trend is driven mainly by global population growth and a consumer shift towards more sustainable and healthier diets. Existing plant-based protein foods and meat mimetics often possess undesirable flavor and sensory properties and there is a need to better understand the formation of desirable meat-like flavors from plant precursors to improve acceptance of novel high-protein plant foods. This study aimed to comprehensively characterize the non-volatile flavor metabolites and the volatiles generated in grilled meat (beef, chicken, and pork) and compare these to commercially available meat substitutes and traditional high-protein plant-based foods (natto, tempeh, and tofu). Solid phase microextraction with gas-chromatography mass-spectrometry was used for elucidation of the flavor volatilome. Untargeted characterization of the non-volatile metabolome was conducted using Orbitrap mass spectrometry and Compound DiscovererTM datamining software. The study revealed greater diversity and higher concentrations of flavor volatiles in plant-based foods in comparison to grilled meat, although the odor activity of specific volatiles was not considered. On average, the total amount of volatiles in plant-based products were higher than in meat. A range of concentrations of free amino acids, dipeptide, tripeptides, tetrapeptides, nucleotides, flavonoids, and other metabolites was identified in meat and plant-based foods.

Enhanced flavor strength of broth prepared from chicken following short-term frozen storage

This study investigated the effect of frozen storage periods (0, 2, 4, 6, or 8 weeks) of raw meat and stewing on the flavor of chicken broth. With the increased storage duration of frozen raw material, the contents of the free amino acids, nucleotides and mineral elements in the broth decreased significantly, especially within the first 4 weeks, and then increased significantly. Meanwhile, the volatile compounds showed the reverse trend. The results from the E-nose, E-tongue and sensory evaluation indicated a progressive difference in overall flavor profiles between the samples. The sensory scores of the meaty and fatty traits reached a maximum as raw chicken meat was stored for 4 weeks at −18 °C, which should be related to the increased contents of aldehydes and 2-pentyl furan. Overall, the limited storage duration of frozen raw meat can enhance the flavor of chicken broth.

Chromatin Organization and Phase Separation of Histones

A concept that formation of heterochromatin is driven by liquid-liquid phase separation (LLPS) has emerged in recent years. Using live cell fluorescence imaging, we show that the linker histone H1 mediates LLPS of heterochromatin in the human cell nucleus. We also find that the core histone H2A readily undergoes LLPS with DNA and nucleosome arrays of varying length in vitro and in doing so does not disrupt the nucleosome core, as observed by FRET measurements. LLPS with histones is enhanced by the presence of free nucleotides like ATP. Consistently, in cells under ATP depleted conditions, the number of H1 rich liquid-like domains in their nuclei are significantly diminished. These findings add yet another dimension to the role of histones in organizing and maintaining the genome inside the nucleus.

Metabolomics and lipidomics analyses delineating Hfq deletion- induced metabolic alterations in Vibrio alginolyticus

Vibrio alginolyticus, a Gram-negative halophilic zoonotic pathogen, can cause bacterial hemorrhagic septicemia in various aquaculture species and wound infections in humans. V. alginolyticus brings about huge economic losses in aquaculture all around the world. Hfq, a ubiquitous RNA-binding protein, is a global post-transcriptional regulator in most bacteria and some archaea, by facilitating base-pairing between sRNAs and their mRNA targets, and is essential for adaptation to different environments and growth conditions. Despite its important roles in the stress adaptation, virulence, nutrient utilization, and colony morphology in V. alginolyticus, further research is required to shed light on the pathways modulated by Hfq. In this study, we used metabolomics and lipidomics analytical approaches based on ultrahigh-performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) to understand the impact of Hfq on the metabolic profile of V. alginolyticus. The deletion of the hfq gene resulted in the growth deficiency and cell elongation of V. alginolyticus cultured in the rich medium. Substantial metabolic alterations, mainly involved in amino acids, nucleotides, fatty acids, and lipids, were observed in the hfq deletion mutant strain (Δhfq) when compared to the wild-type (WT) strain and its complemented strain (hfq+). Notably, the hfq deletion induced accumulations of free fatty acids (FFAs) and nucleotides, but decreased levels of most amino acids (e.g., branched amino acids, aromatic amino acids, and aspartate family amino acids). Meanwhile, the majority of lipids (e.g., lysophosphatidylglycerol, lysophosphatidylethanolamine, phosphatidylcholine, phosphatidic acid, cardiolipin, sphingomyelin, some phosphatidylglycerol and phosphatidylethanolamine with mono- or poly-unsaturated fatty acids) were decreased, while ceramide, diacylglycerol, triacylglycerol and some phosphatidylglycerol and phosphatidylethanolamine with saturated fatty acids increased in the Δhfq strain. These data demonstrated that the hfq deletion triggered de novo synthesis of FFAs and nucleosides and enhanced the catabolism of phospholipid and sphingomyelin to further regulate cell division and growth. Additionally, the hfq deletion could inhibit the synthesis of the branched amino acids, aromatic amino acids and enhance the TCA cycle to influence cell stress defenses and energy metabolism. Our research indicates that Hfq can regulate pathways associated with cell division, growth and stress resistance, being an important metabolic modulator in V. alginolyticus.