HCF-1 Regulates De Novo Lipogenesis through a Nutrient-Sensitive Complex with ChREBP.

Carbohydrate response element binding protein (ChREBP) is a key transcriptional regulator of de novo lipogenesis (DNL) in response to carbohydrates and in hepatic steatosis. Mechanisms underlying nutrient modulation of ChREBP are under active investigation. Here we identify host cell factor 1 (HCF-1) as a previously unknown ChREBP-interacting protein that is enriched in liver biopsies of nonalcoholic steatohepatitis (NASH) patients. Biochemical and genetic studies show that HCF-1 is O-GlcNAcylated in response to glucose as a prerequisite for its binding to ChREBP and subsequent recruitment of OGT, ChREBP O-GlcNAcylation, and activation. The HCF-1:ChREBP complex resides at lipogenic gene promoters, where HCF-1 regulates H3K4 trimethylation to prime recruitment of the Jumonji C domain-containing histone demethylase PHF2 for epigenetic activation of these promoters. Overall, these findings define HCF-1's interaction with ChREBP as a previously unappreciated mechanism whereby glucose signals are both relayed to ChREBP and transmitted for epigenetic regulation of lipogenic genes.

Taxonomic Distribution and Molecular Evolution of Mytilectins.

R-type lectins are a widespread group of sugar-binding proteins found in nearly all domains of life, characterized by the presence of a carbohydrate-binding domain that adopts a ß-trefoil fold. Mytilectins represent a recently described subgroup of ß-trefoil lectins, which have been functionally characterized in a few mussel species (Mollusca, Bivalvia) and display attractive properties, which may fuel the development of artificial lectins with different biotechnological applications. The detection of different paralogous genes in mussels, together with the description of orthologous sequences in brachiopods, supports the formal description of mytilectins as a gene family. However, to date, an investigation of the taxonomic distribution of these lectins and their molecular diversification and evolution was still lacking. Here, we provide a comprehensive overview of the evolutionary history of mytilectins, revealing an ancient monophyletic evolutionary origin and a very broad but highly discontinuous taxonomic distribution, ranging from heteroscleromorphan sponges to ophiuroid and crinoid echinoderms. Moreover, the overwhelming majority of mytilectins display a chimera-like architecture, which combines the ß-trefoil carbohydrate recognition domain with a C-terminal pore-forming domain, suggesting that the simpler structure of most functionally characterized mytilectins derives from a secondary domain loss.

I-branched carbohydrates as emerging effectors of malignant progression.

Cell surface carbohydrates, termed "glycans," are ubiquitous posttranslational effectors that can tune cancer progression. Often aberrantly displayed or found at atypical levels on cancer cells, glycans can impact essentially all progressive steps, from malignant transformation to metastases formation. Glycans are structural entities that can directly bind promalignant glycan-binding proteins and help elicit optimal receptor-ligand activity of growth factor receptors, integrins, integrin ligands, lectins, and other type-1 transmembrane proteins. Because glycans play an integral role in a cancer cell's malignant activity and are frequently uniquely expressed, preclinical studies on the suitability of glycans as anticancer therapeutic targets and their promise as biomarkers of disease progression continue to intensify. While sialylation and fucosylation have predominated the focus of cancer-associated glycan modifications, the emergence of blood group I antigens (or I-branched glycans) as key cell surface moieties capable of modulating cancer virulence has reenergized investigations into the role of the glycome in malignant progression. I-branched glycans catalyzed principally by the I-branching enzyme GCNT2 are now indicated in several malignancies. In this Perspective, the putative role of GCNT2/I-branching in cancer progression is discussed, including exciting insights on how I-branches can potentially antagonize the cancer-promoting activity of ß-galactose-binding galectins.

Manipulating mtDNA in vivo reprograms metabolism via novel response mechanisms.

Mitochondria have been increasingly recognized as a central regulatory nexus for multiple metabolic pathways, in addition to ATP production via oxidative phosphorylation (OXPHOS). Here we show that inducing mitochondrial DNA (mtDNA) stress in Drosophila using a mitochondrially-targeted Type I restriction endonuclease (mtEcoBI) results in unexpected metabolic reprogramming in adult flies, distinct from effects on OXPHOS. Carbohydrate utilization was repressed, with catabolism shifted towards lipid oxidation, accompanied by elevated serine synthesis. Cleavage and translocation, the two modes of mtEcoBI action, repressed carbohydrate rmetabolism via two different mechanisms. DNA cleavage activity induced a type II diabetes-like phenotype involving deactivation of Akt kinase and inhibition of pyruvate dehydrogenase, whilst translocation decreased post-translational protein acetylation by cytonuclear depletion of acetyl-CoA (AcCoA). The associated decrease in the concentrations of ketogenic amino acids also produced downstream effects on physiology and behavior, attributable to decreased neurotransmitter levels. We thus provide evidence for novel signaling pathways connecting mtDNA to metabolism, distinct from its role in supporting OXPHOS.

Potential Carbohydrate Regulation Mechanism Underlying Starvation-Induced Abscission of Tomato Flower.

Tomato flower abscission is a critical agronomic problem directly affecting yield. It often occurs in greenhouses in winter, with the weak light or hazy weather leading to insufficient photosynthates. The importance of carbohydrate availability in flower retention has been illustrated, while relatively little is understood concerning the mechanism of carbohydrate regulation on flower abscission. In the present study, we analyzed the responding pattern of nonstructural carbohydrates (NSC, including total soluble sugars and starch) and the potential sugar signal pathway involved in abscission regulation in tomato flowers under shading condition, and their correlations with flower abscission rate and abscission-related hormones. The results showed that, when plants suffer from short-term photosynthesis deficiency, starch degradation in flower organs acts as a self-protection mechanism, providing a carbon source for flower growth and temporarily alleviating the impact on flower development. Trehalose 6-phosphate (T6P) and sucrose non-fermenting-like kinase (SnRK1) signaling seems to be involved in adapting the metabolism to sugar starvation stress through regulating starch remobilization and crosstalk with IAA, ABA, and ethylene in flowers. However, a continuous limitation of assimilating supply imposed starch depletion in flowers, which caused flower abscission.

Transcription Profiles Reveal Age-Dependent Variations of Photosynthetic Properties and Sugar Metabolism in Grape Leaves (Vitis vinifera L.).

Leaves, considered as the 'source' organs, depend on the development stages because of the age-dependent photosynthesis and assimilation of leaves. However, the molecular mechanisms of age-dependent limitations on the function of leaves are seldom reported. In the present study, the photosynthesis-related characteristics and photoassimilates were investigated in grape leaves at six different age groups (Ll to L6) at micro-morphological, biochemical, and molecular levels. These results showed lower expression levels of genes associated with stomatal development, and chl biosynthesis resulted in fewer stomata and lowered chlorophyll a/b contents in L1 when compared to L3 and L5. The DEGs between L5 and L3/L1 were largely distributed at stomatal movement, carbon fixation, and sucrose and starch metabolism pathways, such as STOMATAL ANION CHANNEL PROTEIN 1 (SLAC1), FRUCTOSE-1,6-BISPHOSPHATE ALDOLASE (FBA1), SUCROSE-PHOSPHATE SYNTHASE (SPP1), and SUCROSE-PHOSPHATE PHOSPHATASE (SPS2, 4). These genes could be major candidate genes leading to increased photosynthesis capacity and sugar content in L5. The accumulation of starch grains in the chloroplast and palisade tissue of L5 and higher transcription levels of genes related to starch biosynthesis in L5 further supported the high ability of L5 to produce photoassimilates. Hence, our results provide insights for understanding different photosynthetic functions in age-dependent leaves in grape plants at the molecular level.

Transcriptomic and metabolomic analyses of non-structural carbohydrates in red maple leaves.

Plant sugars serve to balance nutrition, regulate development, and respond to biotic and abiotic stresses, whereas non-structural carbohydrates (NSCs) are essential energy sources that facilitate plant growth, metabolism, and environmental adaptation. To better elucidate the mechanisms of NSCs in red maple, ultrahigh-performance liquid chromatograph Q extractive mass spectrometry (UHPLC-QE-MS) and high-throughput RNA-sequencing were performed on green, red, and yellow leaves from a selected red maple mutant. In green leaves, the fructose phosphorylation process exhibited greater flux. In yellow leaves, sucrose and starch had a stronger capacity for synthesis and degradation, whereas in red leaves, there was a greater accumulation of trehalose and manninotriose. ArTPS5 positively regulated amylose, which was negatively regulated by ArFBP2, whereas ArFRK2 and ArFBP13 played a positive role in the biosynthesis of Sucrose-6P. Sucrose-6P also regulated anthocyanins and abscisic acid in red maple by affecting transcription factors. The results of this paper can assist with the control and optimization of the biosynthesis of NSCs in red maple, which may ultimately provide the foundation for influencing sugar production in Acer.

Leptin signaling in vagal afferent neurons supports the absorption and storage of nutrients from high-fat diet.

OBJECTIVE: Activation of vagal afferent neurons (VAN) by postprandial gastrointestinal signals terminates feeding and facilitates nutrient digestion and absorption. Leptin modulates responsiveness of VAN to meal-related gastrointestinal signals. Rodents with high-fat diet (HF) feeding develop leptin resistance that impairs responsiveness of VAN. We hypothesized that lack of leptin signaling in VAN reduces responses to meal-related signals, which in turn decreases absorption of nutrients and energy storage from high-fat, calorically dense food. METHODS: Mice with conditional deletion of the leptin receptor from VAN (Nav1.8-Cre/LepRfl/fl; KO) were used in this study. Six-week-old male mice were fed a 45% HF for 4 weeks; metabolic phenotype, food intake, and energy expenditure were measured. Absorption and storage of nutrients were investigated in the refed state. RESULTS: After 4 weeks of HF feeding, KO mice gained less body weight and fat mass that WT controls, but this was not due to differences in food intake or energy expenditure. KO mice had reduced expression of carbohydrate transporters and absorption of carbohydrate in the jejunum. KO mice had fewer hepatic lipid droplets and decreased expression of de novo lipogenesis-associated enzymes and lipoproteins for endogenous lipoprotein pathway in liver, suggesting decreased long-term storage of carbohydrate in KO mice. CONCLUSIONS: Impairment of leptin signaling in VAN reduces responsiveness to gastrointestinal signals, which reduces intestinal absorption of carbohydrates and de novo lipogenesis resulting in reduced long-term energy storage. This study reveals a novel role of vagal afferents to support digestion and energy storage that may contribute to the effectiveness of vagal blockade to induce weight loss.

The shell matrix and microstructure of the Ram's Horn squid: Molecular and structural characterization.

Molluscs are one of the most diversified phyla among metazoans. Most of them produce an external calcified shell, resulting from the secretory activity of a specialized epithelium of the calcifying mantle. This biomineralization process is controlled by a set of extracellular macromolecules, the organic matrix. In spite of several studies, these components are mainly known for bivalves and gastropods. In the present study, we investigated the physical and biochemical properties of the internal planispiral shell of the Ram's Horn squid Spirula spirula. Scanning Electron Microscope investigations of the shell reveal a complex microstructural organization. The saccharides constitute a quantitatively important moiety of the matrix, as shown by Fourier-transform infrared and solid-state nuclear magnetic resonance spectroscopies. NMR identified ß-chitin and additional polysaccharides for a total amount of 80% of the insoluble fraction. Proteomics was applied to both soluble and insoluble matrices and in silico searches were performed, first on heterologous metazoans models, and secondly on an unpublished transcriptome of Spirula spirula. In the first case, several peptides were identified, some of them matching with tyrosinase, chitinase 2, protease inhibitor, or immunoglobulin. In the second case, 39 hits were obtained, including transferrin, a serine protease inhibitor, matrilin, or different histones. The very few similarities with known molluscan shell matrix proteins suggest that Spirula spirula uses a unique set of shell matrix proteins for constructing its internal shell. The absence of similarity with closely related cephalopods demonstrates that there is no obvious phylogenetic signal in the cephalopod skeletal matrix.

Structural annotation of the conserved carbohydrate esterase vb_24B_21 from Shiga toxin-encoding bacteriophage Φ24B.

Shiga toxin-encoding bacteriophages transfer Shiga toxin genes to Escherichia coli and are responsible for the emergence of pathogenic bacterial strains that cause severe foodborne human diseases. Gene vb_24B_21 is the most highly conserved gene across sequenced Shiga bacteriophages. Protein vb_24B_21 (also termed 933Wp42 and NanS-p) is a carbohydrate esterase with homology to the E. coli chromosomally encoded NanS that deacetylates sialic acid in the intestinal mucus. To assist the functional characterization of vb_24B_21, we have studied its molecular structure by homology modelling its esterase domain and by elucidating the crystal structure of its uncharacterized C-terminal domain at the atomic resolution of 0.97 Å. Our modelling confirms that NanS from the E. coli host is the closest structurally characterized homolog to the esterase domain of vb_24B_21. Like NanS, vb_24B_21 has an atypical active site, comprising a simple catalytic dyad Ser-His and a divergent oxyanion hole. The crystal structure of the C-terminal domain reveals a lectin-like, jelly-roll ß-sandwich fold. The domain displays a prominent cleft that bioinformatics analysis predicts to be a carbohydrate binding site without catalytic properties. In summary, our study indicates that vb_24B_21 is a NanS-like atypical esterase that is assisted by a carbohydrate-binding module of yet undetermined binding specificity.

CBS homogenization mutation strategy narrows the glycan binding profile of a GlcNAc-specific lectin AANL.

Glycosylation plays important roles in many cellular processes, such as signal transduction, cell cycle progression and transcriptional regulation. However, the identification and analysis of glycosylation are severely hampered by the low specificity or avidity of antiglycan antibodies and lectins. We have reported that a lectin AANL, which has high specificity for terminal GlcNAc glycans and contains six carbohydrate binding sites (CBSs), was used to enrich O-GlcNAcylated peptides. To further improve AANL binding specificity, we designed a CBS-homogenization strategy and restructured six mutant lectins, known as AANL1-AANL6. Affinity chromatography with GlcNAc and isothermal titration calorimetry analysis indicated that the two mutants (AANL3 and AANL6) all maintained GlcNAc binding activity. AANL6 and AANL3 showed higher specificity for terminal GlcNAc glycans than AANL, as shown by the hemagglutination assay, cell binding assays and glycan microarray analysis, and AANL6 exhibited the highest specificity. The binding activity of AANL6 for O-GlcNAcylated peptides was shown by surface plasmon resonance assays. By AANL6 affinity chromatography enrichment and mass spectrometry analysis, 79 high-confidence and 21 putative O-GlcNAcylated sites were identified on 85 peptides mapped onto 54 proteins. Most of these sites were new sites compared with reported data. These results indicate that the enrichment capacity of AANL6 is higher than that of wild-type AANL. In conclusion, the CBS-homogenization mutation strategy was successful, and AANL6 was identified as a powerful tool for O-GlcNAcylation enrichment. Our research suggests that the CBS-homogenization strategy is valuable for improving the specificity of lectins with multiple CBSs.

Revisiting the Language of Glycoscience: Readers, Writers and Erasers in Carbohydrate Biochemistry.

The roles of carbohydrates in nature are many and varied. However, the lack of template encoding in glycoscience distances carbohydrate structure, and hence function, from gene sequence. This challenging situation is compounded by descriptors of carbohydrate structure and function that have tended to emphasise their complexity. Herein, we suggest that revising the language of glycoscience could make interdisciplinary discourse more accessible to all interested parties.

130 years of Plant Lectin Research.

Lectins are proteins with diverse molecular structures that share the ability to recognize and bind specifically and reversibly to carbohydrate structures without changing the carbohydrate moiety. The history of lectins started with the discovery of ricin about 130 years ago but since then our understanding of lectins has dramatically changed. Over the years the research focus was shifted from 'the characterization of carbohydrate-binding proteins' to 'understanding the biological function of lectins'. Nowadays plant lectins attract a lot of attention especially because of their potential for crop improvement and biomedical research, as well as their application as tools in glycobiology. The present review aims to give an overview of plant lectins and their applications, and how the field evolved in the last decades.

Deciphering carbohydrate metabolism during wheat grain development via integrated transcriptome and proteome dynamics.

Grain development of Triticum aestivum is being studied extensively using individual OMICS tools. However, integrated transcriptome and proteome studies are limited mainly due to complexity of genome. Current study focused to unravel the transcriptome-proteome coordination of key mechanisms underlying carbohydrate metabolism during whole wheat grain development. Wheat grains were manually dissected to obtain grain tissues for proteomics and transcriptomics analyses. Differentially expressed proteins and transcripts at the 11 stages of grain development were compared. Computational workflow for integration of two datasets related to carbohydrate metabolism was designed. For CM proteins, output peptide sequences of proteomic analyses (via LC-MS/MS) were used as source to search corresponding transcripts. The transcript that turned out with higher number of peptides was selected as bona fide ribonucleotide sequence for respective protein synthesis. More than 90% of hits resulted in successful identification of respective transcripts. Comparative analysis of protein and transcript expression profiles resulted in overall 32% concordance between these two series of data. However, during grain development correlation of two datasets gradually increased up to ~ tenfold from 152 to 655 °Cd and then dropped down. Proteins involved in carbohydrate metabolism were divided in five categories in accordance with their functions. Enzymes involved in starch and sucrose biosynthesis showed the highest correlations between proteome-transcriptome profiles. High percentage of identification and validation of protein-transcript hits highlighted the power of omics data integration approach over existing gene functional annotation tools. We found that correlation of two datasets is highly influenced by stage of grain development. Further, gene regulatory networks would be helpful in unraveling the mechanisms underlying the complex and significant traits such as grain weight and yield.

The sugar code: letters and vocabulary, writers, editors and readers and biosignificance of functional glycan-lectin pairing.

Ubiquitous occurrence in Nature, abundant presence at strategically important places such as the cell surface and dynamic shifts in their profile by diverse molecular switches qualifies the glycans to serve as versatile biochemical signals. However, their exceptional structural complexity often prevents one noting how simple the rules of objective-driven assembly of glycan-encoded messages are. This review is intended to provide a tutorial for a broad readership. The principles of why carbohydrates meet all demands to be the coding section of an information transfer system, and this at unsurpassed high density, are explained. Despite appearing to be a random assortment of sugars and their substitutions, seemingly subtle structural variations in glycan chains by a sophisticated enzymatic machinery have emerged to account for their specific biological meaning. Acting as 'readers' of glycan-encoded information, carbohydrate-specific receptors (lectins) are a means to turn the glycans' potential to serve as signals into a multitude of (patho)physiologically relevant responses. Once the far-reaching significance of this type of functional pairing has become clear, the various modes of spatial presentation of glycans and of carbohydrate recognition domains in lectins can be explored and rationalized. These discoveries are continuously revealing the intricacies of mutually adaptable routes to achieve essential selectivity and specificity. Equipped with these insights, readers will gain a fundamental understanding why carbohydrates form the third alphabet of life, joining the ranks of nucleotides and amino acids, and will also become aware of the importance of cellular communication via glycan-lectin recognition.

dbCAN2: a meta server for automated carbohydrate-active enzyme annotation.

Complex carbohydrates of plants are the main food sources of animals and microbes, and serve as promising renewable feedstock for biofuel and biomaterial production. Carbohydrate active enzymes (CAZymes) are the most important enzymes for complex carbohydrate metabolism. With an increasing number of plant and plant-associated microbial genomes and metagenomes being sequenced, there is an urgent need of automatic tools for genomic data mining of CAZymes. We developed the dbCAN web server in 2012 to provide a public service for automated CAZyme annotation for newly sequenced genomes. Here, dbCAN2 (http://cys.bios.niu.edu/dbCAN2) is presented as an updated meta server, which integrates three state-of-the-art tools for CAZome (all CAZymes of a genome) annotation: (i) HMMER search against the dbCAN HMM (hidden Markov model) database; (ii) DIAMOND search against the CAZy pre-annotated CAZyme sequence database and (iii) Hotpep search against the conserved CAZyme short peptide database. Combining the three outputs and removing CAZymes found by only one tool can significantly improve the CAZome annotation accuracy. In addition, dbCAN2 now also accepts nucleotide sequence submission, and offers the service to predict physically linked CAZyme gene clusters (CGCs), which will be a very useful online tool for identifying putative polysaccharide utilization loci (PULs) in microbial genomes or metagenomes.

Galactofuranose-Related Enzymes: Challenges and Hopes.

Galactofuranose is a rare form of the well-known galactose sugar, and its occurrence in numerous pathogenic micro-organisms makes the enzymes responsible for its biosynthesis interesting targets. Herein, we review the role of these carbohydrate-related proteins with a special emphasis on the galactofuranosidases we recently characterized as an efficient recombinant biocatalyst.

Asymmetrically Branched Precision Glycooligomers Targeting Langerin.

Asymmetrically branched precision glycooligomers are synthesized by solid-phase polymer synthesis for studying multivalent carbohydrate-protein interactions. Through the stepwise assembly of Fmoc-protected oligo(amidoamine) building blocks and Fmoc/Dde-protected lysine, straightforward variation of structural parameters such as the number and length of arms, as well as the number and position of carbohydrate ligands, is achieved. Binding of 1-arm and 3-arm glycooligomers toward lectin receptors langerin and concanavalin A (ConA) was evaluated where the smallest 3-arm glycooligomer shows the highest binding toward langerin, and stepwise elongation of one, two, or all three arms leads to decreased binding. When directly comparing binding toward langerin and ConA, we find that structural variation of the scaffold affects glycomimetic ligand binding differently for the different targets, indicating the potential to tune such ligands not only for their avidity but also for their selectivity toward different lectins.

Carbohydrate antigen expression and anti-pig antibodies in New World capuchin monkeys: Relevance to studies of xenotransplantation.

BACKGROUND: Old World non-human primates (OWNHPs) are used for preclinical pig-to-NHP studies. However, like pigs, OWNHPs express Neu5Gc, and therefore do not develop natural anti-Neu5Gc antibodies. New World NHPs (NWNHPs) have been reported not to express Neu5Gc. We investigated the potential of NWNHPs in xenotransplantation research. METHODS: We investigated expression of Gal, Neu5Gc, and Sda antigens on RBCs and PBMCs from humans, selected OWNHPs, and capuchin monkeys (a NWNHP). Serum anti-Gal and anti-Neu5Gc IgM and IgG levels were measured by ELISA. Binding of primate serum IgM and IgG to pig RBCs was measured by flow cytometry. RESULTS: (a) Neither humans, OWNHPs, or capuchin monkeys expressed Gal on their RBCs, but capuchins expressed Gal on PBMCs. Humans and capuchins did not express Neu5Gc on either RBCs or PBMCs, but OWNHPs expressed Neu5Gc on both cells. Sda was not expressed on any RBCs or PBMCs. (b) By ELISA, human and OWNHP, but not capuchin, sera showed IgM and IgG binding to Gal. Human and capuchin, but not OWNHP, sera demonstrated some binding to Neu5Gc. (c) Anti-Sda IgM/IgG antibodies were detected in OWNHP sera. Knockout of Sda on pig RBCs did not significantly reduce human and capuchin antibody binding. CONCLUSION: Capuchin monkeys could be surrogates for humans in experiments using RBCs, islets, neuronal cells, etc, from triple-knockout pigs (but may be too small to be used as recipients of pig organ grafts).

Evolution in primates by "Catastrophic-selection" interplay between enveloped virus epidemics, mutated genes of enzymes synthesizing carbohydrate antigens, and natural anti-carbohydrate antibodies.

"Catastrophic-selection" is an evolutionary mechanism, by which entire parental-populations are eliminated but very few mutated offspring survive and replace extinct parental-populations. The human natural anti-carbohydrate antibodies, anti-Gal and anti-Neu5Gc suggest the occurrence of catastrophic-selection events in primate evolution. Parental-populations synthesizing corresponding carbohydrate-antigens underwent extinction in viral epidemics, and few offspring survived. These offspring carried accidental mutations that inactivated carbohydrate-antigen synthesis and produced natural-antibody against the lost antigen. Such natural anti-carbohydrate antibody was produced against environmental carbohydrate-antigens (e.g., gastrointestinal bacteria). The carbohydrate-antigen in infected parental-populations was also synthesized on viruses by the host glycosylation-machinery. The natural-antibody in the offspring bound to the carbohydrate-antigen on infecting viruses produced in parental-populations, destroyed the viruses and protected these offspring from extinction. This process occurred in ancestral Old-World monkeys and apes synthesizing α-gal epitopes, which were replaced 20-30 million-years-ago by offspring lacking α-gal epitopes and producing natural anti-Gal antibody against this antigen, and later in hominins synthesizing the sialic-acid antigen Neu5Gc, which were replaced by offspring lacking Neu5Gc and producing anti-Neu5Gc antibody. A present-day example for accidental mutations in very few humans that lost a common carbohydrate-antigen and produce a natural antibody against it is the rare blood-group "Bombay" individuals. These individuals lack the H-antigen (blood-group O) which is synthesized in all other humans, and produce the natural anti-H antibody against blood-group O. Overall, it is suggested that natural anti-carbohydrate antibodies played a critical role in preventing complete extinction of mammalian species in epidemics of highly virulent viruses and may have similar role in future events.