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Traditional methodologies often fall short in addressing the complexity of biological systems. In this regard, system biology omics have brought invaluable tools for conducting comprehensive analysis. Current sequencing capabilities have revolutionized genetics and genomics studies, as well as the characterization of transcriptional profiling and dynamics of several species and sample types. Biological systems experience complex biochemical processes involving thousands of molecules. These processes occur at different levels that can be studied using mass spectrometry-based (MS-based) analysis, enabling high-throughput proteomics, glycoproteomics, glycomics, metabolomics, and lipidomics analysis. Here, we present the most up-to-date techniques utilized in the completion of omics analysis. Additionally, we include some interesting examples of the applicability of multi omics to a variety of biological systems.

Glyphosate, the active ingredient in many herbicides, has been widely used in agriculture since the 1970s. Despite initial beliefs in its safety for humans and animals due to the absence of the shikimate pathway, recent studies have raised concerns about its potential health effects. This study aimed to identify glycomic changes indicative of glyphosate-induced toxicity. Specifically, the study focused on profiling N-glycosylation, a protein post-translational modification increasingly recognized for its involvement in various disorders, including neurological conditions. A comprehensive analysis of rat serum N-glycomics following chronic exposure to glyphosate-based herbicides (GBH) was conducted using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results revealed significant changes in the N-glycan profile, particularly in sialylated and sialofucosylated N-glycans. The analysis of N-glycans across gender subgroups provided insights into gender-specific responses to GBH exposure, with the male rats exhibiting a higher susceptibility to these N-glycan changes compared to females. The validation of significantly altered N-glycans using parallel reaction monitoring (PRM) confirmed their expression patterns. This study provides novel insights into the impact of chronic GBH exposure on serum N-glycan composition, with implications for assessing glyphosate toxicity and its potential neurological implications.

Abstract Chia (Salvia hispanica) is an emerging crop considered a functional food containing important substances with multiple potential applications. However, the molecular basis of some relevant chia traits, such as seed mucilage and polyphenol content, remains to be discovered. This study generates an improved chromosome-level reference of the chia genome, resolving some highly repetitive regions, describing methylation patterns, and refining genome annotation. Transcriptomic analysis shows that seeds exhibit a unique expression pattern compared to other organs and tissues. Thus, a metabolic and proteomic approach is implemented to study seed composition and seed-produced mucilage. The chia genome exhibits a significant expansion in mucilage synthesis genes (compared to Arabidopsis), and gene network analysis reveals potential regulators controlling seed mucilage production. Rosmarinic acid, a compound with enormous therapeutic potential, was classified as the most abundant polyphenol in seeds, and candidate genes for its complex pathway are described. Overall, this study provides important insights into the molecular basis for the unique characteristics of chia seeds.

Glyphosate-based herbicides (GBHs) have gained extensive popularity in recent decades. For many years, glyphosate has been regarded as harmless or minimally toxic to mammals due to the absence of its primary target, the shikimic acid pathway in humans. Nonetheless, mounting evidence suggests that glyphosate may cause adverse health effects in humans via other mechanisms. In this study, we described the metabolomic changes in the serum of experimental rats exposed to chronic GBH using the highly sensitive LC-MS/MS technique. We investigated the possible relationship between chronic exposure to GBH and neurological disorders. Our findings suggest that chronic exposure to GBH can alter spatial learning memory and the expression of some important metabolites that are linked to neurophysiological disorders in young rats, with the female rats showing higher susceptibility compared to the males. This indicates that female rats are more likely to show early symptoms of the disorder on exposure to chronic GBH compared to male rats. We observed that four important metabolites (paraxanthine, epinephrine, L-(+)-arginine, and D-arginine) showed significant changes and involvement in neurological changes as suggested by ingenuity pathway analysis. In conclusion, our results indicate that chronic exposure to GBH can increase the risk of developing neurological disorders.

Glycoproteomic analysis is always challenging because of low abundance and complex site-specific heterogeneity. Glycoproteins are involved in various biological processes such as cell signaling, adhesion, and cell–cell communication and may serve as potential biomarkers when analyzing different diseases. Here, we investigate glycoproteins in narcolepsy type 1 (NT1) disease, a form of narcolepsy characterized by cataplexy—the sudden onset of muscle paralysis that is typically triggered by intense emotions. In this study, 27 human blood serum samples were analyzed, 16 from NT1 patients and 11 from healthy individuals serving as controls. We quantified hydrophilic interaction liquid chromatography (HILIC)-enriched glycopeptides from low-abundance serum samples of controls and NT1 patients via LC-MS/MS. Twenty-eight unique N-glycopeptides showed significant changes between the two studied groups. The sialylated N-glycopeptide structures LPTQNITFQTESSVAEQEAEFQSPK HexNAc6, Hex3, Neu5Ac2 (derived from the ITIH4 protein) and the structure IVLDPSGSMNIYLVLDGSDSIGASNFTGAK HexNAc5, Hex4, Fuc1 (derived from the CFB protein), with p values of 0.008 and 0.01, respectively, were elevated in NT1 samples compared with controls. In addition, the N-glycopeptide protein sources Ceruloplasmin, Complement factor B, and ITH4 were observed to play an important role in the complement activation and acute-phase response signaling pathways. This may explain the possible association between the biomarkers and pathophysiological effects.

The highly glycosylated S protein plays a vital role in host cell invasion, making it the principal target for vaccine development. Differences in mutations observed on the spike (S) protein of SARS-CoV-2 variants may result in distinct glycosylation patterns, thus influencing immunological evasion, infectivity, and transmissibility. The glycans can mask key epitopes on the S1 protein and alter its structural conformation, allowing the virus to escape the immune system. Therefore, we comprehensively characterize O-glycosylation in eleven variants of SARS-CoV-2 S1 subunits to understand the differences observed in the biology of the variants. In-depth characterization was performed with a double digestion strategy and an efficient LC-MS/MS approach. We observed that O-glycosylation is highly conserved across all variants in the region between the NTD and RBD, whereas other domains and regions exhibit variation in O-glycosylation. Notably, omicron has the highest number of O-glycosylation sites on the S1 subunit. Also, omicron has the highest level of sialylation in the RBD and RBM functional motifs. Our findings may shed light on how differences in O-glycosylation impact viral pathogenicity in variants of SARS-CoV-2 and facilitate the development of a robust vaccine with high protective efficacy against the variants of concern.

The SARS-CoV-2 virus rapidly spread worldwide, threatening public health. Since it emerged, the scientific community has been engaged in the development of effective therapeutics and vaccines. The subunit S1 in the spike protein of SARS-CoV-2 mediates the viral entry into the host and is therefore one of the major research targets. The S1 protein is extensively glycosylated, and there is compelling evidence that glycans protect the virus’ active site from the human defense system. Therefore, investigation of the S1 protein glycome alterations in the different virus variants will provide a view of the glycan evolution and its relationship with the virus pathogenesis. In this study, we explored the N-glycosylation expression of the S1 protein for eleven SARS-CoV-2 variants: five variants of concern (VOC), including alpha, beta, gamma, delta, and omicron, and six variants of interest (VOI), including epsilon, eta, iota, lambda, kappa, and mu. The results showed significant differences in the N-glycome abundance of all variants. The N-glycome of the VOC showed a large increase in the abundance of sialofucosylated glycans, with the greatest abundance in the omicron variant. In contrast, the results showed a large abundance of fucosylated glycans for most of the VOI. Two glycan compositions, GlcNAc4,Hex5,Fuc,NeuAc (4-5-1-1) and GlcNAc6,Hex8,Fuc,NeuAc (6-8-1-1), were the most abundant structures across all variants. We believe that our data will contribute to understanding the S1 protein’s structural differences between SARS-CoV-2 mutations.

Mild cognitive impairment (MCI) is an early stage of memory loss that affects cognitive abilities, such as language or virtual/spatial comprehension. This cognitive decline is mostly observed with the aging of individuals. Recently, MCI has been considered as a prodromal phase of Alzheimer’s disease (AD), with a 10–15% conversion rate. However, the existing diagnostic methods fail to provide precise and well-timed diagnoses, and the pathophysiology of MCI is not fully understood. Alterations of serum N-glycan expression could represent essential contributors to the overall pathophysiology of neurodegenerative diseases and be used as a potential marker to assess MCI diagnosis using non-invasive procedures. Herein, we undertook an LC-MS/MS glycomics approach to determine and characterize potential N-glycan markers in depleted blood serum samples from MCI patients. For the first time, we profiled the isomeric glycome of the low abundant serum glycoproteins extracted from serum samples of control and MCI patients using an LC-MS/MS analytical strategy. Additionally, the MRM validation of the identified data showed five isomeric N-glycans with the ability to discriminate between healthy and MCI patients: the sialylated N-glycans GlcNAc5,Hex6,Neu5Ac3 and GlcNAc6,Hex7,Neu5Ac4 with single AUCs of 0.92 and 0.87, respectively, and a combined AUC of 0.96; and the sialylated-fucosylated N-glycans GlcNAc4,Hex5,Fuc, Neu5Ac, GlcNAc5,Hex6,Fuc, Neu5Ac2, and GlcNAc6,Hex7,Fuc, Neu5Ac3 with single AUCs of 0.94, 0.67, and 0.88, respectively, and a combined AUC of 0.98. According to the ingenuity pathway analysis (IPA) and in line with recent publications, the identified N-glycans may play an important role in neuroinflammation. It is a process that plays a fundamental role in neuroinflammation, an important process in the progression of neurodegenerative diseases.

Currently, surveillance strategies have inadequate performance for cirrhosis and early detection of hepatocellular carcinoma (HCC). The glycosylation of serum haptoglobin has shown to have significant differences between cirrhosis and HCC, thus can be used for diagnosis. We performed a comprehensive liquid chromatography—parallel reaction monitoring—mass spectrometry (LC-PRM-MS) approach, where a targeted parallel reaction monitoring (PRM) strategy was coupled to a powerful LC system, to study the site-specific isomerism of haptoglobin (Hp) extracted from cirrhosis and HCC patients. We found that our strategy was able to identify a large number of isomeric N-glycopeptides, mainly located in the Hp glycosylation site Asn207. Four N-glycopeptides were found to have significant changes in abundance between cirrhosis and HCC samples (p < 0.05). Strategic combinations of the significant N-glycopeptides, either with alpha-fetoprotein (AFP) or themselves, better estimate the areas under the curve (AUC) of their respective receiver operating characteristic (ROC) curves with respect to AFP. The combination of AFP with the isomeric sialylated fucosylated N-glycopeptides Asn207 + 5-6-1-2 and Asn207 + 5-6-1-3, resulted with an AUC value of 0.98, while the AUC value for AFP alone was 0.85. When comparing cirrhosis vs. early HCC, the isomeric N-glycopeptide Asn207 + 5-6-0-1 better estimated AUC with respect to AFP (AUCAFP = 0.81, and AUCAsn207 + 5-6-0-1 = 0.88, respectively).

Cerebrospinal fluid (CSF) contains valuable biological and neurological information. However, its glycomics analysis is hampered due to the low amount of protein in the biofluid, as has been demonstrated by other glycomics studies using a substantial amount of CSF. In this work, we investigated different N-glycan sample preparation approaches to develop a more sensitive method. These methods, one with an increased amount of buffer solution during the N-glycan release step with a lower amount of sample volume and the other with Filter-Aided N-Glycan Separation (FANGS), were compared with recent work to demonstrate their effectiveness. It was demonstrated that an increased amount of buffer solution showed higher intensity in comparison to the previously published method and FANGS. This suggested that digestion efficiency during the N-glycan release step was not in an optimal condition from the previously published method, and that there is a substantial loss of sample with FANGS when preparing N-glycans from CSF.

IntroductionGlyphosate-based herbicides (GBHs) have been shown to have significant neurotoxic effects, affecting both the structure and function of the brain, and potentially contributing to the development of neurodegenerative disorders. Despite the known importance of glycosylation in disease progression, the glycome profile of systems exposed to GBH has not been thoroughly investigated.MethodsIn this study, we conducted a comprehensive glycomic profiling using LC-MS/MS, on the hippocampus and prefrontal cortex (PFC) of juvenile rats exposed to GBH orally, aiming to identify glyco-signature aberrations after herbicide exposure.ResultsWe observed changes in the glycome profile, particularly in fucosylated, high mannose, and sialofucosylated N-glycans, which may be triggered by GBH exposure. Moreover, we found major significant differences in the N-glycan profiles between the GBH-exposed group and the control group when analyzing each gender independently, in contrast to the analysis that included both genders. Notably, gender differences in the behavioral test of object recognition showed a decreased performance in female animals exposed to GBH compared to controls (p 0.05).ConclusionThese findings suggest that glycans may play a role in the neurotoxic effect caused by GBH. The result suggests that gender variation may influence the response to GBH exposure, with potential implications for disease progression and specifically the neurotoxic effects of GBHs. Understanding these gender-specific responses could enhance knowledge of the mechanisms underlying GBH-induced toxicity and its impact on brain health. Overall, our study represents the first detailed analysis of N-glycome profiles in the hippocampus and PFC of rats chronically exposed to GBH. The observed alterations in the expression of N-glycan structures suggest a potential neurotoxic effect associated with chronic GBH exposure, highlighting the importance of further research in this area.