Rhizobium etli CFN42 and Sinorhizobium meliloti 1021 bioinformatic transcriptional regulatory networks from culture and symbiosis.

Rhizobium etli CFN42 proteome-transcriptome mixed data of exponential growth and nitrogen-fixing bacteroids, as well as Sinorhizobium meliloti 1021 transcriptome data of growth and nitrogen-fixing bacteroids, were integrated into transcriptional regulatory networks (TRNs). The one-step construction network consisted of a matrix-clustering analysis of matrices of the gene profile and all matrices of the transcription factors (TFs) of their genome. The networks were constructed with the prediction of regulatory network application of the RhizoBindingSites database (http://rhizobindingsites.ccg.unam.mx/). The deduced free-living Rhizobium etli network contained 1,146 genes, including 380 TFs and 12 sigma factors. In addition, the bacteroid R. etli CFN42 network contained 884 genes, where 364 were TFs, and 12 were sigma factors, whereas the deduced free-living Sinorhizobium meliloti 1021 network contained 643 genes, where 259 were TFs and seven were sigma factors, and the bacteroid Sinorhizobium meliloti 1021 network contained 357 genes, where 210 were TFs and six were sigma factors. The similarity of these deduced condition-dependent networks and the biological E. coli and B. subtilis independent condition networks segregates from the random Erdös-Rényi networks. Deduced networks showed a low average clustering coefficient. They were not scale-free, showing a gradually diminishing hierarchy of TFs in contrast to the hierarchy role of the sigma factor rpoD in the E. coli K12 network. For rhizobia networks, partitioning the genome in the chromosome, chromids, and plasmids, where essential genes are distributed, and the symbiotic ability that is mostly coded in plasmids, may alter the structure of these deduced condition-dependent networks. It provides potential TF gen-target relationship data for constructing regulons, which are the basic units of a TRN.

MAYEX is an old long noncoding RNA recruited for X chromosome dosage compensation in a reptile.

Long noncoding RNAs (lncRNAs) are essential regulatory elements of sex chromosomes that act to equalize gene expression levels between males and females. XIST, RSX, and roX2 regulate X chromosomes in placental mammals, marsupials, and Drosophila, respectively. Because the green anole (Anolis carolinensis) shows complete dosage compensation of its X chromosome, we tested whether a lncRNA was involved. We found an ancient lncRNA, MAYEX, that gained male-specific expression more than 89 million years ago. MAYEX evolved a notable association with the acetylated histone 4 lysine 16 (H4K16ac) epigenetic mark and the ability to loop its locus to the totality of the X chromosome to increase expression levels. MAYEX is the first lncRNA in reptiles linked to a dosage compensation mechanism that balances the expression of sex chromosomes.

RhizoBindingSites v2.0 Is a Bioinformatic Database of DNA Motifs Potentially Involved in Transcriptional Regulation Deduced From Their Genomic Sites.

RhizoBindingSites is a de novo depurified database of conserved DNA motifs potentially involved in the transcriptional regulation of the Rhizobium, Sinorhizobium, Bradyrhizobium, Azorhizobium, and Mesorhizobium genera covering 9 representative symbiotic species, deduced from the upstream regulatory sequences of orthologous genes (O-matrices) from the Rhizobiales taxon. The sites collected with O-matrices per gene per genome from RhizoBindingSites were used to deduce matrices using the dyad-Regulatory Sequence Analysis Tool (RSAT) method, giving rise to novel S-matrices for the construction of the RizoBindingSites v2.0 database. A comparison of the S-matrix logos showed a greater frequency and/or re-definition of specific-position nucleotides found in the O-matrices. Moreover, S-matrices were better at detecting genes in the genome, and there was a more significant number of transcription factors (TFs) in the vicinity than O-matrices, corresponding to a more significant genomic coverage for S-matrices. O-matrices of 3187 TFs and S-matrices of 2754 TFs from 9 species were deposited in RhizoBindingSites and RhizoBindingSites v2.0, respectively. The homology between the matrices of TFs from a genome showed inter-regulation between the clustered TFs. In addition, matrices of AraC, ArsR, GntR, and LysR ortholog TFs showed different motifs, suggesting distinct regulation. Benchmarking showed 72%, 68%, and 81% of common genes per regulon for O-matrices and approximately 14% less common genes with S-matrices of Rhizobium etli CFN42, Rhizobium leguminosarum bv. viciae 3841, and Sinorhizobium meliloti 1021. These data were deposited in RhizoBindingSites and the RhizoBindingSites v2.0 database (http://rhizobindingsites.ccg.unam.mx/).

The role of HR-HPV integration in the progression of premalignant lesions into different cancer types.

High-risk human papillomavirus (HR-HPV) is associated with the development of different types of cancer, such as cervical, head and neck (including oral, laryngeal, and oropharyngeal), vulvar, vaginal, penile, and anal cancers. The progression of premalignant lesions to cancer depends on factors associated with the host cell and the different epithelia infected by HPV, such as basal cells of the flat epithelium and the cells of the squamocolumnar transformation zone (STZ) found in the uterine cervix and the anal canal, which is rich in heparan sulfate proteoglycans and integrin-like receptors. On the other hand, factors associated with the viral genotype, infection with multiple viruses, viral load, viral persistence, and type of integration determine the viral breakage pattern and the sites at which the virus integrates into the host cell genome (introns, exons, intergenic regions), inducing the loss of function of tumor suppressor genes and increasing oncogene expression. This review describes the role of viral integration and the molecular mechanisms induced by HR-HPV in different types of tissues. The purpose of this review is to identify the common factors associated with the role of integration events in the progression of premalignant lesions in different types of cancer.

Bacterial protein acetylation: mechanisms, functions, and methods for study.

Lysine acetylation is an evolutionarily conserved protein modification that changes protein functions and plays an essential role in many cellular processes, such as central metabolism, transcriptional regulation, chemotaxis, and pathogen virulence. It can alter DNA binding, enzymatic activity, protein-protein interactions, protein stability, or protein localization. In prokaryotes, lysine acetylation occurs non-enzymatically and by the action of lysine acetyltransferases (KAT). In enzymatic acetylation, KAT transfers the acetyl group from acetyl-CoA (AcCoA) to the lysine side chain. In contrast, acetyl phosphate (AcP) is the acetyl donor of chemical acetylation. Regardless of the acetylation type, the removal of acetyl groups from acetyl lysines occurs only enzymatically by lysine deacetylases (KDAC). KATs are grouped into three main superfamilies based on their catalytic domain sequences and biochemical characteristics of catalysis. Specifically, members of the GNAT are found in eukaryotes and prokaryotes and have a core structural domain architecture. These enzymes can acetylate small molecules, metabolites, peptides, and proteins. This review presents current knowledge of acetylation mechanisms and functional implications in bacterial metabolism, pathogenicity, stress response, translation, and the emerging topic of protein acetylation in the gut microbiome. Additionally, the methods used to elucidate the biological significance of acetylation in bacteria, such as relative quantification and stoichiometry quantification, and the genetic code expansion tool (CGE), are reviewed.

Bioinformatics Analysis of Human Papillomavirus 16 Integration in Cervical Cancer: Changes in MAGI-1 Expression in Premalignant Lesions and Invasive Carcinoma.

HPV 16 integration is crucial for the onset and progression of premalignant lesions to invasive squamous cell carcinoma (ISCC) because it promotes the amplification of proto-oncogenes and the silencing of tumor suppressor genes; some of these are proteins with PDZ domains involved in homeostasis and cell polarity. Through a bioinformatics approach based on interaction networks, a group of proteins associated with HPV 16 infection, PDZ domains, and direct physical interaction with E6 and related to different hallmarks of cancer were identified. MAGI-1 was selected to evaluate the expression profile and subcellular localization changes in premalignant lesions and ISCC with HPV 16 in an integrated state in cervical cytology; the profile expression of MAGI-1 diminished according to lesion grade. Surprisingly, in cell lines CaSki and SiHa, the protein localization was cytoplasmic and nuclear. In contrast, in histological samples, a change in subcellular localization from the cytoplasm in low-grade squamous intraepithelial lesions (LSIL) to the nucleus in the high-grade squamous intraepithelial lesion (HSIL) was observed; in in situ carcinomas and ISCC, MAGI-1 expression was absent. In conclusion, MAGI-1 expression could be a potential biomarker for distinguishing those cells with normal morphology but with HPV 16 integrated from those showing morphology-related uterine cervical lesions associated with tumor progression.

Dynamics of Mitochondrial Proteome and Acetylome in Glioblastoma Cells with Contrasting Metabolic Phenotypes.

Glioblastoma, a type of cancer affecting the central nervous system, is characterized by its poor prognosis and the dynamic alteration of its metabolic phenotype to fuel development and progression. Critical to cellular metabolism, mitochondria play a pivotal role, where the acetylation of lysine residues on mitochondrial enzymes emerges as a crucial regulatory mechanism of protein function. This post-translational modification, which negatively impacts the mitochondrial proteome's functionality, is modulated by the enzyme sirtuin 3 (SIRT3). Aiming to elucidate the regulatory role of SIRT3 in mitochondrial metabolism within glioblastoma, we employed high-resolution mass spectrometry to analyze the proteome and acetylome of two glioblastoma cell lines, each exhibiting distinct metabolic behaviors, following the chemical inhibition of SIRT3. Our findings reveal that the protein synthesis machinery, regulated by lysine acetylation, significantly influences the metabolic phenotype of these cells. Moreover, we have shed light on potential novel SIRT3 targets, thereby unveiling new avenues for future investigations. This research highlights the critical function of SIRT3 in mitochondrial metabolism and its broader implications for cellular energetics. It also provides a comparative analysis of the proteome and acetylome across glioblastoma cell lines with opposing metabolic phenotypes.

High variability of perezone content in rhizomes of Acourtia cordata wild plants, environmental factors related, and proteomic analysis.

With the aim of exploring the source of the high variability observed in the production of perezone, in Acourtia cordata wild plants, we analyze the influence of soil parameters and phenotypic characteristics on its perezone content. Perezone is a sesquiterpene quinone responsible for several pharmacological effects and the A. cordata plants are the natural source of this metabolite. The chemistry of perezone has been widely studied, however, no studies exist related to its production under natural conditions, nor to its biosynthesis and the environmental factors that affect the yield of this compound in wild plants. We also used a proteomic approach to detect differentially expressed proteins in wild plant rhizomes and compare the profiles of high vs. low perezone-producing plants. Our results show that in perezone-producing rhizomes, the presence of high concentrations of this compound could result from a positive response to the effects of some edaphic factors, such as total phosphorus (Pt), total nitrogen (Nt), ammonium (NH4), and organic matter (O. M.), but could also be due to a negative response to the soil pH value. Additionally, we identified 616 differentially expressed proteins between high and low perezone producers. According to the functional annotation of this comparison, the upregulated proteins were grouped in valine biosynthesis, breakdown of leucine and isoleucine, and secondary metabolism such as terpenoid biosynthesis. Downregulated proteins were grouped in basal metabolism processes, such as pyruvate and purine metabolism and glycolysis/gluconeogenesis. Our results suggest that soil parameters can impact the content of perezone in wild plants. Furthermore, we used proteomic resources to obtain data on the pathways expressed when A. cordata plants produce high and low concentrations of perezone. These data may be useful to further explore the possible relationship between perezone production and abiotic or biotic factors and the molecular mechanisms related to high and low perezone production.

The transmembrane proteins (TMEM) and their role in cell proliferation, migration, invasion, and epithelial-mesenchymal transition in cancer.

Transmembrane proteins (TMEM) are located in the different biological membranes of the cell and have at least one passage through these cellular compartments. TMEM proteins carry out a wide variety of functions necessary to maintain cell homeostasis TMEM165 participates in glycosylation protein, TMEM88 in the development of cardiomyocytes, TMEM45A in epidermal keratinization, and TMEM74 regulating autophagy. However, for many TMEM proteins, their physiological function remains unknown. The role of these proteins is being recently investigated in cancer since transcriptomic and proteomic studies have revealed that exits differential expression of TMEM proteins in different neoplasms concerning cancer-free tissues. Among the cellular processes in which TMEM proteins have been involved in cancer are the promotion or suppression of cell proliferation, epithelial-mesenchymal transition, invasion, migration, intravasation/extravasation, metastasis, modulation of the immune response, and response to antineoplastic drugs. Inclusive data suggests that the participation of TMEM proteins in these cellular events could be carried out through involvement in different cell signaling pathways. However, the exact mechanisms not clear. This review shows a description of the involvement of TMEM proteins that promote or decrease cell proliferation, migration, and invasion in cancer cells, describes those TMEM proteins for which both a tumor suppressor and a tumor promoter role have been identified, depending on the type of cancer in which the protein is expressed. As well as some TMEM proteins involved in chemoresistance. A better characterization of these proteins is required to improve the understanding of the tumors in which their expression and function are altered; in addition to improving the understanding of the role of these proteins in cancer will show those TMEM proteins be potential candidates as biomarkers of response to chemotherapy or prognostic biomarkers or as potential therapeutic targets in cancer.

Differential Protein Expression of Taenia crassiceps ORF Strain in the Murine Cysticercosis Model Using Resistant (C57BL/6) Mice.

A cysticercosis model of Taenia crassiceps ORF strain in susceptible BALB/c mice revealed a Th2 response after 4 weeks, allowing for the growth of the parasite, whereas resistant C57BL/6 mice developed a sustained Th1 response, limiting parasitic growth. However, little is known about how cysticerci respond to an immunological environment in resistant mice. Here, we show that the Th1 response, during infection in resistant C57BL/6 mice, lasted up to 8 weeks and kept parasitemia low. Proteomics analysis of parasites during this Th1 environment showed an average of 128 expressed proteins; we chose 15 proteins whose differential expression varied between 70 and 100%. A total of 11 proteins were identified that formed a group whose expression increased at 4 weeks and decreased at 8 weeks, and another group with proteins whose expression was high at 2 weeks and decreased at 8 weeks. These identified proteins participate in tissue repair, immunoregulation and parasite establishment. This suggests that T. crassiceps cysticerci in mice resistant under the Th1 environment express proteins that control damage and help to establish a parasite in the host. These proteins could be targets for drugs or vaccine development.

Rhizobium etli CFN42 proteomes showed isoenzymes in free-living and symbiosis with a different transcriptional regulation inferred from a transcriptional regulatory network.

A comparative proteomic study at 6 h of growth in minimal medium (MM) and bacteroids at 18 days of symbiosis of Rhizobium etli CFN42 with the Phaseolus vulgaris leguminous plant was performed. A gene ontology classification of proteins in MM and bacteroid, showed 31 and 10 pathways with higher or equal than 30 and 20% of proteins with respect to genome content per pathway, respectively. These pathways were for energy and environmental compound metabolism, contributing to understand how Rhizobium is adapted to the different conditions. Metabolic maps based on orthology of the protein profiles, showed 101 and 74 functional homologous proteins in the MM and bacteroid profiles, respectively, which were grouped in 34 different isoenzymes showing a great impact in metabolism by covering 60 metabolic pathways in MM and symbiosis. Taking advantage of co-expression of transcriptional regulators (TF's) in the profiles, by selection of genes whose matrices were clustered with matrices of TF's, Transcriptional Regulatory networks (TRN´s) were deduced by the first time for these metabolic stages. In these clustered TF-MM and clustered TF-bacteroid networks, containing 654 and 246 proteins, including 93 and 46 TFs, respectively, showing valuable information of the TF's and their regulated genes with high stringency. Isoenzymes were specific for adaptation to the different conditions and a different transcriptional regulation for MM and bacteroid was deduced. The parameters of the TRNs of these expected biological networks and biological networks of E. coli and B. subtilis segregate from the random theoretical networks. These are useful data to design experiments on TF gene-target relationships for bases to construct a TRN.

Proteomic analysis of Burkholderia zhejiangensis CEIB S4-3 during the methyl parathion degradation process.

Methyl parathion is an organophosphorus pesticide widely employed worldwide to control pests in agricultural and domestic environments. However, due to its intensive use, high toxicity, and environmental persistence, methyl parathion is recognized as an important ecosystem and human health threat, causing severe environmental pollution events and numerous human poisoning and deaths each year. Therefore, identifying and characterizing microorganisms capable of fully degrading methyl parathion and its degradation metabolites is a crucial environmental task for the bioremediation of pesticide-polluted sites. Burkholderia zhejiangensis CEIB S4-3 is a bacterial strain isolated from agricultural soils capable of immediately hydrolyzing methyl parathion at a concentration of 50 mg/L and degrading the 100% of the released p-nitrophenol in a 12-hour lapse when cultured in minimal salt medium. In this study, a comparative proteomic analysis was conducted in the presence and absence of methyl parathion to evaluate the biological mechanisms implicated in the methyl parathion biodegradation and resistance by the strain B. zhejiangensis CEIB S4-3. In each treatment, the changes in the protein expression patterns were evaluated at three sampling times, zero, three, and nine hours through the use of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and the differentially expressed proteins were identified by mass spectrometry (MALDI-TOF). The proteomic analysis allowed the identification of 72 proteins with differential expression, 35 proteins in the absence of the pesticide, and 37 proteins in the experimental condition in the presence of methyl parathion. The identified proteins are involved in different metabolic processes such as the carbohydrate and amino acids metabolism, carbon metabolism and energy production, fatty acids ß-oxidation, and the aromatic compounds catabolism, including enzymes of the both p-nitrophenol degradation pathways (Hydroquinone dioxygenase and Hydroxyquinol 1,2 dioxygenase), as well as the overexpression of proteins implicated in cellular damage defense mechanisms such as the response and protection of the oxidative stress, reactive oxygen species defense, detoxification of xenobiotics, and DNA repair processes. According to these data, B. zhejiangensis CEIB S4-3 overexpress different proteins related to aromatic compounds catabolism and with the p-nitrophenol  degradation pathways, the higher expression levels observed in the two subunits of the enzyme Hydroquinone dioxygenase, suggest a preferential use of the Hydroquinone metabolic pathway in the p-nitrophenol degradation process. Moreover the overexpression of several proteins implicated in the oxidative stress response, xenobiotics detoxification, and DNA damage repair reveals the mechanisms employed by B. zhejiangensis CEIB S4-3 to counteract the adverse effects caused by the methyl parathion and p-nitrophenol exposure.

Specialized acyl carrier protein used by serine palmitoyltransferase to synthesize sphingolipids in Rhodobacteria.

Serine palmitoyltransferase (SPT) catalyzes the first and committed step in sphingolipid biosynthesis condensating L-serine and acyl-CoA to form 3-oxo-sphinganine. Whenever the structural gene for SPT is present in genomes of Rhodobacteria (α-, ß-, and γ-Proteobacteria), it co-occurs with genes coding for a putative acyl carrier protein (ACP) and a putative acyl-CoA synthetase (ACS). In the α-proteobacterium Caulobacter crescentus, CC_1162 encodes an SPT, whereas CC_1163 and CC_1165 encode the putative ACP and ACS, respectively, and all three genes are known to be required for the formation of the sphingolipid intermediate 3-oxo-sphinganine. Here we show that the putative ACP possesses a 4'-phosphopantetheine prosthetic group, is selectively acylated by the putative ACS and therefore is a specialized ACP (AcpR) required for sphingolipid biosynthesis in Rhodobacteria. The putative ACS is unable to acylate coenzyme A or housekeeping ACPs, but acylates specifically AcpR. Therefore, it is a specialized acyl-ACP synthetase (AasR). SPTs from C. crescentus, Escherichia coli B, or Sphingomonas wittichii use preferentially acyl-AcpR as thioester substrate for 3-oxo-sphinganine synthesis. Whereas acyl-AcpR from C. crescentus is a good substrate for SPTs from distinct Rhodobacteria, acylation of a specific AcpR is achieved by the cognate AasR from the same bacterium. Rhodobacteria might use this more complex way of 3-oxo-sphinganine formation in order to direct free fatty acids toward sphingolipid biosynthesis.

Curcumin prevents proteins expression changes of oxidative phosphorylation, cellular stress response, and lipid metabolism proteins in liver of mice fed a high-fructose diet.

Increased fructose consumption has been associated with the development of metabolic diseases due to the modification in protein expression, altering metabolic and signaling pathways. Curcumin is a natural compound with a regulatory effect on genes and metabolic pathways. To identify the fructose-induced protein expression changes and the effect of curcumin on the change of protein expression in the liver of mice fed a standard diet and a high fructose diet, to elucidate the global role of curcumin. Four groups (n = 4/group) of male mice (C57BL6J) of six-weeks-old were formed. One group received a standard diet (C); another received curcumin at 0.75% w/w in the feed (C + C); one more received 30% w/v fructose in drinking water (F); and one group received 30% w/v fructose in drinking water and 0.75% w/w curcumin in food (F + C); for 15 weeks. Proteomic analysis was performed by LC-MS/MS, using the label-free technique with the MaxQuant programs for identification and Perseus for expression change analysis. Differentially expressed proteins (fold change ≥1.5 and p < 0.5) were analyzed by gene ontology and KEGG. A total of 1047 proteins were identified, of which 113 changed their expression in mice fed fructose, compared to the control group, and curcumin modified the expression of 64 proteins in mice fed fructose and curcumin compared to mice that only received fructose. Curcumin prevented the change of expression of 13 proteins involved in oxidative phosphorylation (NDUFB8, NDUFB3, and ATP5L) in the cellular response to stress (PSMA5, HIST1H1D) and lipid metabolism (THRSP, DGAT1, ECI1, and ACOT13). Curcumin in mice fed the standard diet increased the expression of proteins related to oxidative phosphorylation, ribosomes, and PPAR pathways. In addition to fructose, increased expression of proteins involved in oxidative phosphorylation, ribosomes, lipid metabolism, and carbon metabolism. However, curcumin prevented expression change in 13 hepatic proteins of fructose-fed mice involved in oxidative phosphorylation, cellular stress response, and lipid metabolism. SIGNIFICANCE: Curcumin is a natural compound with a regulatory effect on proteins and metabolic pathways. So, curcumin prevents the change of expression in 13 hepatic proteins of fructose-fed mice involved in oxidative phosphorylation, cellular stress response and lipid metabolism, as a supplement with protector activity on fructose-induced toxic effects.

Curcumin Prevents the Glycation of Tricarboxylic Acid Cycle and Cell Respiration Proteins in the Heart of Mice Fed with a High-fructose Diet.

BACKGROUND: A high fructose diet (HFD) induces protein glycation. The latter is related to a higher risk of cardiovascular disease. Curcumin is a natural pleiotropic compound that may possess antiglycant properties. OBJECTIVE: The study aims to analyze the effect of curcumin on the content of glycated proteins in the hearts of 6-week-old mice fed with a HFD for 15 weeks. METHODS: Mice were allocated into four groups (n = 6/group): a control group that received a standard diet (CT); a group that received 30% w/v fructose in water (F); a group that received 0.75% w/w curcumin supplemented in food (C); a group that received 30% w/v fructose in water and 0.75% w/w curcumin supplemented in food (F+C). The content of glycated proteins in the heart was determined by Western Blot (whereas the spots were detected by 2D-PAGE) using anti-AGE and anti-CML antibodies. Densitometric analysis was performed using the ImageLab software. Glycated proteins were identified by MALDI-TOF-MS, and an ontological analysis was performed in terms of biological processes and molecular function based on the STRING and DAVID databases. RESULTS: Fourteen glycated protein spots were detected, two of them with anti-AGE and the other 12 with anti- CML. In total, eleven glycated proteins were identified, out of which three had decreased glycation levels due to curcumin exposure. The identified proteins participate in processes such as cellular respiration, oxidative phosphorylation, lipid metabolism, carbohydrate metabolism, the tricarboxylic acid cycle (TAC), and the organization of intermediate filaments. CONCLUSION: Curcumin decreased the fructose-induced glycation level of the ACO2, NDUFS7, and DLAT proteins.

Quantitative Proteomic Analysis of Cervical Cancer Tissues Identifies Proteins Associated With Cancer Progression.

BACKGROUND/AIM: To date, several proteomics studies in cervical cancer (CC) have focused mainly on squamous cervical cancer (SCC). Our study aimed to discover and clarify differences in SCC and CAD that may provide valuable information for the identification of proteins involved in tumor progression, in CC as a whole, or specific for SCC or CAD. MATERIALS AND METHODS: Total protein extracts from 15 individual samples corresponding to 5 different CC tissue types were compared with a non-cancerous control group using bidimensional liquid chromatography-mass spectrometry (2D LC-MS/MS), isobaric tags for relative and absolute quantitation (ITRAQ), principal component analysis (PCA) and gene set enrichment analysis (GSEA). RESULTS: A total of 622 statistically significant different proteins were detected. Exocytosis-related proteins were the most over-represented, accounting for 25% of the identified and quantified proteins. Based on the experimental results, reticulocalbin 3 (RCN3) and Ras-related protein Rab-14 (RAB14) were chosen for further downstream in vitro and vivo analyses. RCN3 was overexpressed in all CC tissues compared to the control and RAB14 was overexpressed in squamous cervical cancer (SCC) compared to invasive cervical adenocarcinoma (CAD). In the tumor xenograft experiment, RAB14 protein expression was positively correlated with increased tumor size. In addition, RCN3-expressing HeLa cells induced a discrete size increment compared to control, at day 47 after inoculation. CONCLUSION: RAB14 and RCN3 are suggested as potential biomarkers and therapeutic targets in the treatment of CC.

Proteomic analysis reveals the metabolic versatility of Amycolatopsis sp. BX17: A strain native from milpa agroecosystem soil.

Amycolatopsis sp. BX17 is an actinobacterium isolated from milpa soils, which antagonizes the phytopathogenic fungus Fusarium graminearum. Metabolites secreted by the actinobacterium cultured in glucose-free medium inhibited 100% of the mycelial growth of F. graminearum RH1, while the inhibition rate was 65% in medium supplemented with 20 g/L glucose. With the aim of studying how the metabolism of strain BX17 is modulated by glucose as the main carbon source, media with 0 and 20 g/L glucose were selected to analyze the intracellular proteins by quantitative label-free proteomic analysis. Data are available via ProteomeXchange with identifier PXD028644. Proteins identified in bacteria cultured in medium without glucose were involved in glutamate metabolism, the Krebs cycle and the shikimate pathway, suggesting that amino acids are metabolized to synthesize antifungal compounds. In glucose-containing medium, carbon flux was directed mainly toward the synthesis of energy and cell growth. This study shows the metabolic versatility of Amycolatopsis BX17, and strengthens its potential use in designing biotechnological strategies for phytopathogen control. SIGNIFICANCE: Amycolatopsis BX17 is a bacterium isolated from milpa agroecosystems that antagonizes the phytopathogenic fungus Fusarium graminearum. Currently, there is scarce information about the metabolism involved in the biosynthesis of antifungal agents by this genus. We used a label-free proteomic approach to identify the differences in metabolic routes for antifungal biosynthesis in Amycolatopsis BX17 grown in media with 0 and 20 g/L glucose. Taken together the results suggest that the BX17 strain could be synthesizing the antifungal metabolite(s) from the Shikimate pathway through the synthesis and degradation of the amino acid tyrosine, which is a known precursor of glycopeptides with antibiotic and antifungal activity. While the lower antifungal activity of the metabolites secreted by Amycolatopsis BX17 when grown in a medium with glucose as the main carbon source, may be correlated with a lower synthesis of antifungal compounds, due to the directing of carbon flux toward metabolic pathways involved with energy synthesis and cell growth. Likewise, it is possible that the bacteria synthesize other compounds with biological activity, such as glycopeptides with antibiotic activity. These findings are relevant because they represent the first stage to understand the metabolic regulation involved in the biosynthesis of antifungal metabolites by the genus Amycolatopsis. Finally, improving our understanding of the metabolic regulation involved in the biosynthesis of antifungal metabolites is essential to design of strategies in agricultural biotechnology for phytopathogen control.

Lysine Acetylation Stoichiometry Analysis at the Proteome Level.

Lysine acetylation is a widespread posttranslational modification (PTM) in all kingdoms of live. A large number of proteins involved in most of biological pathways are targets of this PTM. The lysine acetylation is a reversible modification controlled by two main groups of enzymes, lysine acetyltransferases responsible for transferring the acetyl group of acetylCoA to the side chain of lysine residues and lysine deacetylases which effectively remove the acetyl tag. Dysregulation of enzymes that control acetylation and/or target proteins have been associated with a growing number of human pathologies. Lysine acetylation is largely a modification that occurs at low stoichiometry at its target sites. Here we describe a method to identify lysine acetylation sites and estimate their site occupancy at the proteome scale. The method relies on a high-resolution mass spectrometry-based proteomics approach, which includes a specific chemical acetylation reaction on unmodified lysine residues that carry heavy isotopes. The procedures described here have been applied to cell line cultures and to clinically relevant samples stored as both snap-frozen and formalin-fixed paraffin-embedded (FFPE) tissues.

Protein lysine acetylation and its role in different human pathologies: a proteomic approach.

INTRODUCTION: Lysine acetylation is a reversible post-translational modification (PTM) regulated through the action of specific types of enzymes: lysine acetyltransferases (KATs) and lysine deacetylases (HDACs), in addition to bromodomains, which are a group of conserved domains which identify acetylated lysine residues, several of the players in the process of protein acetylation, including enzymes and bromodomain-containing proteins, have been related to the progression of several diseases. The combination of high-resolution mass spectrometry-based proteomics, and immunoprecipitation to enrich acetylated peptides has contributed in recent years to expand the knowledge about this PTM described initially in histones and nuclear proteins, and is currently reported in more than 5000 human proteins, that are regulated by this PTM. AREAS COVERED: This review presents an overview of the main participant elements, the scenario in the development of protein lysine acetylation, and its role in different human pathologies. EXPERT OPINION: Acetylation targets are practically all cellular processes in eukaryotes and prokaryotes organisms. Consequently, this modification has been linked to many pathologies like cancer, viral infection, obesity, diabetes, cardiovascular, and nervous system-associated diseases, to mention a few relevant examples. Accordingly, some intermediate mediators in the acetylation process have been projected as therapeutic targets.

Phenotypic plasticity and a new small molecule are involved in a fungal-bacterial interaction.

Nitrogen-fixing bacteria have been extensively studied in the context of interactions with their host plants; however, little is known about the phenotypic plasticity of these microorganisms in nonmutualistic interactions with other eukaryotes. A dual-species coculture model was developed by using the plant symbiotic bacterium Rhizobium etli and the well-studied eukaryote Saccharomyces cerevisiae as a tractable system to explore the molecular mechanisms used by R. etli in nonmutual interactions. Here, we show that the fungus promotes the growth of the bacterium and that together, these organisms form a mixed biofilm whose biomass is ~ 3 times greater and is more structured than that of either single-species biofilm. We found that these biofilm traits are dependent on a symbiotic plasmid encoding elements involved in the phenotypic plasticity of the bacterium, mitochondrial function and in the production of a yeast-secreted sophoroside. Interestingly, the promoters of 3 genes that are key in plant bacteria-interaction (nifH, fixA and nodA) were induced when R. etli coexists with yeast. These results show that investigating interactions between species that do not naturally coexist is a new approach to discover gene functions and specialized metabolites in model organisms.