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MLife Jun 2024The microbial synthesis of sulfonolipids within the human body is likely involved in maintaining human health or causing diseases. However, the enzymes responsible for...
The microbial synthesis of sulfonolipids within the human body is likely involved in maintaining human health or causing diseases. However, the enzymes responsible for their biosynthesis remain largely unknown. In this study, we identified and verified the role of 3-ketocapnine reductase, the third-step enzyme, in the four-step conversion of l-phosphoserine into sulfobacin B both in vivo and in vitro. This finding builds upon our previous research into sulfonolipid biosynthesis, which focused on the vaginal bacterium DSM 16776 and the gut bacterium DSM 17242. Through comprehensive gene mapping, we demonstrate the widespread presence of potential sulfonolipid biosynthetic genes across diverse bacterial species inhabiting various regions of the human body. These findings shed light on the prevalence of sulfonolipid-like metabolites within the human microbiota, suggesting a potential role for these lipid molecules in influencing the intricate biointeractions within the complex microbial ecosystem of the human body.
PubMed: 38948141
DOI: 10.1002/mlf2.12134 -
Nature Jun 2024Insertion sequence (IS) elements are the simplest autonomous transposable elements found in prokaryotic genomes. We recently discovered that IS110 family elements encode...
Insertion sequence (IS) elements are the simplest autonomous transposable elements found in prokaryotic genomes. We recently discovered that IS110 family elements encode a recombinase and a non-coding bridge RNA (bRNA) that confers modular specificity for target DNA and donor DNA through two programmable loops. Here we report the cryo-electron microscopy structures of the IS110 recombinase in complex with its bRNA, target DNA and donor DNA in three different stages of the recombination reaction cycle. The IS110 synaptic complex comprises two recombinase dimers, one of which houses the target-binding loop of the bRNA and binds to target DNA, whereas the other coordinates the bRNA donor-binding loop and donor DNA. We uncovered the formation of a composite RuvC-Tnp active site that spans the two dimers, positioning the catalytic serine residues adjacent to the recombination sites in both target and donor DNA. A comparison of the three structures revealed that (1) the top strands of target and donor DNA are cleaved at the composite active sites to form covalent 5'-phosphoserine intermediates, (2) the cleaved DNA strands are exchanged and religated to create a Holliday junction intermediate, and (3) this intermediate is subsequently resolved by cleavage of the bottom strands. Overall, this study reveals the mechanism by which a bispecific RNA confers target and donor DNA specificity to IS110 recombinases for programmable DNA recombination.
Topics: Catalytic Domain; Cryoelectron Microscopy; DNA; DNA Transposable Elements; Models, Molecular; Nucleic Acid Conformation; Protein Multimerization; Recombinases; Recombination, Genetic; RNA, Untranslated; Substrate Specificity
PubMed: 38926616
DOI: 10.1038/s41586-024-07570-2 -
BioRxiv : the Preprint Server For... Jun 2024The receptor tyrosine kinase EphA2 drives cancer malignancy by facilitating metastasis. EphA2 can be found in different self-assembly states: as a monomer, dimer, and...
The receptor tyrosine kinase EphA2 drives cancer malignancy by facilitating metastasis. EphA2 can be found in different self-assembly states: as a monomer, dimer, and oligomer. However, our understanding remains limited regarding which EphA2 state is responsible for driving pro-metastatic signaling. To address this limitation, we have developed SiMPull-POP, a single-molecule method for accurate quantification of membrane protein self-assembly. Our experiments revealed that a reduction of plasma membrane cholesterol strongly promoted EphA2 self-assembly. Indeed, low cholesterol caused a similar effect to the EphA2 ligand ephrinA1-Fc. These results indicate that cholesterol inhibits EphA2 assembly. Phosphorylation studies in different cell lines revealed that low cholesterol increased phospho-serine levels, the signature of oncogenic signaling. Investigation of the mechanism that cholesterol uses to inhibit the assembly and activity of EphA2 indicate an in-trans effect, where EphA2 is phosphorylated by protein kinase A downstream of beta-adrenergic receptor activity, which cholesterol also inhibits. Our study not only provides new mechanistic insights on EphA2 oncogenic function, but also suggests that cholesterol acts as a molecular safeguard mechanism that prevents uncontrolled self-assembly and activation of EphA2.
PubMed: 38915729
DOI: 10.1101/2024.06.10.598255 -
BioRxiv : the Preprint Server For... Jun 2024Protein post-translational modifications, such as phosphorylation, are important regulatory signals for diverse cellular functions. In particular, intrinsically...
Protein post-translational modifications, such as phosphorylation, are important regulatory signals for diverse cellular functions. In particular, intrinsically disordered protein regions (IDRs) are subject to phosphorylation as a means to modulate their interactions and functions. Toward understanding the relationship between phosphorylation in IDRs and specific functional outcomes, we must consider how phosphorylation affects the IDR conformational ensemble. Various experimental techniques are suited to interrogate the features of IDR ensembles; molecular simulations can provide complementary insights and even illuminate ensemble features that may be experimentally inaccessible. Therefore, we sought to expand the tools available to study phosphorylated IDRs by all-atom Monte Carlo simulations. To this end, we implemented parameters for phosphoserine (pSer) and phosphothreonine (pThr) into the OPLS version of the continuum solvent model, ABSINTH, and assessed their performance in all-atom simulations compared to published findings. We simulated short (< 20 residues) and long (> 80 residues) phospho-IDRs that, collectively, survey both local and global phosphorylation-induced changes to the ensemble. Our simulations of four well-studied phospho-IDRs show near-quantitative agreement with published findings for these systems via metrics including changes to radius of gyration, transient helicity, and persistence length. We also leveraged the inherent advantage of sequence control in molecular simulations to explore the conformational effects of diverse combinations of phospho-sites in two multi-phosphorylated IDRs. Our results support and expand on prior observations that connect phosphorylation to changes in the IDR conformational ensemble. Herein, we describe phosphorylation as a means to alter sequence chemistry, net charge and charge patterning, and intramolecular interactions, which can collectively modulate the local and global IDR ensemble features.
PubMed: 38915510
DOI: 10.1101/2024.06.10.598315 -
MicroPublication Biology 2024Abnormal synaptic aggregation of α-synuclein is linked to cognitive deficits in Parkinson's disease (PD). While the impacts of excess α-synuclein on synaptic function...
Abnormal synaptic aggregation of α-synuclein is linked to cognitive deficits in Parkinson's disease (PD). While the impacts of excess α-synuclein on synaptic function are well established, comparatively less is known about the effects on local mitochondria. Here, we examined morphological features of synaptic mitochondria treated with wild type (WT) or phosphoserine 129 (pS129) α-synuclein, a variant with prominent synaptic accumulation in PD. Acute introduction of pS129 α-synuclein to lamprey synapses caused an activity-dependent swelling and bursting of mitochondria, which did not occur with WT α-synuclein. These pS129-induced effects on mitochondria likely contribute to the synaptic deficits observed in PD.
PubMed: 38854632
DOI: 10.17912/micropub.biology.001206 -
Scientific Reports May 2024The non-essential amino acid L-serine is involved in a number of metabolic pathways and in the brain its level is largely due to the biosynthesis from the glycolytic...
The non-essential amino acid L-serine is involved in a number of metabolic pathways and in the brain its level is largely due to the biosynthesis from the glycolytic intermediate D-3-phosphoglycerate by the phosphorylated pathway (PP). This cytosolic pathway is made by three enzymes proposed to generate a reversible metabolon named the "serinosome". Phosphoserine phosphatase (PSP) catalyses the last and irreversible step, representing the driving force pushing L-serine synthesis. Genetic defects of the PP enzymes result in strong neurological phenotypes. Recently, we identified the homozygous missense variant [NM_004577.4: c.398A > G p.(Asn133Ser)] in the PSPH, the PSP encoding gene, in two siblings with a neurodevelopmental syndrome and a myelopathy. The recombinant Asn133Ser enzyme does not show significant alterations in protein conformation and dimeric oligomerization state, as well as in enzymatic activity and functionality of the reconstructed PP. However, the Asn133Ser variant is less stable than wild-type PSP, a feature also apparent at cellular level. Studies on patients' fibroblasts also highlight a strong decrease in the level of the enzymes of the PP, a partial nuclear and perinuclear localization of variant PSP and a stronger perinuclear aggregates formation. We propose that these alterations contribute to the formation of a dysfunctional serinosome and thus to the observed reduction of L-serine, glycine and D-serine levels (the latter playing a crucial role in modulating NMDA receptors). The characterization of patients harbouring the Asn133Ser PSP substitution allows to go deep into the molecular mechanisms related to L-serine deficit and to suggest treatments to cope with the observed amino acids alterations.
Topics: Humans; Serine; Mutation, Missense; Phosphoric Monoester Hydrolases; Fibroblasts; Male; Neurodevelopmental Disorders; Female
PubMed: 38816452
DOI: 10.1038/s41598-024-63164-y -
Nature Microbiology May 2024Adaptation to hypoxia is a major challenge for the survival of Mycobacterium tuberculosis (Mtb) in vivo. Interferon (IFN)-γ-producing CD8 T cells contribute to control...
Adaptation to hypoxia is a major challenge for the survival of Mycobacterium tuberculosis (Mtb) in vivo. Interferon (IFN)-γ-producing CD8 T cells contribute to control of Mtb infection, in part by promoting antimicrobial activities of macrophages. Whether Mtb counters these responses, particularly during hypoxic conditions, remains unknown. Using metabolomic, proteomic and genetic approaches, here we show that Mtb induced Rv0884c (SerC), an Mtb phosphoserine aminotransferase, to produce D-serine. This activity increased Mtb pathogenesis in mice but did not directly affect intramacrophage Mtb survival. Instead, D-serine inhibited IFN-γ production by CD8 T cells, which indirectly reduced the ability of macrophages to restrict Mtb upon co-culture. Mechanistically, D-serine interacted with WDR24 and inhibited mTORC1 activation in CD8 T cells. This decreased T-bet expression and reduced IFN-γ production by CD8 T cells. Our findings suggest an Mtb evasion mechanism where pathogen metabolic adaptation to hypoxia leads to amino acid-dependent suppression of adaptive anti-TB immunity.
PubMed: 38806671
DOI: 10.1038/s41564-024-01701-1 -
International Journal of Biological... Jun 2024Endogenous phospholipase A (PLA) plays an important role in phospholipids degradation during cured meat products manufacturing. The present study was undertaken to...
Endogenous phospholipase A (PLA) plays an important role in phospholipids degradation during cured meat products manufacturing. The present study was undertaken to reveal more information about the endogenous PLA in muscles and its role in degradation of intramuscular phospholipids. With the catalytic domain of pork calcium-independent PLA (iPLAcd), impacts of physic-chemical factors on the activity were investigated and substrate specificity of the enzyme were tested respectively. The optimum temperature and pH of pork iPLAcd were 40 °C and 7.5, respectively. The iPLAcd could be stimulated by adequate contents of NaCl and ATP, and inhibited by CaCl and NaNO. For native phospholipids, the iPLAcd was of a little higher affinity towards phosphatidylcholine (PC) than phosphatidylethanolamine (PE), phosphoserine (PS) and phosphatidylinositol (PI). The iPLAcd could preferentially hydrolyze peroxidized PC over the native PC. The results would help better understand the degradation of phospholipids and the role played by endogenous enzymes during meat products manufacturing.
Topics: Animals; Hydrolysis; Phosphatidylcholines; Swine; Catalytic Domain; Phospholipases A2; Hydrogen-Ion Concentration; Substrate Specificity; Temperature; Hydrogen Peroxide
PubMed: 38768921
DOI: 10.1016/j.ijbiomac.2024.132516 -
Nature May 2024Plant pattern-recognition receptors perceive microorganism-associated molecular patterns to activate immune signalling. Activation of the pattern-recognition receptor...
Plant pattern-recognition receptors perceive microorganism-associated molecular patterns to activate immune signalling. Activation of the pattern-recognition receptor kinase CERK1 is essential for immunity, but tight inhibition of receptor kinases in the absence of pathogen is crucial to prevent autoimmunity. Here we find that the U-box ubiquitin E3 ligase OsCIE1 acts as a molecular brake to inhibit OsCERK1 in rice. During homeostasis, OsCIE1 ubiquitinates OsCERK1, reducing its kinase activity. In the presence of the microorganism-associated molecular pattern chitin, active OsCERK1 phosphorylates OsCIE1 and blocks its E3 ligase activity, thus releasing the brake and promoting immunity. Phosphorylation of a serine within the U-box of OsCIE1 prevents its interaction with E2 ubiquitin-conjugating enzymes and serves as a phosphorylation switch. This phosphorylation site is conserved in E3 ligases from plants to animals. Our work identifies a ligand-released brake that enables dynamic immune regulation.
Topics: Animals; Chitin; Homeostasis; Ligands; Oryza; Phosphorylation; Plant Immunity; Plant Proteins; Ubiquitin; Ubiquitin-Conjugating Enzymes; Ubiquitin-Protein Ligases; Ubiquitination; Phosphoserine; Conserved Sequence
PubMed: 38750355
DOI: 10.1038/s41586-024-07418-9 -
American Journal of Translational... 2024Breast cancer is the most common cancer and the leading cause of cancer-related death among women. An Estrogen Receptor (ER) antagonist called tamoxifen is used as an...
OBJECTIVES
Breast cancer is the most common cancer and the leading cause of cancer-related death among women. An Estrogen Receptor (ER) antagonist called tamoxifen is used as an adjuvant therapy for ER-positive breast cancers. Approximately 40% of patients develop tamoxifen resistance (TAMR) while receiving treatment. Cancer cells can rewire their metabolism to develop resistant phenotypes, and their metabolic state determines how receptive they are to chemotherapy.
METHODS
Metabolite extraction from human MCF-7 and MCF-7/TAMR cells was done using the methanol-methanol-water extraction method. After treating the dried samples with methoxamine hydrochloride in pyridine, the samples were derivatized with 2,2,2-Trifluoro-N-methyl-N-(trimethylsilyl)-acetamide, and Chlorotrimethylsilane (MSTFA + 1% TMCS). The Gas chromatography/mass spectrometry (GC-MS) raw data were processed using MSdial and Metaboanalyst for analysis.
RESULTS
Univariate analysis revealed that 35 metabolites were elevated in TAMR cells whereas 25 metabolites were downregulated. N-acetyl-D-glucosamine, lysine, uracil, tyrosine, alanine, and o-phosphoserine were upregulated in TAMR cells, while hydroxyproline, glutamine, N-acetyl-L-aspartic acid, threonic acid, pyroglutamic acid, glutamine, o-phosphoethanolamine, oxoglutaric acid, and myoinositol were found to be downregulated. Multivariate analysis revealed a distinct separation between the two cell lines, as evidenced by their metabolite levels. The enriched pathways of deregulated metabolites included valine, leucine, and isoleucine degradation, Citric Acid Cycle, Warburg effect, Malate-Aspartate shuttle, glucose-alanine cycle, propanoate metabolism, and Phospholipid biosynthesis.
CONCLUSION
This study revealed dysregulation of various metabolic processes in TAMR cells, which may be crucial in elucidating the molecular basis of the mechanisms underlying acquired tamoxifen resistance.
PubMed: 38715825
DOI: 10.62347/MJLN5908