Multiplexing of TMT labeling reveals folate-deficient diet-specific proteome changes in NTDs

Abstract

Introduction: In the early stage of embryonic development, the neural tube (NT) cannot be closed properly due to some complex factors, including environmental factors, genetic factors, and the relationship between various factors, leading to the occurrence of neural tube defects (NTDs).Methods: In this study, we induced a mouse model of NTDs by feeding mice with a low-folate diet and intraperitoneally injecting them with 1.5 mg/kg methotrexate on E7.5. Fetal mice were achieved at E13.5, and we extracted proteins from brain tissues with trypsin digestion. After enzymatic digestion, peptides were labeled with TMT/iTRAQ and separated in high-performance liquid chromatography (HPLC) for subsequent liquid chromatography tandem mass spectroscopy (LC-MS/MS) analysis. We used gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation to analyze proteomic changes and analyze the functional enrichment of differentially expressed proteins (DEPs) in the NTD mice tissues.Results: A low-folate-induced mouse model was successfully constructed. Folate was used as a sensitizing agent, and the teratogenicity rate of the NTD fetal mice increased to 36.5% when the concentration of methotrexate was at 1.5 mg/kg. Mass spectrometry was used to identify 6,614 proteins, and among them, 5,656 proteins were quantified. In the following proteomic analysis, GO classification and KEGG pathway enrichment analysis were conducted, and heatmaps were drawn for differentially expressed proteins (DEPs). The main pathways associated with NTDs, such as the Hedgehog, Wnt, p53, and Hippo signaling pathways and the one-carbon pool mediated by folate, can be identified through a protein–protein interaction (PPI) network. It was also found that the regulation of ribosomal proteins, such as RPL13 and RPL14, which are upregulated in NTDs, has a certain impact on neural tube development.Discussion: Our results revealed proteomic changes in the tissues of low-folate-induced NTD mice. Validation showed that ribosomal proteins play a regulatory role during the development of NTDs and provides new ideas for the pathogenesis and preventive measures of NTDs.

Umami Altering Salivary Proteome: A Study across a Sensitivity Spectrum on Subjects

“…Enhanced interactions between ALDH2 and ADH1C improve the group’s ability to metabolize nitrogenous waste and manage oxidative stress, crucial for countering the oxidative challenges associated with high MSG intake . The downregulation of the ribosomal protein RPL14 and its interactions indicates a strategic modulation of protein synthesis, likely aimed at conserving energy in response to elevated MSG levels. Additionally, adaptive changes in the biosynthesis of organic and carboxylic acids may alter membrane composition or signaling mechanisms essential for umami taste transduction .…”

“…The active roles of proteins such as PKLR and RPL14, through their synergistic interactions, are critical in optimizing protein synthesis in response to increased glucose metabolism and vital for sustaining a high turnover of proteins necessary for robust sensory and metabolic responses to umami . Furthermore, the interaction between RPL14 and EEF1D accentuates the importance of translation elongation factors, ensuring that the protein synthesis machinery is finely tuned to meet the metabolic demands imposed by MSG exposure …”

“…42 Furthermore, the interaction between RPL14 and EEF1D accentuates the importance of translation elongation factors, ensuring that the protein synthesis machinery is finely tuned to meet the metabolic demands imposed by MSG exposure. 35 In the LMS group, the adaptation strategy is characterized by more conservative metabolic regulation, especially under lower MSG stimuli. This is evidenced by the downregulation of key proteins such as ALDH2 with HADHA, alongside a noticeable decrease in protein synthesis and mitochondrial respiratory chain functions, suggesting a strategic recalibration of cellular resources in response to lesser demands of umami components.…”