RESUMO
Epitranscriptomic mechanisms linking tRNA function and the brain proteome to cognition and complex behaviors are not well described. Here, we report bi-directional changes in depression-related behaviors after genetic disruption of neuronal tRNA cytosine methylation, including conditional ablation and transgene-derived overexpression of Nsun2 in the mouse prefrontal cortex (PFC). Neuronal Nsun2-deficiency was associated with a decrease in tRNA m5C levels, resulting in deficits in expression of 70% of tRNAGly isodecoders. Altogether, 1488/5820 proteins changed upon neuronal Nsun2-deficiency, in conjunction with glycine codon-specific defects in translational efficiencies. Loss of Gly-rich proteins critical for glutamatergic neurotransmission was associated with impaired synaptic signaling at PFC pyramidal neurons and defective contextual fear memory. Changes in the neuronal translatome were also associated with a 146% increase in glycine biosynthesis. These findings highlight the methylation sensitivity of glycinergic tRNAs in the adult PFC. Furthermore, they link synaptic plasticity and complex behaviors to epitranscriptomic modifications of cognate tRNAs and the proteomic homeostasis associated with specific amino acids.
Assuntos
Transtorno Depressivo/fisiopatologia , Epigênese Genética/genética , Metiltransferases/genética , Proteoma/metabolismo , RNA de Transferência/genética , Transmissão Sináptica/genética , Animais , Transtorno Depressivo/genética , Transtorno Depressivo/metabolismo , Perfilação da Expressão Gênica/métodos , Metiltransferases/deficiência , Metiltransferases/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Neurônios/metabolismo , Córtex Pré-Frontal/citologia , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/fisiologia , Proteômica/métodos , RNA de Transferência/metabolismo , Transdução de Sinais/genéticaRESUMO
BACKGROUND: To validate a test for independent assessment of insulin secretion and insulin sensitivity during the same occasion for metabolic studies in clinical practice, i.e. combined glucagon-stimulated C-peptide test and insulin tolerance test (GITT). SUBJECTS AND METHODS: We measured C-peptide response to 0.5 mg of intravenous glucagon followed 30 min later by administration of 0.05 U kg(-1) insulin (insulin tolerance test, ITT). Ten subjects with normal glucose tolerance participated on different days in an ITT, glucagon-C-peptide test, ITT followed by glucagon-C-peptide test and glucagon-C-peptide test followed by ITT to establish whether and how the tests could be combined. The test was then repeated in nine patients with type 2 diabetes to investigate its reproducibility. In 20 subjects with varying degrees of glucose tolerance, the test was compared with the Botnia clamp (an intravenous glucose tolerance test combined with a euglycaemic hyperinsulinemic clamp). RESULTS: When ITT preceded the glucagon test, C-peptide response was blunted. Therefore, we first administered glucagon and then insulin (GITT). The K(ITT) from the GITT was reproducible (CV = 13 %) and correlated strongly with the glucose disposal rate from the Botnia clamp (r = 0.87, r(2) = 0.75, P < 0.001). The C-peptide response to glucagon was reproducible (CV = 13 %). The disposition index, providing a measure of beta-cell function adjusted for insulin sensitivity, calculated from the GITT showed good discrimination between individuals with varying degrees of glucose tolerance. CONCLUSIONS: The GITT provides simple, reproducible and independent estimates of insulin sensitivity and secretion on the same occasion for metabolic studies in individuals with normal and abnormal glucose tolerance.
