The publication of the complete human genome sequence in 2004 has marked the beginning of a new era. Since then, the famous Crick dogma that all genes generate active factors, known as proteins, through an intermediate known as mRNA, has led to the categorization of the mRNAs expressed in virtually all cells of the human body. The enormous effort was facilitated by the fact that the analysis of mRNAs is technologically simple, whereas the analysis of proteins is not.
One might, however, ask, “Can protein amounts be deduced by mRNA concentration measurements?” The answer is now “No,” because mRNA levels by themselves are not sufficient to predict protein levels in many scenarios. In short, the presence of an mRNA is necessary but not sufficient for the expression of the respective protein. Indeed, it has been recently demonstrated that there is a rather poor correlation between mRNA and protein levels (Kim et al. 2014). This fact implies the presence of post-transcriptional and translational control mechanisms of protein-abundance regulation.
The lack of correlation between mRNA and proteins is especially important during highly dynamic phases, such as cellular differentiation or cellular responses to stimuli where post-transcriptional processes lead to stronger deviations from an ideal correlation. In these cases, the translation of preexisting mRNA transcripts can help to quickly synthesize a newly required protein. Intriguingly, 60 years ago, Wilson suggested that “cytoplasmic components” in the egg dictate the rapid fate of newly formed embryonic cells. Today, we know that the cytoplasmic components referred by Wilson are preexisting molecules of messenger RNAs (mRNAs) lacking the corresponding proteins. These mRNAs are ready to be translated into proteins in order to direct the rapid synthesis and assembly of cellular components required to sustain embryos during cell division.
T cells represent an outstanding example of highly dynamic cells. Naïve T cells circulate in the body in a resting state, but upon recognition of foreign antigens (antigen engagement) and receipt of proper costimulatory signals, they become promptly activated. They undergo a rapid proliferation, develop into different T cell types and, in doing so, they fight infections. Intimately integrated into this program of T cell activation is the awakening of cellular metabolism. The general notion is that naïve T cells start from a metabolic quiescent phenotype and upon T cell receptor (TCR) activation undergo a radical shift in their metabolic profile, simplified by a switch from oxidative phosphorylation (OXPHOS) to aerobic glycolysis. Among all the signals that mediate T cell metabolic changes following TCR activation, the most critical converges on the mammalian target of rapamycin (mTOR), an evolutionary conserved serine/threonine kinase that acts as a sensor of environmental cues, such as nutrient levels, energy status, and growth factors.
According to the current view, the activation of antigen-engaged naive T cells requires a rapid mTORC1-dependent readjustment of the metabolic machinery towards glycolysis, i.e. the breakage of glucose to generate the intermediates required for energy production and proliferation. The effect of mTORC activation on the metabolic transition towards glycolysis is most likely mediated by the induction of Myc, a transcription factor that regulates cellular gene expression through binding to specific target genes in the nucleus. In T cells, Myc activates the transcription of mRNAs encoding for glycolytic enzymes, thereby diverting glucose away from oxidative phosphorylation and towards glycolysis. In short, the model predicts that naïve T cells are “awakened” through a cascade of events including mTORC1 activation, which is followed by Myc-mediated transcription of mRNAs that encode for glycolytic and fatty acid synthesis enzymes, which are finally translated into proteins.
However, several facts remain unexplained. For instance, the metabolic changes of T cells occur quickly in response to environmental cues and involve transition states that could not be fully explained by a linear flow of events that funnels into transcription and precedes translation. Rather, and more plausibly, post-transcriptional control mechanisms are also at stake. The main aim of our recent work was to investigate if this could be the case. In short, we discovered that, during their quiescence, naïve T cells accumulate specific mRNAs that can be immediately translated.
In particular, in our study, we devised a novel approach to identify the mechanistic network coordinating such metabolic switch, which consisted in combining a comprehensive analysis of publicly available transcriptomics and proteomics datasets of human lymphocytes subsets with a mass spectrometry analysis (MS) of metabolites. We studied the expression of ribosomal, translation and metabolic genes in human primary lymphocyte subtypes isolated from the blood of healthy donors and validated the results using T cells activated in vitro and advanced protein synthesis measurement techniques. A coordinated analysis of the ribosomal and translational machinery showed that CD4+ naïve T cells have an unexpected high translational capability, poised at the pre-initiation step.
In spite of quiescence, naïve cells accumulate mRNAs encoding for anabolic rate-limiting enzymes required for metabolic reprogramming. Strikingly, the mRNAs are there but the proteins not. Upon activation, prompt protein synthesis of the glucose transporter GLUT1 from the corresponding preaccumulated mRNA ignites metabolic awakening and reboots the naïve state. Thanks to GLUT1, glucose can flow into T cells and “awake” them. Next, translational activation of Acetyl-CoA carboxylase ACC1 sustains a metabolic feedforward loop that completes the metabolic reprogramming to an effector phenotype. Altogether, the results of our study suggest for the first time that translational control dictates the metabolic status of quiescence of CD4+ T lymphocytes and regulates naïve T cell exit from their dormant state.
Considering that the whole field of immunology is dominated by a “transcriptocentric” view, the translational view we disclosed offers new perspectives in the field. For instance, capturing interindividual differences in the translated genome will provide new insights into the genes and regulatory pathways underlying human diseases, such as cancer. Thus, translation factors can be considered as good targets for drug development efforts. Indeed, it is well known that the efficacy of antitumor responses by tumor-infiltrating T lymphocytes (TILs) is limited by their progressive loss of function in the tumor microenvironment due to inability to regulate metabolism.
These findings are described in the article entitled The Translational Machinery of Human CD4+ T Cells Is Poised for Activation and Controls the Switch from Quiescence to Metabolic Remodeling, recently published in the journal Cell Metabolism. This work was conducted by Sara Ricciardi, Nicola Manfrini, Roberta Alfieri, Piera Calamita, Maria Cristina Crosti, Simone Gallo, Rolf Müller, Massimiliano Pagani, Sergio Abrignani and Stefano Biffo from the National Institute of Molecular Genetics, Milan, Italy.
- Kim, M.S. et al. A draft map of the human proteome. Nature 509, 575-581 (2014)
- Wilson, E.B. The Cell in Development and Heredity. Macmillan, New York (1925)