Andrea Galmozzi

Identification of adipocyte functional pathways using chemoproteomics methods

Efficient intracellular trafficking of small-molecule metabolites, such as lipids, intermediary metabolites, and enzyme cofactors, is critical to maintain cellular and organismal function. Alterations in metabolite homeostasis are linked to multiple diseases, including developmental, immunological, inflammatory, and metabolic disorders. However, our understanding of how key metabolites are handled within the cell remains limited. Using a platform integrating chemoproteomic tools and cellular assays we intend to reveal intracellular mobilization pathways of metabolites that alter the activity of transcription factors, and thus gene expression. Depending on their subcellular localization, the same metabolites can have very diverse functions. For instance, lipids can enter the mitochondria to produce energy, can be released by cells into the bloodstream, or they can modulate protein functions to trigger specific cellular responses. Therefore, knowing where metabolites accumulate in the cells can guide our understanding of their biological function(s). Signaling metabolites may act as second messengers, allosteric regulators, or as ligands for nuclear receptors (NR), a class of ligand-activated transcription factors that sense environmental signals. Metabolite-sensing NRs, with the exception of endocrine receptors, reside in the nucleus and respond to a broad range of ligands derived from endogenous and exogenous sources, such as diet (vitamins, bile acids) and intermediary metabolism (porphyrins, retinoids, fatty acids). However, the intracellular levels of free endogenous NR ligands are often below the concentration required to induce NR activation, and this is especially true for dietary and intermediate metabolites. This observation hints at the existence of chaperones that bind NR ligands at their site of synthesis or entry into the cell, transport them to the nucleus, and deliver them to NRs (Figure 1).

Such transport mechanisms may have evolved to protect cells from the intrinsic reactivity and cytotoxicity of these metabolites. An example of one such chaperone is PGRMC2, activity of the heme-responsive NR Rev-Erbα. Ligand chaperones with tissue-specific expression can also drive the ultimate effect of metabolites that regulate multiple NRs. For instance, retinoic acid carried by CRABP-II activates RAR, while retinoic acid delivered by FABP5 stimulates PPARβ/δ.

In spite of great advances made in understanding metabolite-protein crosstalk, knowledge of the intracellular trafficking of signaling metabolites remains limited. Elucidation of the molecular pathways that traffic signaling metabolites may reveal fundamental mechanisms in cell biology and provide new avenues to modulate gene expression for therapeutic purposes. To this end, we integrate cutting-edge mass spectrometry, chemical biology, and molecular genetics to uncover intracellular trafficking pathways for signaling metabolites and develop new tools for the scientific community to explore protein-metabolite interactions in live cells.