Microtubule methylation by SET domain protein methyltransferases – MICROMETHYL

The alpha-beta-tubulin heterodimer protein is the microtubule building block. The hydrolysis of the GTP molecule bound to the beta subunit that accompanies assembly makes microtubules intrinsically dynamic. This dynamics is controlled in the cell by several families of regulatory proteins, most of them having either a stabilizing or a destabilizing effect. Another level of regulation comes from the variability of the tubulin molecules themselves. Indeed, tubulin is the product of the expression of different isotypes; it also undergoes post-translational modifications. By analogy with the epigenetic histone code, these molecular patterns define the “tubulin code”. The objective of the MICROMETHYL project is to characterize a type of tubulin modifications that has been discovered only recently, the methylation of lysine residues.

Post-translational modifications of macromolecules depend on “writer” and “eraser” enzymes, which respectively add and remove modifications. These chemical marks are then recognized by “reader” proteins. In tubulin, the flexible C-terminal tail of both subunits, a region of about 15 residues, concentrates many post-translational modifications, some of them being specific to tubulin or found in only a few others proteins. By comparison, methylation is a more widespread modification of macromolecules and is found in the “core” part of tubulin. So far, three SET-domain containing methyltransferases have been identified to target different lysine residues of alpha-tubulin. SETD2 was described to methylate Lys40, a mark recognized by the reader protein PBRM1. In addition, SET8 modifies Lys311, an activity that is enhanced in the presence of the transcription factor LSF protein, and SMYD2 targets Lys394. Because these three methyltransferases also modify histones, these findings raise the hypothesis of a cross-talk between the epigenome and the cytoskeleton.

In this general framework, the MICROMETHYL project aims to characterize the catalytic mechanism of the methylation of tubulin, the consequences of these modifications on the dynamics of the microtubules, and their functions in the cell. We will develop enzymology and structural biology approaches, associated with microtubule assembly experiments, both in bulk and at the single microtubule level by TIRF microscopy, and with cell biology experiments. Therefore, the MICROMETHYL project is first and foremost a project of fundamental research. However, because the dysregulation of the proteins involved in tubulin methylation, whether the methyltransferases, the oncogenic LSF protein or the PBRM1 reader protein, is associated with human diseases, and to cancers in particular, our project will also have more applied benefits in the medical field.