PML- and Smyd1-dependent regulatory mechanisms and effects on inflammatory responses of vascular endothelial cells

• Applicant: Dr. Janine Berkholz
• Subject Area: Anatomy and Physiology
• Term: since 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 418689898

A single layer of endothelial cells (ECs) lines the interior surface of blood vessels and lymphatic vessels. The loss of proper endothelial function can occur as a chronic inflammatory disease as in arteriosclerosis with specific anatomical localization or as in diabetes mellitus. But they can also arise acut as in the case of a sepsis or in the case of ischemic-reperfusion injury. In these inherently very different situations, inflammatory changes of the endothelium play a central role in the functional integrity of the endothelium of the affected blood vessel. The overall goal of the proposed project is to investigate the role of Smyd1 and PML in inflammatory responses in vascular endothelial cells (ECs). We will validate the hypothesis that Smyd1 and PML (or PML nuclear bodies) are coordinately involved in the perception of inflammatory stimuli to then elicit inflammatory responses in ECs. Smyd1 is a transcription factor that can also catalyze the trimethylation of histone protein 3 at lysine 4 (H3K4me3). PML is the main component of highly dynamic multiprotein complexes called PML nuclear bodies, which are assembled and disassembled in the nucleus and cause, among other things, the SUMOylation of proteins. In the first funding period, we showed that Smyd1 and PML in ECs can interact with each other in PML nuclear bodies and control their mutual activities as signaling molecules. The interaction of the two proteins was regulated by lipopolysaccharides or the inflammatory mediators TNF-α and interferon-γ and affected the production and secretion of IL-6 from ECs. The Smyd1-PML module therefore offers, among other things, a new approach to explain the sequence of "cytokine storm" and "compensatory anti-inflammatory response syndrome" in the course of sepsis or other inflammatory situations. The continuation of the project aims to systematically investigate the involvement of the Smyd1-PML module in inflammatory reactions in ECs. Inflammatory responses will be experimentally triggered in cell cultures either by exotoxins, inflammatory mediators, hyperglycemia, pathological flow conditions or hypoxia to subsequently determine the activation level of the Smyd1-PML module (Smyd1-dependent histone modifications and recruitment of HDACs; SUMOylation of proteins controlled by PML nuclear bodies). In addition, we will experimentally analyze the influence of Smyd1 and PML on inflammation-related endothelial cell functions (release of inflammatory cytokines, PD1-PDL1 axis, inflammasome activation, hemostasis regulation, and leukocyte adhesion) by experimentally varying the concentration and/or biological activity of Smyd1 and PML in ECs. For this purpose, we will also use new Smyd1 inhibitors that we identified in the first funding period. Furthermore, the establishment of translational approaches should be prepared through the development of complex cell culture models.

• DFG Programme: Research Grants