Principal Investigator: Regine Heller (Institute of Molecular Cell Biology, University Hospital Jena)
Rohama Zahid (PhD):Functional characterization of carboxymethylated proteins in endothelial cells
Ageing is known to be associated with the accumulation of advanced glycation end products such as carboxymethylated proteins in cells and tissues. However, little is known about how cells respond to glycative stress and how glycation of specific proteins influences cell signaling and function. Previous studies of our group revealed that treatment of endothelial cells with the reactive dicarbonyl glyoxal induced robust formation of carboxymethyllysine (CML) as well as alterations in protein abundance. More than 120 CML-modified proteins mainly related to metabolism and cytoskeleton regulation and the respective sites were identified by mass spectrometry analysis. One of the identified carboxymethylated proteins is tubulin whose modification seems to lead to altered microtubule dynamics. The current project is aimed at characterizing microtubule dynamics in living endothelial cells using fluorescence-labeled tubulin and relate it to possible microtubule dysfunction. Furthermore, CML modification of tubulin will be characterized in different models of endothelial ageing and
Previous studies of our group revealed that treatment of endothelial cells with glyoxal induced robust formation of carboxymethyllysine (CML), an advanced glycation end product (AGE) as well as alterations in the abundance of cellular proteins. Among others we found an increased expression and activity of AMP-activated protein kinase (AMPK), an important sensor and regulator of cellular energy metabolism, and an upregulation of heme oxygenase 1 (HO-1). The current project will reveal mechanisms and functional consequences of AMPK upregulation in response to enhanced AGE formation with a special focus on autophagy regulation by AMPK. In addition, we will test, which role the Nrf2-HO-1 pathway plays in counteracting glycative stress. In this context, we will link CML modification of selected proteins to their respective functions and test their role in replicative senescent or chronologically aged endothelial cells.
Pia Senst (MD):Characterisation of the glyoxalase system in endothelial cells