Our group is driven by the curiosity to identify molecular and cellular mechanisms of post-translational protein modifications in neurodegenerative and protein aggregation disorders that have a neuroinflammatory component. Special focus is given on protein aggregation in Alzheimer’s and Parkinson’s disease and on neuroinflammation in stroke. These clinical conditions share glutaminyl cyclase-catalyzed pyroglutamate modification of aggregation-prone and/or pro-inflammatory proteins and its reversal by antagonizing enzymes. This provides a basis for pharmacological interference with pathological pathways characteristic for these clinical entities. We combine cellular and animal experimental models with the analyses of human brain tissue from the respective clinical conditions
Glutaminyl cyclase (glutaminyl peptide cyclotransferase; QPCT, QC) and its isoenzyme QPCT-like (QPCTL, isoQC) catalyze the formation of N-terminal pyroglutamate (pGlu) on a number of neuropeptides, peptide hormones, chemokines and cell surface receptors. This post-translational modification protects these proteins from proteolytical degradation and, in some instances, enhances their biological activity. The substrate specificity of QCs arises from the distinct subcellular localization of QC along the secretory pathway and of isoQC to the Golgi apparatus. In addition to the importance of QCs in physiological processes, they are implicated in the pathological pGlu modification of Abeta in Alzheimer’s disease, of a-Synuclein in Parkinson’s disease and of the chemokine CCL2 in inflammatory conditions such as stroke. In specific research projects outlined below we investigate the importance and relative contributions of QCs.
Signatures of modified Abeta variants in Alzheimer's disease
We hypothesize that defined Abeta peptide variants contribute to the pathogenesis and progression of Alzheimer’s disease (AD) in a specific manner. Abeta peptides represent a heterogeneous group of peptides with differing biophysical and cell biological characteristics that are even more diversified by specific post-translational modifications. We employ a set of existing and novel monoclonal antibodies for the comparative analyses of phosphorylated, nitrated, pyroglutamate- and isoaspartate-modified Abeta peptides in brain parenchyma and blood vessels. We analyze post mortem human brain tissue from pre-symptomatic and symptomatic AD cases in comparison to brain tissue from control subjects and subjects who suffered from other types of dementia. The sequence of appearance of the respective Abeta peptides is also assessed in different transgenic mouse models with amyloid pathology. Additionally, we will reveal aggregation characteristics and neurotoxic profiles of modified Abeta peptides.
Post-translational α-synuclein modification in synucleinopathies
We recently discovered a pyroglutamate (pGlu) modified fragment of a-synuclein (αSyn) which promotes oligomer formation and neurotoxicity in synucleinopathies. The synthesis of the newly identified pGlu79-αSyn fragment requires two enzymatic activities to generate the N-terminal Q79 residue and to subsequently catalyze glutamine cyclization into pGlu. The best candidates for these enzymes are matrix metalloproteinase-3 (MMP-3) and glutaminyl cyclase (QC). We analyze the co-localization of pGlu79-αSyn with MMP-3 and QC in human brain tissue of Parkinson’s disease and dementia with Lewy body subjects and in transgenic mouse models of synucleinopathies. In addition, we investigate structural characteristics of pGlu-modified αSyn by using Small-angle X-ray scattering and Synchrotron radiation circular dichroism. QC might have a similar pathogenic profile in synucleinopathies as in amyloid pathologies such as Alzheimer´s disease and, therefore, might be considered as a drug target for these clinical conditions as well.
Reversal of post-translational protein modification in Alzheimer's disease and Parkinson's disease
We hypothesize that neurotoxic post-translational protein modification of Abeta and of α-synuclein can be enzymatically reverted. Alzheimer’s disease (AD) and Parkinson’s disease (PD) are characterized by protein aggregations of amyloid β (Abeta) and α-synuclein (aSyn), respectively. Both, Abeta and αSyn can undergo a number of pathogenic post-translational modifications (PTMs). Among these PTMs, phosphorylation, nitration and pyroglutamination are prominent examples. We hypothesize that specific enzymes reverting these post-translational protein modifications also detoxify post-translationally modified Abeta and αSyn protein species. To test this hypothesis, we analyze brain samples of the human clinical conditions – AD and PD – and their transgenic mouse models for co-localization of the modified peptides with their putatively degrading enzymes. Furthermore, pharmacological interference and genetic ablation will be helpful to analyze molecular mechanisms of the degradation of modified Abeta and αSyn variants.
Inflammation in stroke
We hypothesize that the biological activity of CCL2 is regulated by enzymatic stabilization and degradation resulting in endogenous downregulation of inflammation. The biological activity of the pro-inflammatory chemokine CCL2 is regulated enzymatically in a number of clinical conditions. On the one hand, N-terminal CCL2 truncation by dipeptidyl peptidase-4 (DP4) initiates further proteolytical degradation and abolishes its biological activity. On the other hand, N-terminal pyroglutamate modification catalyzed by QC/isoQC protects CCL2 from proteolytical degradation and increases its biological activity. We investigate the consequences of QC/isoQC, DP4 and CCL2 knock-out in an inflammatory stroke mouse model. By analyzing neurological motor deficits, infarct size and response of immune cells within the brain, the project targets an important aspect related to the endogenous downregulation of inflammation by enzyme processing pathways.
Alteration of structure-stabilizing proteins after stroke
Together with Prof. Dominik Michalski, Department of Neurology, University of Leipzig, we explore the importance of cell structure-stabilizing proteins, vascular and extracellular matrix components after cerebral infarction to find starting points for future neuroprotective therapeutic strategies. Ischemic stroke leads to long-term tissue damage within affected brain areas, depending on the extent and duration of the energy and oxygen deficiency due to vascular occlusion. To date, detailed information on the importance of cell structure stabilizing elements in the intra- and extracellular area during the development of this tissue damage is lacking. By using multiple stroke models, we aim to investigate the role and dynamic interrelationship of diverse components of the neurovascular unit (NVU).