A balanced and functional set of proteins is crucial for cell viability, as proteins are
the key players in biological processes that direct almost all cellular functions. Ag-
gregation of proteins into highly ordered amyloid fibrils is associated with loss of
protein function as well as gain of toxic function and is, therefore, considered a se-
rious hazard for cells. Fibrils of the human protein Tau are a hallmark of several
neurodegenerative diseases termed Tauopathies, including Alzheimer’s disease as
a prominent example. Despite intensive research, the underlying molecular mecha-
nisms and cellular strategies for preventing Tauopathies are still not well under-
stood. Recently, Tau as well as Tau fibrils were identified as substrates of the human
serine protease HTRA1. Defining features such as combination of protease and
chaperone function, ATP independence, and tightly regulated reversible activation
make HTRA1 a remarkable factor of the protein quality control system. In this study,
I investigated the interactions of HTRA1 with pathological Tau species and thus
gained detailed mechanistic insights into the process of fibril degradation.

Hyperphosphorylated and fibrillar Tau were generated as disease-relevant Tau spe-
cies, and their interactions with HTRA1 were analyzed using different biochemical
assays. Comparison with native Tau revealed that HTRA1 distinguishes native and
pathological Tau species, and specifically targets fibrils by conformation-specific
recognition. Interactions between HTRA1 and Tau fibrils result in activation of
HTRA1 and, subsequently, efficient degradation of the fibrils. Visualization of the
degradation process by atomic force microscopy revealed a mechanism with simul-
taneous degradation of Tau molecules along the entire length of a fibril by multiple
HTRA1 molecules. Combining time-resolved and cross-linking mass spectrometry,
I identified initial interactions between loop L3 of the HTRA1 protease domain and
the C-terminal region of the Tau molecules within the fibril. Subsequently, the tightly
packed fibril core is completely degraded into small peptides within minutes, making
reassembly seem unfeasible. Preliminary experiments indicate similar degradation
of fibrils formed by amyloid-β peptide, suggesting a general role of HTRA1 in coun-
teracting amyloid-associated pathologies. Overall, this study provides new insights
into how fibrils, characterized by strong inter- and intramolecular interactions, are degraded and contributes to a deeper understanding of the mechanisms cells have
evolved to counteract protein aggregation.