Doctor of Philosophy
Dr. Michael J. Strong
Dr. Arthur Brown
Aberrant phosphorylation and pathological deposition of the microtubule associated protein tau (tau protein) is associated with toxicity and cellular death in a number of neurodegenerative diseases (tauopathies). Specific phosphorylation sites are of interest in the processes leading to tau protein toxicity. One site of interest on tau protein is Thr175 (pThr175), which has been identified in diseased brain tissue from individuals with amyotrophic lateral sclerosis with cognitive impairment (ALSci) and Alzheimer’s disease. In vitro, pseudophosphorylation at this residue has been shown to induce the formation of pathological tau fibrils and, apoptotic cell death.
In my thesis, I have investigated the mechanism of cellular toxicity following phosphorylation of tau protein at Thr175. After showing that Thr175 pseudophosphorylation alone is insufficient to initiate tau protein fibrillization, I demonstrated that tau phosphorylation at Thr175 directly leads to the induction of kinase glycogen synthase kinase-3β (GSK3β) which in turn phosphorylates tau protein at Thr231. Both of these steps are necessary for the cytotoxicity of pThr175 tau to be manifest. I have shown that the pharmacological inhibition of this process leading to Thr231 phosphorylation prevents both fibril formation and cell death. To determine the extent to which this pathological process of Thr231 phosphorylation was applicable across the tauopathies in general, I characterized the presence of pThr175, activated GSK3b, pThr231tau and oligomeric tau formation across multiple tauopathies. In doing so, I demonstrated that this pathway may play an integral role in the generation of pathological tau deposition beyond that discovered for ALSci.
I then characterized pThr175 tau protein pathology in the trauma-associated neurodegenerative disease chronic traumatic encephalopathy (CTE) and CTE with amyotrophic lateral sclerosis (CTE-ALS), demonstrating the presence of pThr175 tau protein in pathology associated with these diseases as well. In order to determine whether the induction of pathological pThr175 tau was a primary event in the induction of this neuropathology, I used a rat model of moderate traumatic brain injury in which I demonstrated that after a single cortical impact, phosphorylation of endogenous tau protein at Thr175 was persistently elevated. pThr175 tau was present in axonal pathology as well as tau protein fibrillar neuronal pathology.
In order to definitively prove that pThr175tau was sufficient to induce tau pathology in vivo, I undertook somatic gene transfer of a rAAV9 construct expressing pseudophosphorylated human pThr175 tau (Thr175-Asp tau) in young adult rat hippocampus. I observed that one year following the stereotactic inoculation of this modified viral vector, rats developed tau pathology in construct-expressing hippocampal neurons along with caspase-3 cleavage. While the construct was similarly expressed in control rats, including empty vector and wild-type human tau, none of these latter rats developed pathology.
These findings indicate that phosphorylation of human tau at Thr175 triggers the pathological phosphorylation of tau protein at Thr231 through activation of GSK3b, and that this cascade leads to pathological fibril formation in vitro and in vivo. I have further demonstrated that this pathological process may have broader applicability than to the pathogenesis of ALSci, and includes a broad range of tauopathies in addition to CTE and CTE-ALS.
Moszczynski, Alexander, "Phosphorylation of tau protein at Thr175 is a toxic event associated with neurodegeneration" (2017). Electronic Thesis and Dissertation Repository. 4892.
Available for download on Saturday, August 31, 2019