Selective neurodegeneration in Alzheimer's disease and Parkinson's disease

Alzheimer’s disease (AD) and Parkinson’s disease (PD) are featured by cholinergic and dopaminergic neuron loss, respectively. As a unique pathological hallmark of AD, neuritic plaques contain aggregated amyloid β protein (Aβ), generated from amyloid β precursor protein (APP). APP mutations cause familial AD; mutations in the alpha-synuclein (SNCA) and leucine-rich repeat kinase 2 (LRRK2) genes are associated with PD. Recent studies suggest that the level of LRRK2 affects its toxicity in neurons. Therefore, understanding the mechanisms underlying LRRK2 expression would help to examine its pathogenic effects on PD. However, the features of the LRRK2 promoter remain elusive. In the first project, we cloned and characterized the LRRK2 promoter. There were two functional cis-acting specificity protein 1(Sp1)-responsive elements in its promoter. Our study demonstrates that LRRK2 transcription and translation were facilitated by Sp1 overexpression and blocked by an Sp1 inhibitor in vitro. The Lewy bodies primarily consist of α-synuclein protein, encoded by SNCA, and SNCAA₅₃T mutation promotes α-synuclein aggregation. The Swedish APP mutation (APPSWE) promotes Aβ generation and AD pathogenesis. However, the mechanisms underlying selective neurodegeneration in AD and PD are still unknown. In the second project, we stably overexpressed wildtype and mutated APP and SNCA genes in cholinergic SN56 and dopaminergic MN9D cells. APPSWE and SNCAA₅₃T mutations enhanced Aβ generation and α-synuclein inclusion formation in SN56 and MN9D cells, respectively. Aβ₄₂ and mutant α-synuclein oligomers caused severe cell death in SN56-APPSWE and MN9D-SNCAA53T cells, respectively. Furthermore, syndecan 3 (SDC3) and fibroblast growth factor receptor like 1 (FGFRL1) genes were identified as two of the differentially expressed genes in APP- and SNCA- related stable cells by microarrays. SDC3 was increased in the cholinergic nucleus of APPSWE knock-in mouse brains, whereas FGFRL1 was elevated in dopaminergic neurons in SNCAA₅₃T transgenic mice. Finally, knockdown of SDC3 and FGFRL1 attenuated oxidative stress-induced cell death in SN56-APPSWE and MN9D-SNCAA₅₃T cells. Overall, these demonstrate that SDC3 and FGFRL1 mediated the specific effects of APPSWE and SNCAA₅₃T on cholinergic and dopaminergic neurodegeneration in AD and PD, respectively. Our study suggests that SDC3 and FGFRL1 could be potential targets to alleviate the selective neurodegeneration in AD and PD.Medicine, Faculty ofGraduat

Regulation of LRRK2 promoter activity and gene expression by Sp1

Background: The dopaminergic neurodegeneration in the nigrostriatal pathway is a prominent neuropathological feature of Parkinson’s disease (PD). Mutations in various genes have been linked to familial PD, and leucine-rich repeat kinase 2 (LRRK2) gene is one of them. LRRK2 is a large complex protein, belonging to the ROCO family of proteins. Recent studies suggest that the level of LRRK2 protein is one of the contributing factors to PD pathogenesis. However, it remains elusive how LRRK2 is regulated at the transcriptional and translational level. Results In this study, we cloned a 1738 bp 5’-flanking region of the human LRRK2 gene. The transcriptional start site (TSS) was located to 135 bp upstream of translational start site and the fragment −118 to +133 bp had the minimum promoter activity required for transcription. There were two functional Sp1- responsive elements on the human LRRK2 gene promoter revealed by electrophoretic mobility shift assay (EMSA). Sp1 overexpression promoted LRRK2 transcription and translation in the cellular model. On the contrary, application of mithramycin A inhibited LRRK2 transcriptional and translational activities. Conclusion This is the first study indicating that Sp1 signaling plays an important role in the regulation of human LRRK2 gene expression. It suggests that controlling LRRK2 level by manipulating Sp1 signaling may be beneficial to attenuate PD-related neuropathology.Medicine, Faculty ofPsychiatry, Department ofReviewedFacult

Transcriptional activation of USP16 gene expression by NFκB signaling

Ubiquitin Specific Peptidase 16 (USP16) has been reported to contribute to somatic stem-cell defects in Down syndrome. However, how this gene being regulated is largely unknown. To study the mechanism underlying USP16 gene expression, USP16 gene promoter was cloned and analyzed by luciferase assay. We identified that the 5′ flanking region (− 1856 bp ~ + 468 bp) of the human USP16 gene contained the functional promotor to control its transcription. Three bona fide NFκB binding sites were found in USP16 promoter. We showed that p65 overexpression enhanced endogenous USP16 mRNA level. Furthermore, LPS and TNFα, strong activators of the NFκB pathway, upregulated the USP16 transcription. Our data demonstrate that USP16 gene expression is tightly regulated at transcription level. NFκB signaling regulates the human USP16 gene expression through three cis-acting elements. The results provide novel insights into a potential role of dysregulation of USP16 expression in Alzheimer’s dementia in Down Syndrome.Arts, Faculty ofNon UBCPsychology, Department ofReviewedFacult