Biology subjects

Yacoubian, T. A.

Publications and source records attributed to Yacoubian, T. A..

5 recordsLinked to original sources

bioRxiv2025

Knockout of Rab27b exacerbates neuropathology in alpha-synuclein mouse models

Parkinsons Disease (PD) and other synucleinopathies are characterized by the formation of inclusions comprised of alpha-synuclein (syn) among other proteins, but the mechanisms by which these inclusions form and cause toxicity are not well understood. We have previously reported that the small GTPase Rab27b modulates autophagic-lysosomal function in neurons and supports lysosomal degradation of syn across multiple syn cellular models. Knockout (KO) and knockdown of Rab27b damage lysosomal degradative capacity and exacerbate syn pathology, while Rab27b overexpression is conversely protective in cellular syn models. Elevations of Rab27b seen in human synucleinopathies suggest a compensatory role for Rab27b in these disorders. Here, we examined the role Rab27b plays in vivo in the context of both A53T genetic syn overexpression and viral AAV syn overexpression mouse models. Rab27b knockout in A53T+ mice did not alter motor behavior or survival. However, Rab27b knockout increased proteinase-K resistant syn in the cortex, striatum, and substantia nigra of A53T mice starting as early as six months of age. Additionally, Rab27b KO increased phosphorylated S129 syn in the cortex and nigra. Astrocyte and microglial activation were also observed upon Rab27b KO in the A53T model. In the AAV syn model, Rab27b KO resulted in accelerated dopaminergic cell loss in the nigra. Collectively, we report that loss of Rab27b results in elevated neuropathology in PD-relevant brain regions, validating its role as a therapeutic target in synucleinopathies.

neuroscience↗

bioRxiv2024

Rab27b promotes lysosomal function and alpha-synuclein clearance in neurons

Alpha-synuclein (syn) is the key pathogenic protein implicated in synucleinopathies including Parkinsons Disease (PD) and Dementia with Lewy Bodies (DLB). In these diseases, syn is thought to spread between cells where it accumulates and induces pathology; however, mechanisms that drive its propagation or aggregation are poorly understood. We have previously reported that the small GTPase Rab27b is elevated in human PD and DLB and that it can mediate the autophagic clearance and toxicity of syn in a paracrine syn cell culture neuronal model. Here, we expanded our previous work and further characterized a role for Rab27b in neuronal lysosomal processing and syn clearance. We found that Rab27b KD in this syn inducible neuronal model resulted in lysosomal dysfunction and increased syn levels in lysosomes. Similar lysosomal proteolytic defects and enzymatic dysfunction were observed in both primary neuronal cultures and brain lysates from Rab27b knockout (KO) mice. Syn aggregation was exacerbated in Rab27b KO neurons upon treatment with syn preformed fibrils. We found no changes in lysosomal counts or lysosomal pH in either model, but we did identify defects in acidic vesicle trafficking in Rab27b KO primary neurons which may drive lysosomal dysfunction and promote syn aggregation. Rab27b OE enhanced lysosomal activity and reduced insoluble syn accumulation. Finally we found elevated Rab27b levels in human postmortem incidental Lewy Body Disease (iLBD) subjects relative to healthy controls. These data suggest a role for Rab27b in neuronal lysosomal activity and identify it as a potential therapeutic target in synucleinopathies.

neuroscience↗

bioRxiv2023

14-3-3 phosphorylation inhibits 14-3-3theta's ability to regulate LRRK2 kinase activity

LRRK2 mutations are among the most common genetic causes for Parkinsons disease (PD), and toxicity is associated with increased kinase activity. 14-3-3 proteins are key interactors that regulate LRRK2 kinase activity. Phosphorylation of the 14-3-3{theta} isoform at S232 is dramatically increased in human PD brains. Here we investigate the impact of 14-3-3{theta} phosphorylation on its ability to regulate LRRK2 kinase activity. Both wildtype and the non-phosphorylatable S232A 14-3-3{theta} mutant reduced the kinase activity of wildtype and G2019S LRRK2, whereas the phosphomimetic S232D 14-3-3{theta} mutant had minimal effects on LRRK2 kinase activity, as determined by measuring autophosphorylation at S1292 and T1503 and Rab10 phosphorylation. However, wildtype and both 14-3-3{theta} mutants similarly reduced the kinase activity of the R1441G LRRK2 mutant. 14-3-3{theta} phosphorylation did not promote global dissociation with LRRK2, as determined by co-immunoprecipitation and proximal ligation assays. 14-3-3s interact with LRRK2 at several phosphorylated serine/threonine sites, including T2524 in the C-terminal helix, which can fold back to regulate the kinase domain. Interaction between 14-3-3{theta} and phosphorylated T2524 LRRK2 was important for 14-3-3{theta}s ability to regulate kinase activity, as wildtype and S232A 14-3-3{theta} failed to reduce the kinase activity of G2019S/T2524A LRRK2. Molecular modeling showed that 14-3-3{theta} phosphorylation causes a partial rearrangement of its canonical binding pocket, thus affecting the interaction between 14-3-3{theta} and the C-terminus of LRRK2. We conclude that 14-3-3{theta} phosphorylation destabilizes the interaction of 14-3-3{theta} with LRRK2 at T2524, which consequently promotes LRRK2 kinase activity.

neuroscience↗

bioRxiv2020

14-3-3s reduce propagation and toxicity of pathologic alpha-synuclein in the in vivo preformed fibril model

Alpha-synuclein (syn) is the key component of proteinaceous aggregates termed Lewy Bodies (LBs) that pathologically define a group of disorders known as synucleinopathies, including Parkinsons Disease (PD) and Dementia with Lewy Bodies (DLB). Syn is hypothesized to misfold and spread throughout the brain in a prion-like fashion. Transmission of syn necessitates the release of misfolded syn from one cell and the uptake of that syn by another, in which it can template the misfolding of endogenous syn upon cell internalization. 14-3-3 proteins are a family of highly expressed brain proteins that are neuroprotective in multiple PD models. We have previously shown that 14-3-3{theta} acts as a chaperone to reduce syn aggregation, cell-to-cell transmission, and neurotoxicity in the in vitro pre-formed fibril (PFF) model. In this study, we expanded our studies to test the impact of 14-3-3s on syn toxicity in the in vivo syn PFF model. We used both transgenic expression models and adenovirus associated virus (AAV)-mediated expression to examine whether 14-3-3 manipulation impacts behavioral deficits, syn aggregation, and neuronal loss in the PFF model. 14-3-3{theta} transgene overexpression in cortical and amygdala regions rescued social dominance deficits induced by PFFs at 6 months post injection, whereas 14-3-3 inhibition by transgene expression of the competitive 14-3-3 peptide inhibitor difopein in the cortex and amygdala accelerated social dominance deficits. The behavioral rescue by 14-3-3{theta} overexpression was associated with delayed syn aggregation induced by PFFs in these brain regions. Conversely, 14-3-3 inhibition by difopein in the cortex and amygdala accelerated syn aggregation and cortical pyramidal neuron loss induced by PFFs. 14-3-3{theta} overexpression by AAV in the substantia nigra (SN) also delayed syn aggregation in the SN and partially rescued PFF-induced dopaminergic cell loss in the SN. 14-3-3 inhibition in the SN accelerated nigral syn aggregation and increased PFF-induced dopaminergic cell loss. These data indicate a neuroprotective role for 14-3-3{theta} against syn toxicity in vivo.

neuroscience↗

bioRxiv2020

Rab27 GTPases regulate alpha-synuclein uptake, cell-to-cell 1 transmission, and toxicity

Parkinsons disease and Dementia with Lewy Bodies are two common neurodegenerative disorders marked by proteinaceous aggregates composed primarily of the protein -synuclein. -Synuclein is hypothesized to have prion-like properties, by which misfolded -synuclein induces the pathological aggregation of endogenous -synuclein and neuronal loss. Rab27a and Rab27b are two highly homologous Rab GTPases that regulate -synuclein secretion, clearance, and toxicity in vitro. In this study, we tested the impact of Rab27a/b on the transmission of pathogenic -synuclein. Double knockout of both Rab27 isoforms eliminated -synuclein aggregation and neuronal toxicity in primary cultured neurons exposed to fibrillary -synuclein. In vivo, Rab27 double knockout mice lacked fibril-induced -synuclein inclusions, dopaminergic neuron loss, and behavioral deficits seen in wildtype mice with fibril-induced inclusions. Studies using AlexaFluor488-labeled -synuclein fibrils revealed that Rab27a/b knockout prevented -synuclein internalization without affecting bulk endocytosis. Rab27a/b knockout also blocked the cell-to-cell spread of -synuclein pathology in multifluidic, multichambered devices. This study provides critical insight into the role of Rab GTPases in Parkinsons disease and identifies Rab27s as key players in the progression of synucleinopathies.

neuroscience↗