Biology subjects

Gerhardt, E.

Publications and source records attributed to Gerhardt, E..

5 recordsLinked to original sources

bioRxiv2026

LRRK2/LRRK1 interactions modulate Rab7 activity and inhibit lysosomal exocytosis

Mutations in the LRRK2 gene are the most common genetic cause of both familial and sporadic Parkinsons disease (PD). LRRK2 belongs to the leucine-rich repeat kinase (LRRK) family. Two members of the LRRK family exist in humans (LRRK1 and LRRK2). Although there is strong structural similarity between the two proteins, they have attracted very different levels of attention by the scientific community owing to the strong association between LRRK2 and PD. In contrast, the role of LRRK1 is relatively unexplored. LRRK2 is also known to regulate endolysosomal function, but its precise role in this process remains incompletely understood. Our study investigated the interaction between LRRK1 and LRRK2 under different cellular conditions, uncovering their role in modulating the endolysosomal system. We found that LRRK1 and LRRK2 interact and modulate each others activity, and that this interaction is reduced under starvation conditions. We also found that LRRK1 and LRRK2 have contrasting effects on lysosomal size, impacting on lysosomal exocytosis. Together, our findings suggest that LRRK2 regulates endolysosomal homeostasis, at least in part, by modulating LRRK1. Our findings offer new insight into the molecular mechanisms associated with lysosomal function and, ultimately, we anticipate this knowledge will help us better understand the molecular crosstalk between LRRK kinases and their contribution to PD pathogenesis. Graphical abstractStarvation reduces the interaction between LRRK2 and LRRK1 due to a conformational change in LRRK2. Under normal conditions, LRRK2/LRRK1 interaction enhances LRRK1 activity, leading to increased phosphorylation of Rab7. Disruption of the Rab7 cycle impairs lysosomal homeostasis, leading to lysosomal accumulation and an increase in lysosomal diameter. This enlargement negatively impacts lysosomal exocytosis. Created with BioRender.com. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=176 SRC="FIGDIR/small/731951v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@13ce58forg.highwire.dtl.DTLVardef@1034543org.highwire.dtl.DTLVardef@1b84096org.highwire.dtl.DTLVardef@198598f_HPS_FORMAT_FIGEXP M_FIG C_FIG

Cell Biology↗

bioRxiv2024

Synphilin-1 as a modulator of aSyn assembly

Alpha-synuclein (aSyn) is an intrinsically disordered protein that undergoes phase-separation and is associated with several neurodegenerative conditions. However, the function and the pathological role of aSyn are still elusive. Here, we modeled different types of aSyn assemblies in living cells, and developed a model that reports on gel and solid-like inclusions based on the coexpression of aSyn and synphilin-1 (Sph1). We identified striking morphological differences between aSyn-aSyn and Sph1-aSyn assemblies, characterized by distinct antibody recognition patterns, resistance to Proteinase K treatment, and protein mobilities. Importantly, we showed that the interaction between Sph1-aSyn can be manipulated, altering inclusion size and number. Sph1-aSyn interactions were central for inclusion formation and localization, and that inclusions include lysosomes and AP-1 vesicles, consistent with previous studies in human brain tissue. In total, we provide novel insight into the biology of protein aggregation, shedding light on potential therapeutic strategies that extend beyond conventional targets. Deciphering the role of Sph1 and other aSyn-interacting proteins on aSyn biology and pathobiology will be essential for treating synucleinopathies.

molecular biology↗

bioRxiv2021

Common molecular mechanisms underlie the transfer of alpha-synuclein, Tau and huntingtin and modulate spontaneous activity in neuronal cells

The misfolding and accumulation of disease-related proteins are common hallmarks among several neurodegenerative diseases. Alpha-synuclein (aSyn), Tau and huntingtin (wild-type and mutant, 25QHtt and 103QHtt, respectively) were recently shown to be transferred from cell-to-cell through different cellular pathways, thereby contributing to disease progression and neurodegeneration. However, the relative contribution of each of these mechanisms towards the spreading of these different proteins and the overall effect on neuronal function is still unclear. To address this, we exploited different cell-based systems to conduct a systematic comparison of the mechanisms of release of aSyn, Tau and Htt, and evaluated the effects of each protein upon internalization in microglial, astrocytic, and neuronal cells. In the models used, we demonstrate that 25QHtt, aSyn and Tau are released to the extracellular space at higher levels than 103QHtt, and their release can be further augmented with the co-expression of USP19. Furthermore, cortical neurons treated with recombinant monomeric 43QHtt exhibited alterations in neuronal activity that correlated with the toxicity of the polyglutamine expansion. Tau internalization resulted in an increase in neuronal activity, in contrast to slight effects observed with aSyn. Interestingly, all these disease-associated proteins were present at higher levels in ectosomes than in exosomes. The internalization of both types of extracellular vesicles (EVs) by microglial or astrocytic cells elicited the production of pro-inflammatory cytokines and promoted an increase in autophagy markers. Additionally, the uptake of the EVs modulated neuronal activity in cortical neurons. Overall, our systematic study demonstrates the release of neurodegenerative disease-associated proteins through similar cellular pathways. Furthermore, it emphasizes that protein release, both in a free form or in EVs, might contribute to a variety of detrimental effects in receiving cells and to progression of pathology, suggesting they may be exploited as valid targets for therapeutic intervention in different neurodegenerative diseases. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=153 SRC="FIGDIR/small/452825v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1400e8org.highwire.dtl.DTLVardef@1478fccorg.highwire.dtl.DTLVardef@1319abcorg.highwire.dtl.DTLVardef@1a5ad40_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2021

Ectosomes and exosomes are distinct proteomic entities that modulate spontaneous activity in neuronal cells

Extracellular vesicles (EVs) are important mediators in intercellular communication. However, understanding the biological origin and functional effects of EVs subtypes has been challenging due to the moderate differences in their physical properties and absence of reliable markers. Here, we characterize the proteomes of ectosomes and exosomes using an improved differential ultracentrifugation protocol and quantitative proteomics. Cytoskeleton and glycolytic proteins are distinctively present in ectosomes, while endosomal sorting complexes proteins and tetraspanins are enriched in exosomes. Furthermore, annexin-A2 was identified as a specific marker for ectosomes derived from cell media and human cerebrospinal fluid. Expression of EGFP as a cytosolic reporter leads to its incorporation in EVs and enables their imaging with higher resolution. Importantly, ectosomes and exosomes internalization in neuronal cells results in the modulation of neuronal spontaneous activity. Our findings suggest that EVs cargoes reflect core intracellular processes, and their functional properties might regulate basic biological and pathological processes.

neuroscience↗

bioRxiv2021

Cofilin pathology is a new player on α-synuclein-induced spine impairment in models of hippocampal synucleinopathy

Cognitive dysfunction and dementia are presently recognized as major complications in -synucleinopathies, namely in Dementia with Lewy Bodies (DLB) and Parkinsons disease with dementia (PDD). In these disorders, -Synuclein (Syn) accumulation affects severely the hippocampus by inducing synaptic dysfunction which culminates in cognitive impairment. To characterize the mechanisms underlying Syn-induced neuronal dysfunction we analysed the effect of overexpression or extracellular administration of Syn on hippocampal neurons. We observed that Syn induces the dysregulation of the actin-binding protein cofilin and its assembly into rod structures in a mechanism mediated by the cellular prion protein (PrPC). Moreover, we unraveled cofilin pathology as mediator of Syn-induced dendritic spine impairment in hippocampal neurons. Importantly, in a synucleinopathy mouse model with cognitive impairment we validated cofilin dysregulation and synaptic dysfunction at the same age when cognitive deficits were observed. Our data supports cofilin as a novel player on hippocampal synaptic dysfunction triggered by Syn on Lewy Body dementias.

neuroscience↗