Biology subjects

Kirstein, J.

Publications and source records attributed to Kirstein, J..

7 recordsLinked to original sources

bioRxiv2026

Comparative analysis of neuronal proteolytic pathways reveals neuron-specific and sub-compartmental-specific capacities with aging

Proteostasis is essential for maintaining neuronal function, and its dysregulation is a hallmark of aging and neurodegeneration. The ubiquitin-proteasome system (UPS) and macroautophagy are the two major proteolytic pathways responsible for protein degradation. However, their capacity and regulation differ between cell types and across aging. To elucidate the activity of both proteolytic pathways with aging, we performed a comparative analysis of the activity of UPS and macroautophagy in distinct neuronal subcellular compartments, in the cytosol and at synaptic terminals, across aging in neurons of Mus musculus (mouse) and Caenorhabditis elegans (nematode). In mice, our results identified differences between brain areas. While the cortical proteasomal activity declined with aging in both the cytoplasmic as well as synaptic neuronal subcompartments, the cerebellar proteasomal activity decreased only in the cytoplasmic compartment with aging. In C. elegans, we detected a decrease of proteasomal activity in both cytoplasmic and synaptic compartments of neurons. Interestingly, we observed a dysregulation of macroautophagy in both neuronal subcompartments of the cortex and cerebellum in mice as well as in C. elegans neurons with aging. Thus, we uncovered neuron-specific and subcompartmental-specific proteolytic capacities with aging that could manifest in different neuronal vulnerabilities for proteotoxic challenges with aging.

biochemistry↗

bioRxiv2025

HSP-1-Specific Nanobodies Alter Chaperone Function in vitro and in vivo

Targeted regulation of 70 kilodalton Heat Shock Protein (HSP70) chaperones, particularly the essential cognate heat shock protein (HSC70) and its Caenorhabditis elegans ortholog, HSP-1, may hold the key to improving cellular proteostasis and ameliorating aging-associated conditions linked to protein misfolding and aggregation. However, tools to selectively modulate HSP70 chaperone activity remain elusive. In this study, we pioneer the development of two novel nanobodies, B12 and H5, which specifically bind to both recombinant and endogenous HSP-1. We show that these nanobodies, differing by only two amino acids in their complementarity-determining regions, bind specifically to HSP-1 and effectively reduce both HSP-1 ATPase activity and protein folding capacity in a dose-dependent manner in vitro. We further demonstrate in vivo expression of B12, but not H5, in transgenic C. elegans strains reduces heat-stress survival and proteotoxic-stress resistance, mirroring the effects of hsp-1 knockdown via RNA interference. Our findings suggest that these nanobodies can serve as effective and specific tools for modulating HSP-1 chaperone activity in vivo. These discoveries provide a foundation for future research exploring the therapeutic potential of HSP70-targeting nanobodies in aging and protein misfolding diseases.

biochemistry↗

bioRxiv2025

Impairment of neuronal activity occurs at the early stages of the aggregation cascade of Ab1-42 and mutant Tau

Alzheimers disease (AD) is a progressive neurodegenerative disease that is characterized by the accumulation of amyloid-{beta} (A{beta}) plaques and neurofibrillary Tau tangles, ultimately leading to brain atrophy and death. To elucidate the relationship between the aberrant folding and aggregation of A{beta} and mutant Tau and neuronal function, we monitored neuronal activity in C. elegans AD models across age. Our findings reveal that expression of both A{beta} and Tau lead to significant reductions in neuronal activity and function in young adult animals preceding the accumulation of amyloid aggregates. Notably, A{beta} expression and aggregation in muscle tissue produced comparable detrimental effects on neuronal activity as its expression in neurons, suggesting that proteotoxic stress in muscle can influence neuronal function. This may occur through the propagation of A{beta} from muscle to neurons or through retrograde signaling pathways. Further, our new sub-stoichiometrically labeled Tau strains highlight that TauP301L,V337M has a significant impact on neuronal activity throughout aging. These results enhance our understanding of the early functional effects of amyloid aggregation in Alzheimers disease.

neuroscience↗

bioRxiv2025

A fluorescent folding reporter uncovers myosin misfolding as a driver of Hypertrophic Cardiomyopathy

Hypertrophic cardiomyopathy (HCM) is a fatal genetic disorder causing the thickening of ventricular walls in the heart muscle. While certain mutations in cardiac myosin deregulate ATPase activity, the pathology mechanism of most HCM mutations is not known. Here, by designing a fluorescent reporter to monitor myosin folding in cells, we uncovered a distinct class of HCM mutations that cause graded defects in myosin maturation. Using C. elegans as a disease model, we found that folding deficient HCM variants cause myofilament disruption, impaired motility, and reduced lifespan. Dietary restrictions resulted in a near-complete recovery from these detrimental defects by activating autophagy pathways through insulin/TOR signaling. In conclusion, our study identifies myosin misfolding as an important driver of HCM, revealing therapeutic opportunities to counteract muscle protein disorders.

cell biology↗

bioRxiv2024

Amyloid Beta Precursor Protein contributes to brain aging and learning decline in short-lived turquoise killifish (Nothobranchius furzeri)

Brain aging is a key risk factor for many neurodegenerative diseases, yet its molecular and cellular mechanisms remain elusive. Amyloid-beta precursor protein (APP) is among the most studied proteins linked to brain pathology; however, its role in non-pathological brain aging remains poorly characterized. Here, we investigate the natural impact of APP on normal brain aging using the short-lived turquoise killifish (Nothobranchius furzeri), which exhibits rapid and spontaneous age-related decline. We found that a pyroglutamated APP derivative (APPpE11) accumulates intra-neuronally in an age-dependent manner, co-localizing with a marker of cell death. We found that intraneuronal APPpE11 is also present in brains from healthy elderly humans, suggesting deep evolutionary conservation. To determine APPs role in spontaneous brain aging, we knock-out "amyloid precursor protein a" (appa) in killifish via CRISPR/Cas9. The lack of appa mitigated brain aging from a proteome-wide perspective, reduced age-related cell death and inflammation, and improved neuronal activity and learning capacity in aged individuals. Our findings show an ancestral and previously unrecognized role of amyloid-beta precursor protein in non-pathological brain aging, making it an ideal target for anti-aging interventions.

neuroscience↗

bioRxiv2024

Nodal Modulator (NOMO) is a force-bearing transmembrane protein required for muscle differentiation

The endoplasmic reticulum (ER) relies on specialized membrane-shaping proteins to maintain a continuous network of sheets and tubules that host distinct biological processes. How this intricate structure of the ER membrane system is maintained under conditions of mechanical strain is incompletely understood. NOMO is an ER-resident transmembrane protein that contributes to ER morphology and is highly expressed in striated muscle. In this study, we identify a critical interface between distal Ig domains that enables NOMO to maintain ER morphology and buffer mechanical forces. By incorporating two independent tension sensors in the luminal domain of NOMO, we demonstrate that NOMO assemblies experience forces in the single piconewton (pN) range, with a significant contribution from the identified interface. These newly defined features are important, if not indispensable, for myogenesis, as interface mutations affecting mechanosensing fail to restore the essential role of NOMO during myogenesis in a C2C12 differentiation model. Moreover, NOMO depletion impairs nematode motility, underscoring a broader functional importance in muscle physiology.

cell biology↗

bioRxiv2021

A versatile mitochondria isolation- and analysis-pipeline generates 3D nano-topographies and mechano-physical surface maps of single organelles

Living eukaryotic cells typically contain large quantities of highly dynamic mitochondria, which sustain the cells energy and redox homeostasis. Growing evidence suggests that mitochondria can functionally differ among but also within cells. The extent and biological significance of mitochondrial diversity is still largely unexplored, due to technical limitations that hamper profiling of individual organelles. Previous measurements of the cells interior have shown that membrane-bound compartments respond to metabolic manipulation by changes in their surface stiffness, suggesting that mechano-physical properties are a valuable readout of mitochondrial function. We here present the establishment of a robust multi-step analysis pipeline that allows one to profile mechano-physical properties of single mitochondria at the nanoscale using Atomic Force Microscopy (AFM). Firstly, we developed a rapid cell-type specific isolation protocol (mRACE), which selectively functionalizes mitochondria with biotin, facilitating isolation by streptavidin decorated microbeads. We established the technique for human and rat cell cultures, the invertebrate Caenorhabditis elegans, and the model plant Arabidopsis thaliana. Based on this versatile tool, we detected diversity of mitochondrially associated proteins among different tissues, reflecting the trophic condition of the source material. Secondly, a rapid filtration-based mitochondria isolation protocol was established, which was combined with mRACE. Lastly, we established an AFM analysis platform, which generates 3D maps of the nano-topography and mechano-physical properties of individual mitochondria. The comparison of mitochondria with each other revealed an unprecedented diversity in their mechano-physical properties and suggests that shape is not the sole determining parameter for outer membrane stiffness. We expect our results to not only introduce a new dimension for basic mitochondrial research, but in addition to open the door for the exploitation of individual mitochondria for diagnostic characterization.

plant biology↗