Biology subjects

Kardos, J.

Publications and source records attributed to Kardos, J..

3 recordsLinked to original sources

bioRxiv2023

Simulation of gap junction formation reveals critical role of cysteines in connexon coupling

Despite the fundamental functions of connexin gap junctions (CxGJs), understanding of molecular mechanisms, governing intercellular CxGJ formation by coupling of connexon hemichannels (CxHCs, connexons) is still in infancy. In silico simulation of intercellular connexon coupling of two Cx43HC models, embedded in membrane bilayers (Cx43HC-HC) successfully modelled the emergence of trans-gap junctional (trans-GJ) stabilization centers (SCs). Investigating the molecular determinants shaping the HC-HC interface, we revealed that the exceptionally high number of cysteine residues located at the interface play a pivotal role in the stabilization of HC and GJ structures. Opening of the disulphide bonds between these cysteines resulted in disappearance of trans-GJ SCs in the Cx43GJ model. In contrast, the Cx43HC form was found to be consistent with open disulphide bonds. Finally, we have shown that the presence of an adjoining HC contributes to disulphide formation and consequently to the emergence of trans-GJ H-bonds. Our results suggest that several connexon channels in vertebrates may undertake intercellular connexon coupling similarly and may bring forward to the targeting of CxGJ-specific coupling.

neuroscience↗

bioRxiv2020

Live cell imaging of single neurotrophin receptor molecules on human neuron in Alzheimer's disease

The changes in the receptor dynamics such as the surface movement of the receptor molecules on the plasma membrane are essential to receptor function. However, whether the receptor dynamics are affected by disease conditions is unknown. Neurotrophin receptors such as TrkA and p75NTR play a critical role in neuronal survival and their functions are highly affected in Alzheimers disease (AD). Using live-cell single-molecule imaging of neurotrophin receptors we examined the surface trafficking of TrKA and p75NTR molecules on human induced pluripotent stem cells (hiPSCs) derived live neurons from presenilin 1 (PSEN1) mutant AD patients and healthy subjects. Here we report that surface trafficking of p75NTR molecules on neurites is faster than that of TrkA molecules in healthy controls. The surface dynamics of TrkA molecules were elevated in AD patients compared to healthy individuals. In contrast, the surface movement of p75NTR was significantly smaller in AD patients compared to healthy individuals. Interestingly, amyloid beta1-42 (A{beta}1-42) administration increased the surface trafficking of both TrkA and p75NTR in healthy hiPSCs neurons. These findings provides the first evidence that the surface diffusion of TrkA and p75NTR molecules are altered in patients suffering from AD. Our data also suggest that A{beta}1-42 may responsible for the alteration of the surface movements of TrkA but not for p75NTR. One Sentence SummaryThe surface movements of neurotrophin receptors such as TrkA and p75NTR are altered in neurons derived from patients suffering from familial Alzheimers disease.

neuroscience↗

bioRxiv2019

High throughput competitive fluorescence polarization assay reveals functionalredundancy in the S100 protein family

The calcium-binding, vertebrate-specific S100 protein family consists of 20 paralogs in humans (referred as the S100ome), with several clinically important members. To assess their interactome, high-throughput, systematic analysis is indispensable, which allows one to get not only qualitative but quantitative insight into their protein-protein interactions (PPIs). We have chosen an unbiased assay, fluorescence polarization (FP) that revealed a partial functional redundancy when the complete S100ome (n=20) was tested against numerous model partners (n=13). Based on their specificity, the S100ome can be grouped into two distinct classes: promiscuous and orphan. In the first group, members bound to several ligands (>4-5) with comparable high affinity, while in the second one, the paralogs bound only one partner weakly, or no ligand was identified (orphan). Our results demonstrate that in vitro FP assays are highly suitable for quantitative ligand binding studies of selected protein families. Moreover, we provide evidence that PPI-based phenotypic characterization can complement the information obtained from the sequence-based phylogenetic analysis of the S100ome, an evolutionary young protein family.\n\nAuthor summaryFunctional similarity among a protein family can be essential in order to understand proteomic data, to find biomarkers, or in inhibitor design. Proteins with similar functions can compensate the loss-of-function of the others, their expression can co-vary under pathological conditions, and simultaneous targeting can lead to better results in the clinics. To investigate this property one can use sequence-based approaches. However, this path can be difficult. In the case of the vertebrate specific, evolutionary young, S100 family, phylogenetic approaches lead to ambiguous results. To overcome this problem, we applied a high-throughput biochemical approach to experimentally measure the binding affinities of a large number of S100 interactions. We performed unbiased fluorescence polarization assay, involving the complete human S100ome (20 paralogs) and 13 known interaction partners. We used this measured 20x13 (260) protein-protein interaction array to reveal the functional relationships within the family. Our work provide a general framework for studies focusing on phenotype-based domain classification.

biochemistry↗