Biology subjects

Marek-Trzonkowska, N.

Publications and source records attributed to Marek-Trzonkowska, N..

4 recordsLinked to original sources

bioRxiv2024

Universal toolset for mass spectrometric analysis of intracellular peptidome and small protein fraction

The analysis of native intracellular peptidome has gained significant attention in recent years. However, there is still a need for more knowledge regarding various sample preparation methods that facilitate efficient and reproducible recovery of peptides, which can then be analyzed using quantitative liquid chromatography-mass spectrometry. A similar situation exists in small proteome research, typically defined as polypeptides with masses of less than 100 amino acids, often too long for easy identification without enzymatic digestion. In this context, we describe a set of methods that involve simple denaturation and solid-phase extraction of polypeptides, applicable for isolating short intracellular polypeptides within the desired length range. Our work demonstrates the efficiency and reproducibility of these methods for quantitative analysis of the peptidome in mammalian cells. Additionally, we investigated the flexibility of adjusting the mass range through ultrafiltration. We have shown that these methods can be adapted for highly efficient enrichment and fractionation of small proteins, resulting in polypeptide isolates suitable for tryptic digestion and intact protein analysis. Moreover, we describe the use of freely available computational tools that can effectively manage the analysis of the resulting data. The research presented here will benefit the global scientific community in both fundamental (protein turnover, proteolytic processing, non-canonical open reading frames, etc.) and applied sciences (bioactive/neuro peptide discovery, precision medicine, vaccines, etc.), and other areas that could benefit from selective analysis of short native polypeptides.

cancer biology↗

bioRxiv2024

Sample-specific network analysis identifies gene coexpression patterns of immunotherapy response in advanced kidney cancer

Immunotherapies have recently emerged as a standard of care for advanced cancers, offering remarkable improvements in patient prognosis. However, only a small proportion of patients respond to the treatment, and no definitive molecular hallmark has been identified for clinical use in predicting treatment outcomes. Here, we propose a sample-specific weighted gene network approach to investigate the heterogeneity of patient clinical outcomes by leveraging multiscale network features derived from gene expression data. Our results show that patients exhibiting similar clinical benefits share comparable gene coexpression patterns. Increased gene connectivity and stronger negative gene-gene associations are also pivotal factors in patients with poor prognoses. Moreover, integrating pathway genes with topological measures enables the identification of the perturbed regulation of biological pathways associated with treatment responses. Additionally, sample-level network features enhance the prediction performance of gene expression values-based machine learning models. Collectively, our approach provides valuable guidance on the use of gene network information to stratify cancer patients and to optimize treatment strategies.

bioinformatics↗

bioRxiv2022

Major Histocompatibility Complex class I heavy chains localize in both cytoplasmic and nuclear compartment

The Major Histocompatibility Complex class I (MHC-I) molecules present antigenic peptides (AP) to CD8+ T cells for self versus non-self recognition. Loading of AP on MHC-I takes place in the endoplasmic reticulum (ER), upon shuttling of cytoplasmic AP substrates to the ER. Understanding of this process has been influenced by the view that MHC-I antigens are produced from the proteasomal degradation of cellular proteins. Recent observations on the intronic and untranslated region-derived peptides as well as on the non-AUG translation products presented on the MHC-I open the possibility that antigenic peptides can derive from pre-spliced mRNAs translated in the nuclear compartment. In this brief report, we show that a fraction of human MHC-I molecules (human leukocyte antigens type A, HLA-A) is present in the nuclei of cells, in the proximity of histone H2B. With this finding, we hope to initiate a new direction of research on the nuclear role of MHC-I and ask whether the loading of antigens can take place in the nuclear compartment.

immunology↗

bioRxiv2022

Keap1-resistant ΔN-Nrf2 isoform does not translocate to the nucleus upon electrophilic stress

The Nrf2 pathway is an essential defense pathway in a cell. It responds to oxidative and electrophilic stress via derepression of Nrf2 from Keap1-Cul3-mediated degradation, accumulation of Nrf2 in the nucleus and transcriptional activation of a number of detoxifying and cell protective Nrf2 target genes. Here we report that normal and cancer cells also express the N-terminally truncated Nrf2 isoform ({Delta}N-Nrf2), which originates from an alternative promoter. Co-immunoprecipitation together with molecular dynamics simulation showed that the binding between {Delta}N-Nrf2 and Keap1 is impaired, resulting in the much higher stability of this form. {Delta}N-Nrf2 is retained in the cytoplasm in response to electrophilic stress, indicating that it does not regulate transcription under the same stress stimuli as the full-length Nrf2. Altogether this data suggests that Nrf2 has other functions in cells than transcriptional activation of genes, which most probably rely on the protein-protein interactions in the cytoplasm. The regulation between these functions takes place on the level of transcription. Significance StatementThis work signifies the importance of alternative transcription in assigning the function to the produced protein. Nrf2 transcripts produced from the second promoter of the Nrf2 gene give rise to the N-terminally truncated Nrf2 form ({Delta}N-Nrf2), which is retained in the cytoplasm upon stress, thus it has a different role in cells than transcriptional regulation. {Delta}N-Nrf2 is resistant to the Keap1-Cul3 degradation pathway and is highly expressed in all tested cell types. This work points to the new, cytoplasmic role of Nrf2 in cells, determined at the level of transcription.

cell biology↗