Biology subjects

Nicolet, B. P.

Publications and source records attributed to Nicolet, B. P..

9 recordsLinked to original sources

bioRxiv2023

Learning the sequence code for mRNA and protein abundance in human immune cells

Accurate protein expression in human immune cells is essential for appropriate cellular function. The mechanisms that define protein abundance are complex and executed on transcriptional, post-transcriptional and post-translational level. Here, we present SONAR, a machine learning pipeline that learns the endogenous sequence code and that defines protein abundance in human cells. SONAR uses thousands of sequence features (SFs) to predict up to 63% of the protein abundance independently of promoter or enhancer information. SONAR uncovered the cell type-specific and activation-dependent usage of SFs. The deep knowledge of SONAR provides a map of biologically active SFs, which can be leveraged to manipulate the amplitude, timing, and cell type-specificity of protein expression. SONAR informed on the design of enhancer sequences to boost T cell receptor expression and to potentiate T cell function. Beyond providing fundamental insights in the regulation of protein expression, our study thus offers novel means to improve therapeutic and biotechnology applications. One Sentence SummarySONAR informs the design of cell type-specific protein expression in human cells

cell biology↗

bioRxiv2022

Re-use of labware reduces CO2 equivalent footprint and running costs in laboratories

Laboratory-based research is resource intensive in terms of financial costs and its carbon footprint. Research laboratories require immense amounts of energy to power equipment, as well as large volumes of materials, particularly of single-use item consumption. In fact, many laboratories have essentially become reliant on single-use plastics. Understanding the full carbon footprint of consumable usage is increasingly important as many research institutes commit to carbon neutrality. To date, no carbon footprint assessment has been conducted to detail the differences between single-use plastics, and reusable glass in a laboratory setting. Here, we analyse the CO2 equivalent (CO2e) footprint of utilising single-use plastics, and re-use of glass or plastic items within laboratory environments. We focused our assessment on four commonly utilised consumables for mammalian cell and bacterial culture, and found that re-use scenarios resulted in substantial reduction in CO2e footprint up to 11-fold. In addition, we estimated the long-term financial costs of re-use and single-use scenarios, and found that re-use had either similar or much lower running costs even when including technical staff wage. We concluded that research facilities must foster re-use in laboratory consumables, while reserving single-use items for select, defined cases. Our study highlights the need to account for indirect CO2e footprint in designing a carbon-neutral lab.

cell biology↗

bioRxiv2021

Mapping RNA-binding proteins in human B cells and Tcells upon differentiation

B cells and T cells are key players in the defence against infections and malignancies. To exert their function, B cells and T cells differentiate into effector and memory cells. Tight regulation of these differentiation processes is key to prevent their malfunction, which can result in life-threatening disease. Lymphocyte differentiation relies on the appropriate timing and dosage of regulatory molecules, and post-transcriptional gene regulation (PTR) is a key player herein. PTR includes the regulation through RNA-binding proteins (RBPs), which control the fate of RNA and its translation into proteins. To date, a comprehensive RBP expression map throughout lymphocyte differentiation is lacking. Using transcriptome and proteome analyses, we here provide an RBP expression map for human B cells and T cells. We observed that even though the overall RBP expression is conserved, the relative RBP expression is distinct between B cells and T cells. Differentiation into effector and memory cells alters the RBP expression, resulting into preferential expression of different classes of RBPs. For instance, whereas naive T cells express high levels of translation-regulating RBPs, effector T cells preferentially express RBPs that modulate mRNA stability. Lastly, we found that cytotoxic CD8+ and CD4+ T cells express a common RBP repertoire. Combined, our study reveals a cell type-specific and differentiation-dependent RBP expression landscape in human lymphocytes, which will help unravel the role of RBPs in lymphocyte function.

immunology↗

bioRxiv2021

CD29 enriches for cytotoxic human CD4+ T cells

CD4+ T cell are key contributors in the induction of adaptive immune responses against pathogens. Even though CD4+ T cells are primarily classified as non-cytotoxic helper T cells, it has become appreciated that a subset of CD4+ T cells is cytotoxic. However, tools to identify these cytotoxic CD4+ T cells are lacking. We recently showed that CD29 (Integrin Beta 1, ITGB1) expression on human CD8+ T cells enriches for the most potent cytotoxic T cells. Here, we questioned whether CD29 expression also associates with cytotoxic CD4+ T cells. We show that human peripheral blood-derived CD29hiCD4+ T cells display a cytotoxic gene expression profile, which closely resembles that of CD29hi cytotoxic CD8+ T cells. This CD29hi cytotoxic phenotype was observed ex vivo and was maintained in in vitro cultures. CD29 expression enriched for CD4+ T cells, which effectively produced the pro-inflammatory cytokines IFN-{gamma}, IL-2, and TNF-, and cytotoxic molecules. Lastly, CD29-expressing CD4+ T cells transduced with a MART-1 specific TCR showed target cell killing in vitro. In conclusion, we here demonstrate that CD29 can be employed to enrich for cytotoxic human CD4+ T cells.

immunology↗

bioRxiv2020

Abundant circular RNA expression in terminal myeloid differentiation does not correlate with frequent circRNA translation, or with circRNA-mediated translation regulation

Each day, about 1012 erythrocytes and platelets are released into the blood stream. This substantial output from hematopoietic stem cells is tightly regulated by transcriptional and epigenetic factors. Whether and how circular RNAs (circRNAs) contribute to the differentiation and/or identity of hematopoietic cells is to date not known. We recently reported that erythrocytes and platelets contain the highest levels and numbers of circRNAs amongst hematopoietic cells. Here, we provide the first detailed analysis of circRNA expression during erythroid and megakaryoid differentiation. CircRNA expression not only significantly increased upon enucleation, but also had limited overlap between progenitor cells and mature cells, suggesting that circRNA expression stems from regulated processes rather than resulting from mere accumulation. To study circRNA function in hematopoiesis, we first compared the expression levels of circRNAs with the translation efficiency of their mRNA-counterpart. We found that only 1 out of 2531 (0.04%) circRNAs associated with mRNA-translation regulation. Furthermore, irrespective of 1000s of identified putative open reading frames, deep ribosome-footprinting sequencing and mass spectrometry analysis provided little evidence for translation of endogenously expressed circRNAs. In conclusion, circRNAs alter their expression profile during terminal hematopoietic differentiation, yet their contribution to regulate cellular processes remains enigmatic.

molecular biology↗

bioRxiv2020

Limited but gene-class specific correlation of mRNA and protein expression in human CD8+ T cells

T cells are key players in our defence against infections and malignancies. When T cells differentiate or become activated, they undergo substantial alterations in gene expression. Even though RNA expression levels are now well documented throughout different stages of T cells, it is not well understood how mRNA expression translates into the protein landscape. By combining paired RNA sequencing and massspectrometry data of primary human CD8+ T cells, we report that mRNA expression is a poor proxy for the overall protein output, irrespective of the differentiation or activation status. Yet, gene class stratification revealed a function-specific correlation of mRNA with protein expression. This gene class-specific expression pattern associated with differences in gene characteristics such as sequence conservation and untranslated region (UTR) lengths. In addition, the presence of AU-rich elements in the 3UTR associated with alterations in mRNA and protein abundance T cell activation dependent, gene class-specific manner. In conclusion, our study highlights the role of gene characteristics as a determinant for gene expression in T cells.

immunology↗

bioRxiv2019

CD29 marks superior cytotoxic human T cells

Cytotoxic CD8+ T cells can effectively kill target cells by producing cytokines, chemokines and granzymes. Expression of these effector molecules is however highly divergent, and tools that identify and pre-select potent killer cells are lacking. Human CD8+ T cells can be divided into IFN-{gamma} and IL-2 producing cells. Unbiased RNA-sequencing and proteomics analysis on cytokine-producing fixed cells revealed that IL-2+ T cells produce helper cytokines, and that IFN-{gamma}+ T cells produce cytotoxic molecules. IFN-{gamma}+ T cells could be identified with the surface marker CD29 already prior to stimulation. CD29 also marked T cells with cytotoxic gene expression from different tissues in single-cell RNA-sequencing data. Notably, the cytotoxic features of CD29+ T cells were maintained during cell culture, suggesting a stable phenotype. Pre-selecting CD29-expressing MART1 TCR-engineered T cells potentiated the killing of target cells. We therefore propose that selecting for CD29+ T cells could boost the anti-tumoral activity of T cell therapeutics.

immunology↗

bioRxiv2018

Critical role of post-transcriptional regulation for IFN-γ in tumor-infiltrating T cells

Protective T cell responses against tumors require the production of Interferon gamma (IFN-{gamma}). However, tumor-infiltrating T cells (TILs) gradually lose their capacity to produce IFN-{gamma} and therefore fail to clear malignant cells. Dissecting the underlying mechanisms that block cytokine production is thus key for improving T cell products. Here we show that although TILs express substantial levels of Ifng mRNA, post-transcriptional mechanisms impede the production of IFN-{gamma} protein due to loss of mRNA stability. CD28 triggering, but not PD1 blocking antibodies, effectively restores the stability of Ifng mRNA. Intriguingly, TILs devoid of AU-rich elements within the 3untranslated region maintain stabilized Ifng mRNA and produce more IFN-{gamma} protein than wild-type TILs. This sustained IFN-{gamma} production translates into effective suppression of tumor outgrowth, which is almost exclusively mediated by direct effects on the tumor cells. We therefore conclude that post-transcriptional mechanisms could be modulated to potentiate effective T cell therapies in cancer.

immunology↗

bioRxiv2018

Circular RNA expression in human hematopoietic cells is widespread and cell-type specific

Hematopoietic stem cells differentiate into a broad range of specialized blood cells. This process is tightly regulated and depends on transcription factors, micro-RNAs, and long non-coding RNAs. Recently, also circular RNA (circRNA) were found to regulate cellular processes. Their expression pattern and their identity is however less well defined. Here, we provide the first comprehensive analysis of circRNA expression in human hematopoietic progenitors, and in differentiated lymphoid and myeloid cells. We here show that the expression of circRNA is cell-type specific, and increases upon maturation. circRNA splicing variants can also be cell-type specific. Furthermore, nucleated hematopoietic cells contain circRNA that have higher expression levels than the corresponding linear RNA. Enucleated blood cells, i.e. platelets and erythrocytes, were suggested to use RNA to maintain their function, respond to environmental factors or to transmit signals to other cells via microvesicles. Here we show that platelets and erythrocytes contain the highest number of circRNA of all hematopoietic cells, and that the type and numbers of circRNA changes during maturation. This cell-type specific expression pattern of circRNA in hematopoietic cells suggests a hithero unappreciated role in differentiation and cellular function.

immunology↗