Biology subjects

Jahn, L.

Publications and source records attributed to Jahn, L..

5 recordsLinked to original sources

bioRxiv2023

Understanding and exploitation of host stress responses to protein production using novel transcriptomic analytics

Predictable expression of heterologous genes in a production host is a fundamental challenge in biotechnology. While traditional methods focus on manipulating expression and the property of the heterologous gene, a systems biology approach can complement with designs to improve the host itself. Previously, Independent Component Analysis (ICA) of the RNAseq data helped reveal independently modulated gene sets (iModulons) in bacteria. This was later applied to identify common stress responses related to heterologous gene expression for Escherichia coli. In this study, we expand this analysis with additional non-enzymatic proteins and apply our findings to design novel protein production optimization. By leveraging the Precise-1K transcriptomics knowledge base, we identify three iModulons as novel transcriptional responses to protein production stress; Cold Shock, gcvB sRNA, and the uncharacterized UC-9 iModulons. By studying the gene membership in the UC-9 iModulon, we discover effective novel design targets for improving protein production. This study demonstrates the value of big data analytics and systems understanding of host responses for designing novel strategies to optimize protein production.

systems biology↗

bioRxiv2023

Leucine zipper-based sorting system enables generation of multi-functional CAR T cells

Resistance to chimeric antigen receptor (CAR) T cell therapy develops through multiple mechanisms including antigen-loss escape and tumor-induced immune suppression. Expression of multiple CARs may overcome multi-antigen-loss escape. Similarly, expression of switch receptors that convert inhibitory immune checkpoint signals into positive costimulatory signals may enhance CAR T cell activity in the tumor microenvironment. Engineering multiple features into one cell product, however, is limited by transgene packaging constraints of current vector systems. Here, we describe a leucine zipper-based cell sorting methodology that enables selective single-step immunomagnetic purification of cells co-transduced with two vectors, designed to potentially double the number of incorporated transgenes. This "Zip-sorting" system facilitated generation of T cells simultaneously expressing up to four CARs and co-expressing up to three switch receptors. These multi-CAR multi-Switch receptor arrays enabled T cells to eliminate antigenically heterogeneous syngeneic leukemia populations co-expressing multiple inhibitory ligands. Zip-sorted multi-CAR multi-Switch receptor T cells represent a potent therapeutic strategy to overcome multiple mechanisms of CAR T cell resistance.

synthetic biology↗

bioRxiv2023

Reconstructing the Transcriptional Regulatory Network of Probiotic L. reuteri is Enabled by Transcriptomics and Machine Learning

ILimosilactobacillus reuteri, a probiotic microbe instrumental to human health and sustainable food production, adapts to diverse environmental shifts via dynamic gene expression. We applied independent component analysis to 117 high-quality RNA-seq datasets to decode its transcriptional regulatory network (TRN), identifying 35 distinct signals that modulate specific gene sets. This study uncovers the fundamental properties of L. reuteris TRN, deepens our understanding of its arginine metabolism, and the co-regulation of riboflavin metabolism and fatty acid biosynthesis. It also sheds light on conditions that regulate genes within a specific biosynthetic gene cluster and the role of isoprenoid biosynthesis in L. reuteris adaptive response to environmental changes. Through the integration of transcriptomics and machine learning, we provide a systems-level understanding of L. reuteris response mechanism to environmental fluctuations, thus setting the stage for modeling the probiotic transcriptome for applications in microbial food production. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/547516v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@118ec4corg.highwire.dtl.DTLVardef@141b0b7org.highwire.dtl.DTLVardef@1b8cf6forg.highwire.dtl.DTLVardef@3ab181_HPS_FORMAT_FIGEXP M_FIG C_FIG Comprehensive iModulon Workflow Overview. Our innovative workflow is grounded in the analysis of the LactoPRECISE compendium, a curated dataset containing 117 internally sequenced RNA-seq samples derived from a diversity of 50 unique conditions, encompassing an extensive range of 13 distinct condition types. We employ the power of Independent Component Analysis (ICA), a cutting-edge machine learning algorithm, to discern the underlying structure of iModulons within this wealth of data. In the subsequent stage of our workflow, the discovered iModulons undergo detailed scrutiny to uncover media-specific regulatory mechanisms governing metabolism, illuminate the context-dependent intricacies of gene expression, and predict pathways leading to the biosynthesis of probiotic secondary metabolites. Our workflow offers an invaluable and innovative lens through which to view probiotic strain design while simultaneously highlighting transformative approaches to data analytics in the field.

bioinformatics↗

bioRxiv2022

A novel lineage of RORγt+Aire+ antigen presenting cells promotes peripheral generation of intestinal regulatory T cells and tolerance during early life

Establishing and maintaining tolerance to self- or innocuous foreign antigens is vital for preservation of organismal health. Within the thymus, medullary thymic epithelial cells (mTECs) expressing AutoImmune Regulator, Aire, play a critical role in self-tolerance through deletion of autoreactive T cells and promotion of thymic regulatory T (Treg) cell development. Within weeks of birth, a separate wave of Treg cell differentiation occurs in the periphery, upon exposure to dietary and commensal microbiota derived antigens, yet the cell types responsible for the generation of peripheral Treg (pTreg) cells are not known. Here we identified a new class of ROR{gamma}t+ antigen-presenting cells (APC), dubbed Thetis cells (TCs), with transcriptional features of both mTECs and dendritic cells (DCs), comprising 4 major sub-groups (TC I-IV). We uncovered a developmental wave of TCs within intestinal lymph nodes during a critical early life window, coincident with the wave of pTreg cell differentiation. While TC I and III expressed the signature mTEC nuclear factor Aire, TC IV lacked Aire expression and were enriched for molecules required for pTreg generation, including the TGF-{beta} activating integrin v{beta}8. Loss of either MHCII or Itgb8 expression by TCs led to a profound impairment in intestinal pTreg differentiation, with onset of intestinal inflammation. In contrast, MHCII expression by ROR{gamma}t+ group 3 innate lymphoid cells (ILC3) and classical DCs was neither sufficient nor required for pTreg generation, further implicating TCs as the tolerogenic ROR{gamma}t+ APC with an essential early life function. Our studies reveal parallel pathways for establishment of tolerance to self and foreign antigen within the thymus and periphery, marked by involvement of shared cellular and transcriptional programs.

immunology↗

bioRxiv2021

Extraction and high-throughput sequencing of oak heartwood DNA: assessing the feasibility of genome-wide DNA methylation profiling

Tree ring features are affected by environmental factors and therefore are the basis for dendrochronological studies to reconstruct past environmental conditions. Oak wood often provides the data for these studies because of the durability of oak heartwood and hence the availability of samples spanning long time periods of the distant past. Wood formation is regulated in part by epigenetic mechanisms such as DNA methylation. Studies of the methylation state of DNA preserved in oak heartwood thus could identify epigenetic tree ring features informing on past environmental conditions. In this study, we aimed to establish protocols for the extraction of DNA, the high-throughput sequencing of whole-genome DNA libraries (WGS) and the profiling of DNA methylation by whole-genome bisulfite sequencing (WGBS) for oak (Quercus spp.) heartwood drill cores taken from the trunks of living standing trees spanning the AD 1776-2014 time period. Heartwood contains little DNA, and large amounts of phenolic compounds known to hinder the preparation of high-throughput sequencing libraries. Whole-genome and DNA methylome library preparation and sequencing consistently failed for oak heartwood samples more than 100 and 50 years of age, respectively. DNA fragmentation increased with sample age and was exacerbated by the additional bisulfite treatment step during methylome library preparation. Relative coverage of the non-repetitive portion of the oak genome was sparse. These results suggest that quantitative methylome studies of oak hardwood will likely be limited to relatively recent samples and will require a high sequencing depth to achieve sufficient genome coverage.

plant biology↗