Biology subjects

Gustafsson, C.

Publications and source records attributed to Gustafsson, C..

14 recordsLinked to original sources

bioRxiv2026

Macroalgal fucoidan can activate the biological carbon pump

Macroalgae secrete complex carbohydrate polymers, their extracellular matrix, as protection against microbial degradation. By resisting breakdown, these carbohydrates can contribute to marine carbon sequestration, though mechanisms, extent, and timescales remain unknown. Using ship-based sampling and experiments, we found that brown macroalgae release 1.7-4.2% of carbon fixation as fucoidan, equivalent to 0.32-0.88 mg fucoidan per gram of dry seaweed tissue per day. A Bayesian model trained on our empirical data, coupled with Monte Carlo simulations suggests an annual global release of 13-37 megatons fucoidan carbon. Moreover, degradation resistance combined with surface-activity enabled fucoidan to act as glue that cross-linked allochthonous organic carbon including microbes and proteins into marine snow. Notably, substantial fucoidan exudation was universally conserved across all tested species and regions. Thus, any brown macroalgal species can be used e.g. via aquafarming to enhance the formation of marine snow.

ecology↗

bioRxiv2026

Orthogonal Transposons for Iterative Genome Engineering of Mammalian Cells.

The contemporary shift toward multispecific antibodies, antibody-drug conjugates (ADCs), and bespoke glycoengineered therapeutics have exposed the limitations of standard genomic engineering tools. This paper presents a novel iterative engineering paradigm utilizing the Leap-In Transposase(R) platform. By leveraging a suite of three mutually orthogonal transposase-transposon systems, we demonstrate the sequential modification of the Chinese Hamster Ovary (CHO) genome to achieve three distinct functional outcomes: (i) First, the creation of a glutamine synthetase (GS)-deficient host (CHO-K1-GS) via targeted knockdown, (ii) Second, the integration of multiple copies of a model therapeutic IgG1 for expression, and (iii) Third, the subsequent knockdown of the fucosylation pathway to modulate the glycan profile of the expressed IgG1. Genetic stability (copy number & sequence) of each integration event was confirmed using Targeted Locus Amplification (TLA) and Next-Generation Sequencing (NGS). Functional stability (expression levels, metabolic phenotype, and glycan phenotypes) was confirmed using standard cell culture and analytical techniques. Crucially, the truly orthogonal nature of the transposase-transposon pairs prevents cross-mobilization and ensures the structural and functional integrity of previously integrated cargo. This study establishes a "What You See Is What You Get" (WYSIWYG) methodology that provides a robust, scalable, and predictable framework for developing next-generation complex biopharmaceutical manufacturing cell lines.

bioengineering↗

bioRxiv2026

Glycoprotein G enables HSV-2 neuroinvasion and provides protection as a glycosylated vaccine antigen

The function of glycoprotein G (gG-2) of herpes simplex virus 2 (HSV-2) during genital infection is unknown. gG-2 is cleaved into a secreted variant (sgG-2) and a membrane associated variant (mgG-2). This work delineates the glycan profile of mgG-2, and demonstrates that mgG-2 is strongly immunogenic, eliciting glycan dependent humoral and Th1-polarized CD4+ T cell responses in a mouse vaccination model. The N- and O-glycosylation of mgG-2 is important to achieve full protection against HSV-2 infection as immunization with deglycosylated mgG-2 resulted in poor CD4+ T cell activation and viral spread to dorsal root ganglia and spinal cord. Furthermore, an mgG-2 negative HSV-2 mutant virus failed to spread to the dorsal root ganglia and the central nervous system of genitally infected mice, despite viral replication in vaginal cells. Our data demonstrate that mgG-2 is important for HSV-2 propagation and that adaptive immune responses targeting the glycosylated protein can prevent neuronal infection, identifying mgG-2 as a promising vaccine candidate.

microbiology↗

bioRxiv2026

Machine Learning Method for Optimizing Coding Sequences in Mammalian Cells

Mammalian cell lines are the preferred hosts for producing commercially relevant therapeutic proteins such as antibodies, multispecifics, and cytokine fusion proteins. Even though significant investment is made to optimize upstream and downstream processes, the optimal gene design parameters for heterologous recombinant protein expression remain poorly understood. We describe here a generic approach to gene optimization in which design features are systematically sampled and modulated iteratively using machine learning (ML). Synthetic genes encoding the Dasher fluorescent protein, differing only in synonymous codons, were used to interrogate the gene-sequence preferences of transient antibody-expressing HEK293 cells. Synonymous codon variations influenced expression by more than two orders of magnitude. This variation in protein yield was used to build ML models relating gene design features, which were then employed to design further-improved genes. The ML models were shown to be expression system-specific. Messenger RNA levels and ribosome occupancy were highly correlated with protein levels, suggesting that mRNA lifetime has a causal relationship with coding bias. Our results illustrate a novel, generally applicable method to improve gene expression via synonymous re-coding for any protein target or host cell.

bioengineering↗

bioRxiv2023

Aging is associated with functional and molecular changes in distinct hematopoietic stem cell subsets

Age is a risk factor for hematologic malignancies. Attributes of the aging hematopoietic system include increased myelopoiesis, impaired adaptive immunity, and a functional decline of the hematopoietic stem cells (HSCs) that maintain hematopoiesis. Changes in the composition of diverse HSC subsets have been suggested to be responsible for age-related alterations, however, the underlying regulatory mechanisms are incompletely understood in the context of HSC heterogeneity. In this study, we investigated how distinct HSC subsets, separated by CD49b, functionally and molecularly change their behavior with age. We demonstrate that blood lineage differentiation progressively shifts to a higher myeloid cellular output in both lymphoid-biased and myeloid-biased HSC subsets during aging. In parallel, we show that HSCs selectively undergo age-dependent gene expression and gene regulatory molecular changes in a progressive manner, which is initiated already in the pre-adult stage. Overall, our studies suggest that aging intrinsically alters both cellular and molecular properties of HSCs. HighlightsO_LIWith age a gradual shift towards myeloid differentiation occurs in both myeloid-biased and lymphoid-biased enriched HSC subsets. C_LIO_LIAge-related molecular changes preferentially occur in HSCs. C_LIO_LIFunctionally distinct HSC subsets with high transcriptional similarity can be distinguished on the epigenetic level. C_LIO_LIHSC aging is associated with a progressive increase in chromatin accessibility. C_LI

cell biology↗

bioRxiv2022

T-RHEX-RNAseq - A tagmentation-based, rRNA blocked, random hexamer primed RNAseq method for generating stranded RNAseq libraries directly from very low numbers of lysed cells.

BackgroundRNA sequencing has become the mainstay for studies of gene expression. Still, analysis of rare cells with random hexamer priming - to allow analysis of a broader range of transcripts - remains challenging. ResultsWe here describe a tagmentation-based, rRNA blocked, random hexamer primed RNAseq approach (T-RHEX-RNAseq) for generating stranded RNAseq libraries from very low numbers of FACS sorted cells without RNA purification steps. ConclusionT-RHEX-RNAseq provides an easy-to-use, time efficient and automation compatible method for generating stranded RNAseq libraries from rare cells.

molecular biology↗

bioRxiv2022

FOXO dictates initiation of B cell development and myeloid restriction in common lymphoid progenitors

The development of B cells relies on an intricate network of transcription factors critical for developmental progression and lineage commitment. In the B cell developmental trajectory, a temporal switch from predominant Foxo3 to Foxo1 expression occurs at the CLP stage. Utilizing VAV-iCre mediated conditional deletion, we found that the loss of FOXO3 impaired B cell development from LMPP down to B cell precursors, while the loss of FOXO1 impaired B cell commitment and resulted in a complete developmental block at the CD25 negative proB cell stage. Strikingly, the combined loss of FOXO1 and FOXO3 resulted in the failure to restrict the myeloid potential of CLPs and the complete loss of the B cell lineage. This is underpinned by the failure to enforce the early B-lineage gene regulatory circuitry upon a predominantly pre-established open chromatin landscape. Altogether, this demonstrates that FOXO3 and FOXO1 cooperatively govern early lineage restriction and initiation of B-lineage commitment in CLPs. SUMMARYCommon lymphoid progenitors co-express the transcription factors FOXO1 and FOXO3. Removing FOXO1 and FOXO3 at this developmental stage results in regained myeloid potential, failed establishment of the early B cell gene regulatory program, and the complete loss of the B cell lineage.

immunology↗

bioRxiv2021

Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift

The recently emerged SARS-CoV-2 Omicron variant harbors 37 amino acid substitutions in the spike (S) protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody therapeutics. Here, we show that the Omicron RBD binds to human ACE2 with enhanced affinity relative to the Wuhan-Hu-1 RBD and acquires binding to mouse ACE2. Severe reductions of plasma neutralizing activity were observed against Omicron compared to the ancestral pseudovirus for vaccinated and convalescent individuals. Most (26 out of 29) receptor-binding motif (RBM)-directed monoclonal antibodies (mAbs) lost in vitro neutralizing activity against Omicron, with only three mAbs, including the ACE2-mimicking S2K146 mAb1, retaining unaltered potency. Furthermore, a fraction of broadly neutralizing sarbecovirus mAbs recognizing antigenic sites outside the RBM, including sotrovimab2, S2X2593 and S2H974, neutralized Omicron. The magnitude of Omicron-mediated immune evasion and the acquisition of binding to mouse ACE2 mark a major SARS-CoV-2 mutational shift. Broadly neutralizing sarbecovirus mAbs recognizing epitopes conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers.

immunology↗

bioRxiv2021

Linked-read whole-genome sequencing resolves common and private structural variants in multiple myeloma

Multiple myeloma (MM) is an incurable and aggressive plasma cell malignancy characterized by a complex karyotype with multiple structural variants (SVs) and copy number variations (CNVs). Linked-read whole-genome sequencing (lrWGS) allows for refined detection and reconstruction of SVs by providing long-range genetic information from standard short-read sequencing. This makes lrWGS an attractive solution for capturing the full genomic complexity of MM. Here we show that high-quality lrWGS data can be generated from low numbers of FACS sorted cells without DNA purification. Using this protocol, we analyzed FACS sorted MM cells from 37 MM patients with lrWGS. We found high concordance between lrWGS and FISH for the detection of recurrent translocations and CNVs. Outside of the regions investigated by FISH, we identified >150 additional SVs and CNVs across the cohort. Analysis of the lrWGS data allowed for resolving the structure of diverse SVs affecting the MYC and t(11;14) loci causing the duplication of genes and gene regulatory elements. In addition, we identified private SVs causing the dysregulation of genes recurrently involved in translocations with the IGH locus and show that these can alter the molecular classification of the MM. Overall, we conclude that lrWGS allows for the detection of aberrations critical for MM prognostics and provides a feasible route for providing comprehensive genetics. Implementing lrWGS could provide more accurate clinical prognostics, facilitate genomic medicine initiatives, and greatly improve the stratification of patients included in clinical trials. KEY POINTS- Linked-read WGS can be performed without DNA purification and allows for resolving the diverse structural variants found in multiple myeloma. - Linked-read WGS can, as a stand-alone assay, provide comprehensive genetics in myeloma and other diseases with complex genomes.

genetics↗

bioRxiv2021

Distance decay 2.0 - a global synthesis of taxonomic and functional turnover in ecological communities.

Understanding the variation in community composition and species abundances, i.e., {beta}-diversity, is at the heart of community ecology. A common approach to examine {beta}-diversity is to evaluate directional turnover in community composition by measuring the decay in the similarity among pairs of communities along spatial or environmental distances. We provide the first global synthesis of taxonomic and functional distance decay along spatial and environmental distance by analysing 149 datasets comprising different types of organisms and environments. We modelled an exponential distance decay for each dataset using generalized linear models and extracted r2 and slope to analyse the strength and the rate of the decay. We studied whether taxonomic or functional similarity has stronger decay across the spatial and environmental distances. We also unveiled the factors driving the rate of decay across the datasets, including latitude, spatial extent, realm, and organismal features. Taxonomic distance decay was stronger along spatial and environmental distances compared with functional distance decay. The rate of taxonomic spatial distance decay was the fastest in the datasets from mid-latitudes while the rate of functional decay increased with latitude. Overall, datasets covering larger spatial extents showed a lower rate of decay along spatial distances but a higher rate of decay along environmental distances. Marine ecosystems had the slowest rate of decay. This synthesis is an important step towards a more holistic understanding of patterns and drivers of taxonomic and functional {beta}-diversity.

ecology↗

bioRxiv2021

Cell-intrinsic functions of the transcription factor Bhlhe40 in activated B cells and T follicular helper cells restrain the germinal center reaction and prevent lymphomagenesis

The generation of high-affinity antibodies against pathogens and vaccines requires the germinal center (GC) reaction - a process that relies on a complex interplay between specialized effector subsets of B and CD4 T lymphocytes - GC B cells and T follicular helper (TFH) cells. Intriguingly, several key positive regulators of the GC reaction are common for both cell types. Here, we report that the transcription factor Bhlhe40 is a crucial cell-intrinsic negative regulator affecting both the B and T cell sides of the GC reaction. In activated CD4 T cells, Bhlhe40 was required to restrain proliferation thus limiting the number of TFH cells. In B cells, Bhlhe40 executed its function in the first days after immunization by selectively restricting the generation of the earliest GC B cells but not of early memory B cells or plasmablasts. Conditional Bhlhe40 inactivation confirmed cell-autonomous functions of Bhlhe40 in both GC B and TFH cells, while the GC phenotype was further enhanced upon loss of Bhlhe40 in both cell types. This negative regulation of the GC reaction by Bhlhe40 was of crucial importance, as Bhlhe40-deficient mice with progressing age succumbed to a B cell lymphoma characterized by accumulation of monoclonal GC B-like cells and polyclonal TFH cells in various tissues.

immunology↗

bioRxiv2021

FOXO1 and FOXO3 cooperatively regulate innate lymphoid cell development

The natural killer (NK) and non-cytotoxic innate lymphoid cells (ILC) lineages play vital role in the regulation of the immune system. Yet understanding of mechanisms controlling NK/ILC development remains incomplete. The evolutionary conserved FOXO family of forkhead transcription factors are critical regulators of cellular processes. We found that the loss of FOXO1 and FOXO3 together caused impaired activation of the NK gene expression program and reduced ETS binding already at the common lymphoid progenitor (CLP) level and a block at the ILC progenitor (ILCP) to NK progenitor transition. FOXO controlled NK cell maturation in organ specific manner and their ability to respond to IL-15. At the ILCP level, disruption of the ILC lineage specific gene programs was associated with broad perturbation of the generation of the non-cytotoxic ILC subsets. We concluded that FOXO1 and FOXO3 cooperatively regulate ILC lineage specification at the progenitor level as well as the generation of mature ILCs.

cell biology↗

bioRxiv2018

The concerted action of E2-2 and HEB is critical for early lymphoid specification

The apparition of adaptive immunity in Gnathostomata correlates with the expansion of the E-protein family to encompass E2-2, HEB and E2A. Within the family, E2-2 and HEB are more closely evolutionarily related but their concerted action in hematopoiesis remains to be explored. Here we show that the combined disruption of E2-2 and HEB results in failure to express the early lymphoid program in CLPs and a near complete block in B-cell development. In the thymus, ETPs were reduced and T-cell development perturbed, resulting in reduced CD4 T- and increased {gamma}{delta} T-cell numbers. In contrast, HSCs, erythro-myeloid progenitors and innate immune cells were unaffected showing that E2-2 and HEB are dispensable for the ancestral hematopoietic lineages. Taken together, this E-protein dependence suggests that the appearance of the full Gnathostomata E-protein repertoire was critical to reinforce the gene regulatory circuits that drove the emergence and expansion of the lineages constituting humoral immunity.

immunology↗

bioRxiv2018

High-Throughput ChIPmentation: freely scalable, single day ChIPseq data generation from very low cell-numbers

Chromatin immunoprecipitation coupled to sequencing (ChIP-seq) is widely used to map histone modifications and transcription factor binding on a genome-wide level. Here, we present high-throughput ChIPmentation (HT-ChIPmentation) that eliminates the need for DNA purification prior to library amplification and reduces reverse-crosslinking time from hours to minutes. The resulting workflow is easily established, extremely rapid, and compatible with requirements for very low numbers of FACS sorted cells, high-throughput applications and single day data generation.

molecular biology↗