Biology subjects

Heidenreich, M.

Publications and source records attributed to Heidenreich, M..

4 recordsLinked to original sources

bioRxiv2024

The phase separation landscape of genome-wide genetic perturbations

Biomolecular organization is central to cell function. While phase separation is a key mechanism orchestrating this organization, we lack a comprehensive view of genes that can globally influence this process in vivo. To identify such genes, we combined functional genomics and synthetic biology. We developed a bioorthogonal system that can identify changes in the intracellular milieu that globally tune phase separation. We measured in vivo phase diagrams of a synthetic system across >25 million cells in 2,888 yeast knockouts, and identified 68 genes whose deletion alters the phase boundaries of the synthetic system, an unexpected result given the systems bioorthogonal design. Genes involved in TORC1 signaling and metabolism, particularly carbohydrate-, amino acid- and nucleotide synthesis were enriched. The mutants that changed phase separation also showed high pleiotropy, suggesting that phase separation interrelates with many aspects of biology. Highlights- A synthetic protein system reveals the genetic and environmental tunability of protein phase separation - Genetic knockouts affecting phase separation are highly pleiotropic - Carbohydrate, amino acid, and nucleotide metabolism contribute to modulating phase separation potential - Protein phase separation is a globally tunable property of the intracellular environment O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/620319v1_ufig1.gif" ALT="Figure 1"> View larger version (81K): org.highwire.dtl.DTLVardef@4b8403org.highwire.dtl.DTLVardef@1c80503org.highwire.dtl.DTLVardef@c10bf2org.highwire.dtl.DTLVardef@1f76c71_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

Structural determinants of co-translational protein complex assembly

The assembly of proteins into functional complexes is critical to lifes processes. While textbooks depict complex assembly as occurring between fully synthesized proteins, we know today that thousands of proteins in the human proteome assemble co-translationally during their synthesis. Why this process takes place, however, remains unknown. We show that co-translational assembly is governed by biophysical and structural characteristics of the protein complex, and involves mutually stabilized, intertwined subunits. Consequently, these subunits are also co-regulated across the central dogma, from transcription to protein degradation. Leveraging structural signatures with AlphaFold2-based predictions enables us to accurately predict co-translational assembly on a proteome-wide scale, which we validated by ribosome profiling, genetic perturbations, and smFISH experiments. Notably, the latter showed that co-translationally assembling subunits exhibit co-localized mRNAs. This work unveils a fundamental connection between protein structure and the translation process, highlighting the overarching impact of three-dimensional structure on gene expression, mRNA localization, and proteostasis. One Sentence SummaryProtein complexes with topologically intertwined subunits require co-translational assembly and synchronized proteostasis of subunits, with implications in protein stability, mRNA localization, and evolution. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=195 SRC="FIGDIR/small/576408v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@712a2org.highwire.dtl.DTLVardef@58894borg.highwire.dtl.DTLVardef@9db5c6org.highwire.dtl.DTLVardef@8eae4_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗

bioRxiv2023

Removal of a genomic duplication by double-nicking CRISPR restores synaptic transmission and behavior in the MyosinVA mutant mouse Flailer.

Copy number variations, and particularly duplications of genomic regions, have been strongly associated with various neurodegenerative conditions including autism spectrum disorder (ASD). These genetic variations have been found to have a significant impact on brain development and function, which can lead to the emergence of neurological and behavioral symptoms. Developing strategies to target these genomic duplications has been challenging, as the presence of endogenous copies of the duplicate genes often complicates the editing strategies. Using the ASD and anxiety mouse model Flailer, that contains a duplication working as a dominant negative for MyoVa, we demonstrate the use of DN-CRISPRs to remove a 700bp genomic duplication in vitro and in vivo. Importantly, DN-CRISPRs have not been used to remove more gene regions <100bp successfully and with high efficiency. We found that editing the flailer gene in primary cortical neurons reverts synaptic transport and transmission defects. Moreover, long-term depression (LTD), disrupted in Flailer animals, is recovered after gene edition. Delivery of DN-CRISPRs in vivo shows that local delivery to the ventral hippocampus can rescues some of the mutant behaviors, while intracerebroventricular delivery, completely recovers Flailer animal phenotype associated to anxiety and ASD. Our results demonstrate the potential of DN-CRISPR to efficiently (>60% editing in vivo) remove large genomic duplications, working as a new gene therapy approach for treating neurodegenerative diseases.

neuroscience↗

bioRxiv2020

Designer protein assemblies with tunable phase diagrams in living cells

The self-organization of proteins into specific assemblies is a hallmark of biological systems. Principles governing protein-protein interactions have long been known. However, principles by which such nanoscale interactions generate diverse phenotypes of mesoscale assemblies, including phase-separated compartments, remains challenging to characterize and understand. To illuminate such principles, we create a system of two proteins designed to interact and form mesh-like assemblies in living cells. We devise a novel strategy to map high-resolution phase diagrams in vivo, which provide mesoscale self-assembly signatures of our system. The structural modularity of the two protein components allows straightforward modification of their molecular properties, enabling us to characterize how point mutations that change their interaction affinity impact the phase diagram and material state of the assemblies in vivo. Both, the phase diagrams and their dependence on interaction affinity were captured by theory and simulations, including out-of-equilibrium effects seen in growing cells. Applying our system to interrogate biological mechanisms of self-assembly, we find that co-translational protein binding suffices to recruit an mRNA to the designed micron-scale structures.Competing Interest StatementThe authors have declared no competing interest.View Full Text

synthetic biology↗