An ancient competition for the conserved branchpoint sequence influences physiological and evolutionary outcomes in splicing DOI Creative Commons

Karen Larissa Pereira de Castro,

J.M. Abril, Kuo‐Chieh Liao

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 9, 2024

Recognition of the intron branchpoint during spliceosome assembly is a multistep process that defines both mRNA structure and amount. A sequence motif UACUAAC variably conserved in eukaryotic genomes, but some organisms more than one protein can recognize it. Here we show SF1 Quaking (QKI) compete for subset branchpoints with ACUAA. activates exon inclusion through this sequence, QKI represses alternatively spliced exons sequence. Using mutant reporters derived from natural two branchpoint-like sequences, find when either mutated, other used as branchpoint, are present, neither due to high affinity binding strong splicing repression by QKI. occupancy at dual site directly prevents subsequent recruitment spliceosome-associated factors. Finally, ectopic expression budding yeast (which lacks

Language: Английский

Mapping C. difficile TcdB interactions with host cell-surface and intracellular factors using proximity-dependent biotinylation labeling DOI Creative Commons

Jennifer S. Ward,

Karl J. Schreiber, John Tam

et al.

mBio, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 17, 2025

Many bacterial toxins exert their cytotoxic effects by enzymatically inactivating one or more cytosolic targets in host cells. To reach intracellular targets, these possess functional domains subdomains that interact with and exploit various factors biological processes. Despite great progress identifying many of the key involved uptake toxins, significant knowledge gaps remain as to how partially characterized newly discovered microbial processes intoxicate target Proximity-dependent biotinylation (e.g., BioID) is a powerful method identify nearby living cells, offering potential toxins. Here, we used BioID interrogate proximal interactors multi-domain Clostridioides difficile TcdB toxin. Expressed fusions TurboID different fragments identified several high-confidence proteins cytosol, including members Rho GTPase signaling network actin cytoskeletal network. Additionally, developed an extracellular proximity labeling using recombinant TurboID-toxin chimeras, which uncovered limited number cell-surface LRP1, was previously receptor TcdB. Our work reveals surface receptors components exploited highlighting vulnerabilities cells.IMPORTANCEBacterial are causative agents human diseases. Further characterizing intoxication mechanisms important for development vaccines treatments toxin-mediated disease. approaches offer orthogonal approach complement genetic screens. evaluate this host-toxin interactions on cell where toxin modifies essential targets. Critically, have highlighted limitations applied protein researchers weigh when considering technique exotoxin studies.

Language: Английский

Citations

1
In vivo HIV-1 nuclear condensates safeguard against cGAS and license reverse transcription DOI Creative Commons

Selen Ay,

Julien Burlaud‐Gaillard, Anastasia D. Gazi

et al.

The EMBO Journal, Journal Year: 2024, Volume and Issue: 44(1), P. 166 - 199

Published: Dec. 2, 2024

Language: Английский

Citations

3
Computational design and evaluation of optimal bait sets for scalable proximity proteomics DOI Creative Commons
Vesal Kasmaeifar,

Saya Sedighi,

Anne‐Claude Gingras

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 4, 2024

Abstract The spatial organization of proteins in eukaryotic cells can be explored by identifying nearby using proximity-dependent biotinylation approaches like BioID. BioID defines the localization thousands endogenous human when used on hundreds bait proteins. However, this high number restricts approach’s usage and gives these datasets limited scalability for context-dependent profiling. To make subcellular proteome mapping across different cell types conditions more practical cost-effective, we developed a comprehensive benchmarking platform multiple metrics to assess how well given subset reproduce an original dataset. We also introduce GENBAIT, which uses genetic algorithm optimize selection, derive subsets predicted retain structure coverage two large less than third baits. This flexible solution is poised improve intelligent selection baits contextual studies.

Language: Английский

Citations

0
An ancient competition for the conserved branchpoint sequence influences physiological and evolutionary outcomes in splicing DOI Creative Commons

Karen Larissa Pereira de Castro,

J.M. Abril, Kuo‐Chieh Liao

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 9, 2024

Recognition of the intron branchpoint during spliceosome assembly is a multistep process that defines both mRNA structure and amount. A sequence motif UACUAAC variably conserved in eukaryotic genomes, but some organisms more than one protein can recognize it. Here we show SF1 Quaking (QKI) compete for subset branchpoints with ACUAA. activates exon inclusion through this sequence, QKI represses alternatively spliced exons sequence. Using mutant reporters derived from natural two branchpoint-like sequences, find when either mutated, other used as branchpoint, are present, neither due to high affinity binding strong splicing repression by QKI. occupancy at dual site directly prevents subsequent recruitment spliceosome-associated factors. Finally, ectopic expression budding yeast (which lacks

Language: Английский

Citations

0