Biology subjects

Hochstoeger, T.

Publications and source records attributed to Hochstoeger, T..

6 recordsLinked to original sources

bioRxiv2025

Quantitative comparison of methodologies for translation site imaging in living cells

Single-molecule imaging of translation sites in living cells has enabled the dynamics of protein synthesis to be investigated with high spatial and temporal resolution. These methodologies utilize the interaction between a multimerized epitope tag and its cognate fluorescent nanobody to detect nascent polypeptides as they emerge from the ribosome. Here, we present a systematic comparison of current methodologies and determine that the ALFA-tag reduces perturbations of mRNA expression and increases the fluorescent signal of translation sites.

biophysics↗

bioRxiv2023

Distinct roles of LARP1 and 4EBP1/2 in regulating translation and stability of 5'TOP mRNAs

A central mechanism of mTOR complex 1 (mTORC1) signaling is the coordinated translation of ribosomal protein and translation factor mRNAs mediated by the 5'-terminal oligopyrimidine motif (5'TOP). Recently, La-related protein 1 (LARP1) has been proposed to be the specific regulator of 5'TOP mRNA translation downstream of mTORC1, while eIF4E-binding proteins (4EBP1/2) were suggested to have a general role in repression. Here, we employ single-molecule translation site imaging of 5'TOP and canonical mRNAs to study the translational dynamics of single mRNAs in living cells. Our data reveals that 4EBP1/2 has a dominant role in translation repression of both 5'TOP and canonical mRNAs during pharmacological inhibition of mTOR. In contrast, we find that LARP1 selectively protects 5'TOP mRNAs from degradation in a transcriptome-wide analysis of mRNA half-lives. Our results clarify the roles of 4EBP1/2 and LARP1 in regulating 5'TOP mRNAs and provides a framework to further study how these factors control cell growth during development and disease.

molecular biology↗

bioRxiv2021

Live imaging of the co-translational recruitment of XBP1 mRNA to the ER and its processing by diffuse, non-polarized IRE1a.

Endoplasmic reticulum (ER) to nucleus homeostatic signalling, known as the unfolded protein response (UPR), relies on the non-canonical splicing of XBP1 mRNA. The molecular switch that initiates splicing is the oligomerization of the ER stress sensor and UPR endonuclease IRE1. While IRE1 can form large clusters that have been proposed to function as XBP1 processing centers on the ER, the actual oligomeric state of active IRE1 complexes as well as the targeting mechanism that recruits XBP1 to IRE1 oligomers, remain unknown. Here, we used a single molecule imaging approach to directly monitor the recruitment of individual XBP1 transcripts to the ER surface. We confirmed that stable ER association of unspliced XBP1 mRNA is established through HR2-dependent targeting and relies on active translation. In addition, we show that IRE1-catalyzed splicing mobilizes XBP1 mRNA from the ER membrane in response to ER stress. Surprisingly, we find that XBP1 transcripts are not recruited into large IRE1 clusters, which only assemble upon overexpression of fluorescently-tagged IRE1 during ER stress. Our findings support a model where ribosome-engaged, ER-poised XBP1 mRNA is processed by functional IRE1 assemblies that are homogenously distributed throughout the ER membrane.

cell biology↗

bioRxiv2018

Revised testing procedures do not elicit magnetoreceptive behavior in C. elegans

A diverse range of species is known to rely on the Earths magnetic field for spatial information. Vidal-Gadea et al. claimed that C. elegans are magneto-sensitive, exploiting the magnetic field to guide their burrowing behavior [1]. Our attempts to replicate their findings were unsuccessful [2], which Vidal-Gadea attributed to the satiety of the animals and the environment in which they were raised. Here, we report our repeated experiments, having adopted several suggestions proposed by Vidal-Gadea et al. [3]. We find that shortening the length of the behavioral assay and raising the animals in a Faraday cage does not result in magnetotactic behavior. We reluctantly conclude that the assays employed by Vidal-Gadea are not robust or C. elegans are not magneto-sensitive.

animal behavior and cognition↗

bioRxiv2017

Are Caenorhabditis elegans magnetoreceptive?

A diverse array of species on the planet employ the Earths magnetic field as a navigational aid. As the majority of these animals are migratory, their utility to interrogate the molecular and cellular basis of the magnetic sense is limited. Vidal-Gadea and colleagues recently argued that the worm C. elegans possesses a magnetic sense that guides their vertical movement in soil. In making this claim they relied on three different behavioural assays that involved magnetic stimuli. Here, we set out to replicate their results employing blinded protocols and double wrapped coils that control for heat generation. We find no evidence supporting the existence of a magnetic sense in C. elegans. We further show that the Vidal-Gadea hypothesis is problematic as the adoption of a correction angle and a fixed trajectory relative to the Earths magnetic inclination does not necessarily result in vertical movement.

neuroscience↗

bioRxiv2016

ISCA1 and CRY4: An improbable proposition

This manuscript was submitted to Nature Materials on December 15th 2015.\n\nIn their recent manuscript Xie and colleagues argue that the pigeon ISCA1/CRY4 complex acts as a magnetic protein biocompass having unambiguously proved that these two proteins interact forming a complex with unique biophysical features (1). We do not think the evidence presented in the manuscript supports this conclusion. Firstly, the authors provide no data (such as a reciprocal co-immunoprecipitation from retinal tissue) that demonstrates that ISCA1 and CRY4 interact in vivo, instead the conclusions are based primarily on overexpression in E.coli and in vitro reconstitution experiments. Secondly, their experiments have not employed the correct full length version of CRY4. The CRY4 protein expressed by Xie and colleagues was only 497 amino acids in length. We extracted mRNA from the pigeon retina (Invitrogen, 610.11), generated cDNA (Clontech, 121311), and performed rap ...

animal behavior and cognition↗