Biology subjects

Martinez-Seidel, F.

Publications and source records attributed to Martinez-Seidel, F..

5 recordsLinked to original sources

bioRxiv2025

RBP47 family members are negative regulators of heat stress tolerance in Arabidopsis thaliana

Stress granules (SGs) are liquid-liquid phase-separated condensates that sequester RNA, proteins, and metabolites to modulate cellular physiology under stress. In Arabidopsis, the RNA-binding protein RBP47b is a canonical SG marker, yet its functional contribution to thermotolerance remains unresolved. Here, we combined mTurboID proximity labeling with multi-omics profiling to define the RBP47b interactome and its physiological impact. mTurboID identified a stress-specific enrichment of 40S ribosomal subunits within RBP47b SGs, implicating these condensates in translational control. Surprisingly, quadruple mutant plants lacking all four RBP47 paralogues (rbp47abcc') displayed enhanced survival, attenuated growth delay, and faster recovery of photosynthetic efficiency after severe heat stress. Integrated transcriptome, proteome, and metabolome analyses revealed that this gain of thermotolerance is associated with (i) accelerated re-initiation of translation, (ii) constitutively elevated jasmonate and oxylipin pools, and (iii) reduced ROS accumulation during heat and recovery. We conclude that the RBP47 family acts as a negative regulator of heat tolerance by sequestering 40S subunits and limiting translational restart; loss of these SG scaffolds pre-primes jasmonate-dependent detoxification pathways and expedites proteome rebuilding, thereby conferring superior thermotolerance.

plant biology↗

bioRxiv2024

Comparative analysis of 43 distinct RNA modifications by nanopore tRNA sequencing

Transfer RNAs are the fundamental adapter molecules of protein synthesis and the most abundant and heterogeneous class of noncoding RNA molecules in cells. The study of tRNA repertoires remains challenging, complicated by the presence of dozens of post transcriptional modifications. Nanopore sequencing is an emerging technology with promise for both tRNA sequencing and the detection of RNA modifications; however, such studies have been limited by the throughput and accuracy of direct RNA sequencing methods. Moreover, detection of the complete set of tRNA modifications by nanopore sequencing remains challenging. Here we show that recent updates to nanopore direct RNA sequencing chemistry (RNA004) combined with our own optimizations to tRNA sequencing protocols and analysis workflows enable high throughput coverage of tRNA molecules and characterization of nanopore signals produced by 43 distinct RNA modifications. We share best practices and protocols for nanopore sequencing of tRNA and further report successful detection of low abundance mitochondrial and viral tRNAs, providing proof of concept for use of nanopore sequencing to study tRNA populations in the context of infection and organelle biology. This work provides a roadmap to guide future efforts towards de novo detection of RNA modifications across multiple organisms using nanopore sequencing.

molecular biology↗

bioRxiv2023

Guidelines for performing ribosome profiling in plants including structural analysis of rRNA fragments

Ribosome profiling (or Ribo-seq) is a technique that provides genome-wide information on the translational landscape (translatome). Across different plant studies, variable methodological setups have been described which raises questions about the general comparability of data that were generated from diverging methodologies. Furthermore, a common problem when performing Ribo-seq are abundant rRNA fragments that are wastefully incorporated into the libraries and dramatically reduce sequencing depth. To remove these rRNA contaminants, it is common to perform preliminary trials to identify these fragments because they are thought to vary depending on nuclease treatment, tissue source, and plant species. Here, we compile valuable insights gathered over years of generating Ribo-seq datasets from different species and experimental setups. We highlight which technical steps are important for maintaining cross experiment comparability and describe a highly efficient approach for rRNA removal. Furthermore, we provide evidence that many rRNA fragments are structurally preserved over diverse nuclease regimes, as well as across plant species. Using a recently published cryo-electron microscopy (cryo-EM) structure of the tobacco 80S ribosome, we show that the most abundant rRNA fragments are spatially derived from the solvent-exposed surface of the ribosome. The guidelines presented here shall aid newcomers in establishing ribosome profiling in new plant species and provide insights that will help in customizing the methodology for individual research goals.

plant biology↗

bioRxiv2022

Remodelled Ribosomes Synthesise a Specific Proteome in Proliferating Plant Tissue during Cold

Plant acclimation to low temperatures occurs through system-wide mechanisms that include proteome shifts, some of which occur at the level of protein synthesis. All proteins are synthesised by ribosomes. Rather than being monolithic, transcript-to-protein translation machines, ribosomes can be selective and cause effective proteome shifts required for successful temperature acclimation. Here, we use apical root meristems of germinating seedlings of the monocotyledonous plant barley as a model to study changes in protein abundance and synthesis rates during cold acclimation. We measure metabolic and physiological parameters that allow us to compare protein synthesis rates in different physiological states, e.g., in cold acclimation compared to the optimal temperature state. We show that ribosomal proteins are independently synthesised and assembled into ribosomal complexes in root proliferative tissue, and assess how the ribo-proteome shifts during cold may be associated with changes in synthesis and accumulation of macromolecular complexes. We demonstrate that translation initiation is the limiting step during cold acclimation and based on our data propose a model of a ribosomal code that depends on a reconfigured ribosome population, where as a mode of cold acclimation, specific ribosomal protein compositions may confer selective association capabilities between 60S subunits and 48S initiation complexes.

plant biology↗

bioRxiv2020

Arabidopsis REI-LIKE proteins activate ribosome biogenesis during cold acclimation

Arabidopsis REIL proteins are cytosolic ribosomal 60S-biogenesis factors. After shift to 10{degrees}C, reil mutants deplete and slowly replenish non-translating eukaryotic ribosome complexes of root tissue, while tightly controlling the balance of non-translating 40S- and 60S-subunits. Reil mutations compensate by hyper-accumulation of non-translating subunits at steady-state temperature; after cold-shift, a KCl-sensitive 80S sub-fraction remains depleted. We infer that Arabidopsis buffers fluctuating translation by pre-existing non-translating ribosomes before de novo synthesis meets temperature-induced demands. Reil1 reil2 double mutants accumulate 43S-preinitiation and pre-60S-maturation complexes and have altered paralog composition of ribosomal proteins in non-translating complexes. With few exceptions, e.g. RPL3B and RPL24C, these changes are not under transcriptional control. Our study suggests requirement of de novo synthesis of eukaryotic ribosomes for long-term cold acclimation, feedback control of NUC2 and eIF3C2 transcription and links new proteins, AT1G03250, AT5G60530, to plant ribosome biogenesis. We propose that Arabidopsis requires biosynthesis of specialized ribosomes for cold acclimation. Highlight of this studyREIL proteins affect paralog composition of eukaryotic ribosomes and suppress accumulation of 43S-preinitiation and pre-60S-maturation complexes, suggesting functions of ribosome heterogeneity and biogenesis in plant cold acclimation.

plant biology↗