Accumulation Of Ribosomes Research Articles

The ubiquitin-proteasome system regulates the formation of specialized ribosomes during high salt stress in yeast.

Rps26-deficient ribosomes are a physiologically relevant ribosome population which arises during osmotic stress to support the translation of mRNAs involved in the response to high salt in yeast. They are formed by binding of the chaperone Tsr2 to fully assembled ribosomes to release Rps26 when intracellular Na+ concentrations rise. Tsr2-mediated Rps26 release is reversible, enabling a rapid response that conserves ribosomes. However, because the concentration of Tsr2 relative to ribosomes is low, how the released Rps26•Tsr2 complex is managed to allow for accumulation of Rps26-deficient ribosomes to nearly 50% of all ribosomes remains unclear. Here we show that released Rps26 is degraded via the Pro/N-degron pathway, enabling the accumulation of Rps26-deficient ribosomes. Substitution of the N-terminal proline of Rps26 to serine increases the stability of free Rps26, limits the accumulation of Rps26-deficient ribosomes and renders yeast sensitive to high salt. The GID-complex, an E3 ubiquitin ligase, and its adaptor Gid4, mediate polyubiquitination of Rps26 at Lys66 and Lys70. Moreover, this ubiquitination event is required for Rps26 degradation, the accumulation of Rps26-deficient ribosomes and the high salt stress resistance. Together, the data show that targeted degradation of released Rps26 from the Rps26•Tsr2 complex allows Tsr2 to be recycled, thus facilitating multiple rounds of Rps26 release.

Metabolism-dependent secondary effect of anti-MAPK cancer therapy on DNA repair.

Amino acid bioavailability impacts mRNA translation in a codon-dependent manner. Here, we report that the anti-cancer MAPK inhibitors (MAPKi) decrease the intracellular concentration of aspartate and glutamate in melanoma cells. This coincides with the accumulation of ribosomes on codons corresponding to these amino acids and triggers the translation-dependent degradation of mRNAs encoding aspartate- and glutamate-rich proteins, involved in DNA metabolism such as DNA replication and repair. Consequently, cells that survive MAPKi degrade aspartate and glutamate likely to generate energy, which simultaneously decreases their requirement for amino acids due to the downregulation of aspartate- and glutamate-rich proteins involved in cell proliferation. Concomitantly, the downregulation of aspartate- and glutamate-rich proteins involved in DNA repair increases DNA damage loads. Thus, DNA repair defects, and therefore mutations, are at least in part a secondary effect of the metabolic adaptation of cells exposed to MAPKi.

Mitochondrial structural alterations in fibromyalgia: a pilot electron microscopy study.

The pathogenesis of fibromyalgia (FM), characterised by chronic widespread pain and fatigue, remains notoriously elusive, hampering attempts to develop disease modifying treatments. Mitochondria are the headquarters of cellular energy metabolism, and their malfunction has been proposed to contribute to both FM and chronic fatigue. Thus, the aim of the current pilot study, was to detect structural changes in mitochondria of peripheral blood mononuclear cells (PBMCs) of FM patients, using transmission electron microscopy (TEM). To detect structural mitochondrial alterations in FM, we analysed PBMCs from seven patients and seven healthy controls, using TEM. Patients were recruited from a specialised Fibromyalgia Clinic at a tertiary medical centre. After providing informed consent, participants completed questionnaires including the widespread pain index (WPI), symptoms severity score (SSS), fibromyalgia impact questionnaire (FIQ), beck depression inventory (BDI), and visual analogue scale (VAS), to verify a diagnosis of FM according to ACR criteria. Subsequently, blood samples were drawn and PBMCs were collected for EM analysis. TEM analysis of PBMCs showed several distinct mitochondrial cristae patterns, including total loss of cristae in FM patients. The number of mitochondria with intact cristae morphology was reduced in FM patients and the percentage of mitochondria that completely lacked cristae was increased. These results correlated with the WPI severity. Moreover, in the FM patient samples we observed a high percentage of cells containing electron dense aggregates, which are possibly ribosome aggregates. Cristae loss and possible ribosome aggregation were intercorrelated, and thus may represent reactions to a shared cellular stress condition. The changes in mitochondrial morphology suggest that mitochondrial dysfunction, resulting in inefficient oxidative phosphorylation and ATP production, metabolic and redox disorders, and increased reactive oxygen species (ROS) levels, may play a pathogenetic role in FM. We describe novel morphological changes in mitochondria of FM patients, including loss of mitochondrial cristae. While these observations cannot determine whether the changes are pathogenetic or represent an epiphenomenon, they highlight the possibility that mitochondrial malfunction may play a causative role in the cascade of events leading to chronic pain and fatigue in FM. Moreover, the results offer the possibility of utilising changes in mitochondrial morphology as an objective biomarker in FM. Further understanding the connection between FM and dysfunction of mitochondria physiology, may assist in developing both novel diagnostic tools as well as specific treatments for FM, such as approaches to improve/strengthen mitochondria function.

Glucose ingestion before and after resistance training sessions does not augment ribosome biogenesis in healthy moderately trained young adults

PurposeResistance training-induced skeletal muscle hypertrophy seems to depend on ribosome biogenesis and content. High glucose treatment may augment ribosome biogenesis through potentiating resistance training-induced adaptations. This was investigated with total RNA and ribosomal RNA abundances as main outcomes, with relevant transcriptional/translational regulators (c-Myc/UBF/rpS6) as a secondary outcome.MethodsSixteen healthy, moderately trained individuals [male/female, n = 9/7; age, 24.1 (3.3)] participated in a within-participant crossover trial with unilateral resistance training (leg press and knee extension, 3 sets of 10 repetitions maximum) and pre- and post-exercise ingestion of either glucose (3 × 30 g, 90 g total) or placebo supplements (Stevia rebaudiana, 3 × 0.3 g, 0.9 g total), together with protein (2 × 25 g, 50 g total), on alternating days for 12 days. Six morning resistance exercise sessions were conducted per condition, and the sessions were performed in an otherwise fasted state. Micro-biopsies were sampled from m. vastus lateralis before and after the intervention.ResultsGlucose ingestion did not have beneficial effects on resistance training-induced increases of ribosomal content (mean difference 7.6% [− 7.2, 24.9], p = 0.34; ribosomal RNA, 47S/18S/28S/5.8S/5S, range 7.6–37.9%, p = 0.40–0.98) or levels of relevant transcriptional or translational regulators (c-MYK/UBF/rpS6, p = 0.094–0.292). Of note, both baseline and trained state data of total RNA showed a linear relationship with UBF; a ∼14% increase in total RNA corresponded to 1 SD unit increase in UBF (p = 0.003).ConclusionGlucose ingestion before and after resistance training sessions did not augment ribosomal RNA accumulation during twelve days of heavy-load resistance training in moderately trained young adults.

Ultrastructure of the bacteriome and bacterial symbionts in the Asian citrus psyllid, Diaphorina citri.

Omics analyses suggested a mutually indispensable tripartite association among the host D. citri and organelle-like bacteriome associates, Carsonella and Profftella, which are vertically transmitted through host generations. This relationship is based on the metabolic complementarity among these organisms, which is partly enabled by horizontal gene transfer between partners. However, little was known about the fine morphology of the symbionts and the bacteriome, the interface among these organisms. As a first step to address this issue, the present study performed transmission electron microscopy, which revealed previously unrecognized ultrastructures, including aggregations of ribosomes in Carsonella, numerous tubes and occasional protrusions of Profftella, apparently degrading Profftella, and host organelles with different abundance and morphology in distinct cell types. These findings provide insights into the behaviors of the symbionts and host cells to maintain the symbiotic relationship in D. citri.

Senescence in yeast is associated with amplified linear fragments of chromosome XII rather than ribosomal DNA circle accumulation.

The massive accumulation of extrachromosomal ribosomal DNA circles (ERCs) in yeast mother cells has been long cited as the primary driver of replicative ageing. ERCs arise through ribosomal DNA (rDNA) recombination, and a wealth of genetic data connects rDNA instability events giving rise to ERCs with shortened life span and other ageing pathologies. However, we understand little about the molecular effects of ERC accumulation. Here, we studied ageing in the presence and absence of ERCs, and unexpectedly found no evidence of gene expression differences that might indicate stress responses or metabolic feedback caused by ERCs. Neither did we observe any global change in the widespread disruption of gene expression that accompanies yeast ageing, altogether suggesting that ERCs are largely inert. Much of the differential gene expression that accompanies ageing in yeast was actually associated with markers of the senescence entry point (SEP), showing that senescence, rather than age, underlies these changes. Cells passed the SEP irrespective of ERCs, but we found the SEP to be associated with copy number amplification of a region of chromosome XII between the rDNA and the telomere (ChrXIIr) forming linear fragments up to approximately 1.8 Mb size, which arise in aged cells due to rDNA instability but through a different mechanism to ERCs. Therefore, although rDNA copy number increases dramatically with age due to ERC accumulation, our findings implicate ChrXIIr, rather than ERCs, as the primary driver of senescence during budding yeast ageing.

Spatial resolution of cellular senescence dynamics in human colorectal liver metastasis.

Hepatic metastasis is a clinical challenge for colorectal cancer (CRC). Senescent cancer cells accumulate in CRC favoring tumor dissemination. Whether this mechanism progresses also in metastasis is unexplored. Here, we integrated spatial transcriptomics, 3D-microscopy, and multicellular transcriptomics to study the role of cellular senescence in human colorectal liver metastasis (CRLM). We discovered two distinct senescent metastatic cancer cell (SMCC) subtypes, transcriptionally located at the opposite pole of epithelial (e) to mesenchymal (m) transition. SMCCs differ in chemotherapy susceptibility, biological program, and prognostic roles. Mechanistically, epithelial (e)SMCC initiation relies on nucleolar stress, whereby c-myc dependent oncogene hyperactivation induces ribosomal RPL11 accumulation and DNA damage response. In a 2D pre-clinical model, we demonstrated that RPL11 co-localized with HDM2, a p53-specific ubiquitin ligase, leading to senescence activation in (e)SMCCs. On the contrary, mesenchymal (m)SMCCs undergo TGFβ paracrine activation of NOX4-p15 effectors. SMCCs display opposing effects also in the immune regulation of neighboring cells, establishing an immunosuppressive environment or leading to an active immune workflow. Both SMCC signatures are predictive biomarkers whose unbalanced ratio determined the clinical outcome in CRLM and CRC patients. Altogether, we provide a comprehensive new understanding of the role of SMCCs in CRLM and highlight their potential as new therapeutic targets to limit CRLM progression.

Peanut AhmTERF1 Regulates Root Growth by Modulating Mitochondrial Abundance.

Mitochondria are responsible for energy generation, as well as key metabolic and signaling pathways, and thus affect the entire developmental process of plants as well as their responses to stress. In metazoans, mitochondrial transcription termination factors (mTERFs) are known to regulate mitochondrial transcription. mTERFs have also been discovered in plants, but only a few of these proteins have been explored for their biological functions. Here, we report a role in root growth for mitochondria-associated protein AhmTERF1 in peanut (Arachis hypogaea L.). Overexpressing AhmTERF1 significantly stimulated the growth of peanut hairy roots and transgenic Arabidopsis. Surprisingly, AhmTERF1 is predominantly expressed in the root meristem where it increases mitochondrial abundance. AhmTERF1 binding to mtDNA was enriched in the RRN18 and RRN26 regions, suggesting it is related to the accumulation of mitochondrial ribosomes. Peanut is one of the main oil crops and the important source of edible oil and AhmTERF1 likely affects agronomic traits related to root growth in different peanut cultivars. We propose that peanut AhmTERF1 is an important protein for root growth due to its role in regulating mitochondrial abundance.

Amplification of Amyloid Protein-induced Aggregation of the Eukaryotic Ribosome.

Alzheimer's disease (AD) is characterized by the aggregation of Tau protein and Amyloid-β peptides (Aβ 1-40 and Aβ 1-42). A loss of ribosomal population is also observed in the neurons in affected regions of AD. Our studies demonstrated that in vitro aggregation of amyloid forming proteins, Aβ peptides and Tau protein variants (AFPs), in the vicinity of yeast 80S ribosome can induce co-aggregation of ribosomal components. In this study, the ability of minute quantities of AFP-ribosome co-aggregates to seed the aggregation of a large excess of untreated 80S ribosomes was explored. The AFPs were purified using ion-exchange chromatography. Seeded aggregation of ribosomes in the presence of minute quantities of ribosome-protein co-aggregates or ribosomal components was studied using agarose gel electrophoretic and SDS-PAGE analysis of the pellets and Sucrose Density Gradient centrifugation of the supernatant obtained after centrifugation of the aggregation reaction mixture. Our studies, therefore, demonstrate that minute quantities of AFP-80S co-aggregate have significant seeding potential and could lead to aggregation of a large excess of fresh 80S ribosomes and this seeding ability is sustained over multiple cycles of ribosome aggregation. The aggregation propensity of ribosomal components alone could contribute towards the seeding of ribosome aggregation. The ability of minute quantities of AFP-80S co-aggregates to seed the aggregation of a large excess of fresh 80S ribosomes would result in the loss of global ribosomal population in Alzheimer's disease afflicted neurons. Hence, subject to further validation by in vivo studies, our in vitro studies indicate a significant mode of toxicity of amyloid aggregates that might be important in Alzheimer's disease pathology.

A noncanonical RNA-binding domain of the fragile X protein, FMRP, elicits translational repression independent of mRNA G-quadruplexes

Loss of functional fragile X mental retardation protein (FMRP) causes fragile X syndrome, the leading form of inherited intellectual disability and the most common monogenic cause of autism spectrum disorders. FMRP is an RNA-binding protein that controls neuronal mRNA localization and translation. FMRP is thought to inhibit translation elongation after being recruited to target transcripts via binding RNA G-quadruplexes (G4s) within the coding sequence. Here, we directly test this model and report that FMRP inhibits translation independent of mRNA G4s. Furthermore, we found that the RGG box motif together with its natural C-terminal domain forms a noncanonical RNA-binding domain (ncRBD) that is essential for translational repression. The ncRBD elicits broad RNA-binding ability and binds to multiple reporter mRNAs and all four homopolymeric RNAs. Serial deletion analysis of the ncRBD identified that the regions required for mRNA binding and translational repression overlap but are not identical. Consistent with FMRP stalling elongating ribosomes and causing the accumulation of slowed 80S ribosomes, transcripts bound by FMRP via the ncRBD cosediment with heavier polysomes and were present in puromycin-resistant ribosome complexes. Together, this work identifies a ncRBD and translational repression domain that shifts our understanding of how FMRP inhibits translation independent of mRNA G4s.

Ribosome accumulation during early phase resistance training in humans.

AimTo describe ribosome biogenesis during resistance training, its relation to training volume and muscle growth.MethodsA training group (n = 11) performed 12 sessions (3‐4 sessions per week) of unilateral knee extension with constant and variable volume (6 and 3‐9 sets per session respectively) allocated to either leg. Ribosome abundance and biogenesis markers were assessed from vastus lateralis biopsies obtained at baseline, 48 hours after sessions 1, 4, 5, 8, 9 and 12, and after eight days of de‐training, and from a control group (n = 8). Muscle thickness was measured before and after the intervention.ResultsTraining led to muscle growth (3.9% over baseline values, 95% CrI: [0.2, 7.5] vs. control) with concomitant increases in total RNA, ribosomal RNA, upstream binding factor (UBF) and ribosomal protein S6 with no differences between volume conditions. Total RNA increased rapidly in response to the first four sessions (8.6% [5.6, 11.7] per session), followed by a plateau and peak values after session 8 (49.5% [34.5, 66.5] above baseline). Total RNA abundance was associated with UBF protein levels (5.0% [0.2, 10.2] per unit UBF), and the rate of increase in total RNA levels predicted hypertrophy (0.3 mm [0.1, 0.4] per %‐point increase in total RNA per session). After de‐training, total RNA decreased (−19.3% [−29.0, −8.1]) without muscle mass changes indicating halted biosynthesis of ribosomes.ConclusionRibosomes accumulate in the initial phase of resistance training with abundances sensitive to training cessation and associated with UBF protein levels. The average accumulation rate predicts muscle training‐induced hypertrophy.

Rebirth of the translational machinery: The importance of recycling ribosomes.

Translation of the genetic code occurs in a cycle where ribosomes engage mRNAs, synthesize protein, and then disengage in order to repeat the process again. The final part of this process-ribosome recycling, where ribosomes dissociate from mRNAs-involves a complex molecular choreography of specific protein factors to remove the large and small subunits of the ribosome in a coordinated fashion. Errors in this process can lead to the accumulation of ribosomes at stop codons or translation of downstream open reading frames (ORFs). Ribosome recycling is also critical when a ribosome stalls during the elongation phase of translation and must be rescued to allow continued translation of the mRNA. Here we discuss the molecular interactions that drive ribosome recycling, and their regulation in the cell. We also examine the consequences of inefficient recycling with regards to disease, and its functional roles in synthesis of novel peptides, regulation of gene expression, and control of mRNA-associated proteins. Alterations in ribosome recycling efficiency have the potential to impact many cellular functions but additional work is needed to understand how this regulatory power is utilized.

Stromal Protein Chloroplast Development and Biogenesis1 Is Essential for Chloroplast Development and Biogenesis in Arabidopsis thaliana.

Although numerous studies have been carried out on chloroplast development and biogenesis, the underlying regulatory mechanisms are still largely elusive. Here, we characterized a chloroplast stromal protein Chloroplast Development and Biogenesis1 (CDB1). The knockout cdb1 mutant exhibits a seedling-lethal and ivory leaf phenotype. Immunoblot and RNA blot analyses show that accumulation of chloroplast ribosomes is compromised in cdb1, resulting in an almost complete loss of plastid-encoded proteins including the core subunits of the plastid-encoded RNA polymerase (PEP) RpoB and RpoC2, and therefore in impaired PEP activity. Orthologs of CDB1 are found in green algae and land plants. Moreover, a protein shows high similarity with CDB1, designated as CDB1-Like (CDB1L), is present in angiosperms. Absence of CDB1L results in impaired embryo development. While CDB1 is specifically located in the chloroplast stroma, CDB1L is localized in both chloroplasts and mitochondria in Arabidopsis. Thus, our results demonstrate that CDB1 is indispensable for chloroplast development and biogenesis through its involvement in chloroplast ribosome assembly whereas CDB1L may fulfill a similar function in both mitochondria and chloroplasts.

Fast and global reorganization of the chloroplast protein biogenesis network during heat acclimation.

Photosynthesis is a central determinant of plant biomass production, but its homeostasis is increasingly challenged by heat. Little is known about the sensitive regulatory principles involved in heat acclimation that underly the biogenesis and repair of chloroplast-encoded core subunits of photosynthetic complexes. Employing time-resolved ribosome and transcript profiling together with selective ribosome proteomics, we systematically deciphered these processes in chloroplasts of Chlamydomonas reinhardtii. We revealed protein biosynthesis and altered translation elongation as central processes for heat acclimation and showed that these principles are conserved between the alga and the flowering plant Nicotiana tabacum. Short-term heat exposure resulted in specific translational repression of chlorophyll a-containing core antenna proteins of photosystems I and II. Furthermore, translocation of ribosome nascent chain complexes to thylakoid membranes was affected, as reflected by the increased accumulation of stromal cpSRP54-bound ribosomes. The successful recovery of synthesizing these proteins under prolonged acclimation of nonlethal heat conditions was associated with specific changes of the co-translational protein interaction network, including increased ribosome association of chlorophyll biogenesis enzymes and acclimation factors responsible for complex assembly. We hypothesize that co-translational cofactor binding and targeting might be bottlenecks under heat but become optimized upon heat acclimation to sustain correct co-translational protein complex assembly.

Translation inhibition and suppression of stress granules formation by cisplatin.

Platinum-based antineoplastic drugs, such as cisplatin, are commonly used to induce tumor cell death. Cisplatin is believed to induce apoptosis as a result of cisplatin-DNA adducts that inhibit DNA and RNA synthesis. Although idea that DNA damage underlines anti-proliferative effects of cisplatin is dominant in cancer research, there is a poor correlation between the degree of the cell sensitivity to cisplatin and the extent of DNA platination. Here, we examined possible effects of cisplatin on post-transcriptional gene regulation that may contribute to cisplatin-mediated cytotoxicity. We show that cisplatin suppresses formation of stress granules (SGs), pro-survival RNA granules with multiple roles in cellular metabolism. Mechanistically, cisplatin inhibits cellular translation to promote disassembly of polysomes and aggregation of ribosomal subunits. As SGs are in equilibrium with polysomes, cisplatin-induced shift towards ribosomal aggregation suppresses SG formation. Our data uncover previously unknown effects of cisplatin on RNA metabolism.

Nucleolar localization of c‐Jun

Nucleoli are well defined for their function in ribosome biogenesis, but only a small fraction of the nucleolar proteome has been characterized. Here, we report that the proto-oncogene, c-Jun, is targeted to the nucleolus. Using live cell imaging in myogenic cells, we document that the c-Jun basic domain contains a unique, evolutionarily conserved motif that determines nucleolar targeting. Fos family Jun dimer partners, such as Fra2, while nuclear, do not co-localize with c-Jun in the nucleolus. A point mutation in c-Jun that mimics Fra2 (M260E) in its Nucleolar Localization sequence (NoLS) results in loss of c-Jun nucleolar targeting while still preserving nuclear localization. Fra2 can sequester c-Jun in the nucleoplasm, indicating that the stoichiometric ratio of heterodimeric partners regulates c-Jun nucleolar targeting. Finally, nucleolar localization of c-Jun modulates nucleolar architecture and ribosomal RNA accumulation. These studies highlight a novel role for Jun family proteins in the nucleolus, having potential implications for a diverse array of AP-1-regulated cellular processes.

Implementation of a Non-Thermal Atmospheric Pressure Plasma for Eradication of Plant Pathogens from a Surface of Economically Important Seeds.

Plant pathogenic bacteria cause significant economic losses in the global food production sector. To secure an adequate amount of high-quality nutrition for the growing human population, novel approaches need to be undertaken to combat plant disease-causing agents. As the currently available methods to eliminate bacterial phytopathogens are scarce, we evaluated the effectiveness and mechanism of action of a non-thermal atmospheric pressure plasma (NTAPP). It was ignited from a dielectric barrier discharge (DBD) operation in a plasma pencil, and applied for the first time for eradication of Dickeya and Pectobacterium spp., inoculated either on glass spheres or mung bean seeds. Furthermore, the impact of the DBD exposure on mung bean seeds germination and seedlings growth was estimated. The observed bacterial inactivation rates exceeded 3.07 logs. The two-minute DBD exposure stimulated by 3–4% the germination rate of mung bean seeds and by 13.4% subsequent early growth of the seedlings. On the contrary, a detrimental action of the four-minute DBD subjection on seed germination and early growth of the sprouts was noted shortly after the treatment. However, this effect was no longer observed or reduced to 9.7% after the 96 h incubation period. Due to the application of optical emission spectrometry (OES), transmission electron microscopy (TEM), and confocal laser scanning microscopy (CLSM), we found that the generated reactive oxygen and nitrogen species (RONS), i.e., N2, N2+, NO, OH, NH, and O, probably led to the denaturation and aggregation of DNA, proteins, and ribosomes. Furthermore, the cellular membrane disrupted, leading to an outflow of the cytoplasm from the DBD-exposed cells. This study suggests the potential applicability of NTAPPs as eco-friendly and innovative plant protection methods.

Not4 and Not5 modulate translation elongation by Rps7A ubiquitination, Rli1 moonlighting, and condensates that exclude eIF5A.

In this work, we show that Not4 and Not5 from the Ccr4-Not complex modulate translation elongation dynamics and change ribosome A-site dwelling occupancy in a codon-dependent fashion. These codon-specific changes in not5Δ cells are very robust and independent of codon position within the mRNA, the overall mRNA codon composition, or changes of mRNA expression levels. They inversely correlate with codon-specific changes in cells depleted for eIF5A and positively correlate with those in cells depleted for ribosome-recycling factor Rli1. Not5 resides in punctate loci, co-purifies with ribosomes and Rli1, but not with eIF5A, and limits mRNA solubility. Overexpression of wild-type or non-complementing Rli1 and loss of Rps7A ubiquitination enable Not4 E3 ligase-dependent translation of polyarginine stretches. We propose that Not4 and Not5 modulate translation elongation dynamics to produce a soluble proteome by Rps7A ubiquitination, dynamic condensates that limit mRNA solubility and exclude eIF5A, and a moonlighting function of Rli1.

Membrane-Enriched Proteomics Link Ribosome Accumulation and Proteome Reprogramming With Cold Acclimation in Barley Root Meristems.

Due to their sessile nature, plants rely on root systems to mediate many biotic and abiotic cues. To overcome these challenges, the root proteome is shaped to specific responses. Proteome-wide reprogramming events are magnified in meristems due to their active protein production. Using meristems as a test system, here, we study the major rewiring that plants undergo during cold acclimation. We performed tandem mass tag-based bottom-up quantitative proteomics of two consecutive segments of barley seminal root apexes subjected to suboptimal temperatures. After comparing changes in total and ribosomal protein (RP) fraction-enriched contents with shifts in individual protein abundances, we report ribosome accumulation accompanied by an intricate translational reprogramming in the distal apex zone. Reprogramming ranges from increases in ribosome biogenesis to protein folding factors and suggests roles for cold-specific RP paralogs. Ribosome biogenesis is the largest cellular investment; thus, the vast accumulation of ribosomes and specific translation-related proteins during cold acclimation could imply a divergent ribosomal population that would lead to a proteome shift across the root. Consequently, beyond the translational reprogramming, we report a proteome rewiring. First, triggered protein accumulation includes spliceosome activity in the root tip and a ubiquitous upregulation of glutathione production and S-glutathionylation (S-GSH) assemblage machineries in both root zones. Second, triggered protein depletion includes intrinsically enriched proteins in the tip-adjacent zone, which comprise the plant immune system. In summary, ribosome and translation-related protein accumulation happens concomitantly to a proteome reprogramming in barley root meristems during cold acclimation. The cold-accumulated proteome is functionally implicated in feedbacking transcript to protein translation at both ends and could guide cold acclimation.

PPR protein Early Chloroplast Development 2 is essential for chloroplast development at the early stage of Arabidopsis development

Chloroplast biogenesis and development regulation have long been a focus of research; however, the underlying mechanisms of these processes have not yet been fully elucidated. Pentatricopeptide repeat (PPR) proteins have been shown to play key roles in chloroplast development. Here, we identified a novel P-type PPR protein, Early Chloroplast Development 2 (ECD2), and the ecd2 mutant resulted in embryo lethality. The RNAi lines of ECD2 showed varying degrees of albino cotyledons and abnormal chloroplast development, but true leaves were similar to the wild-type. Further analysis revealed that ECD2 was responsible for chloroplast gene expression and group II intron splicing of several genes. Transcriptome analysis combined with quantitative real-time PCR showed that ECD2 was associated with the expression of ribosomal genes and accumulation of chloroplast ribosomes. Overall, our results indicate that ECD2 is critically important for early chloroplast development in cotyledon.