Retrograde Communication Research Articles

Resveratrol impinges on retrograde communication without inducing mitochondrial biogenesis in aged rat soleus muscle

The natural polyphenol resveratrol (RSV) might counteract the skeletal muscle age-related loss of muscle mass and strength/function partly acting on mitochondria. This work analysed the effects of a six-week administration of RSV (50 mg/kg/day) in the oxidative Soleus (Sol) skeletal muscle of old rats (27 months old). RSV effects on key mitochondrial biogenesis proteins led to un unchanged amount of SIRT1 protein and a marked decrease (60 %) in PGC-1α protein. In addition, Peroxyredoxin 3 (PRXIII) protein decreased by 50 %, which on overall suggested the absence of induction of mitochondrial biogenesis by RSV in old Sol. A novel direct correlation between PGC-1α and PRXIII proteins was demonstrated by correlation analysis in RSV and ad-libitum (AL) rats, supporting the reciprocally coordinated expression of the proteins. RSV supplementation led to an unexpected 50 % increase in the frequency of the oxidized base OH8dG in mtDNA. Furthermore, RSV supplementation induced a 50 % increase in the DRP1 protein of mitochondrial dynamics. In both rat groups an inverse correlation between PGC-1α and the frequency of OH8dG as well as an inverse correlation between PRXIII and the frequency of OH8dG were also found, suggestive of a relationship between oxidative damage to mtDNA and mitochondrial biogenesis activity. Such results may indicate that the antioxidant activity of RSV in aged Sol impinged on the oxidative fiber-specific, ROS-mediated, retrograde communication, thereby affecting the expression of SIRT1, PGC-1α and PRXIII, reducing the compensatory responses to the age-related mitochondrial oxidative stress and decline.

A viral protein targets mitochondria and chloroplasts by subverting general import pathways and specific receptors.

Many plant proteins and some proteins from plant pathogens are dually targeted to chloroplasts and mitochondria, and are supposed to be transported along the general pathways for organellar protein import, but this issue has not been explored yet. Moreover, organellar translocon receptors exist as families of several members whose functional specialization in different cargos is supposed but not thoroughly studied. This article provides novel insights into such topics showing for the first time that an exogenous protein, the melon necrotic spot virus coat protein, exploits the common Toc/Tom import systems to enter both mitochondria and chloroplasts while identifying the involved specific receptors.

Redox regulation, thioredoxins and glutaredoxins in retrograde signalling and in gene transcription.

Integration of reactive oxygen species (ROS)-mediated signal transduction pathways via redox sensors and the thiol-dependent signalling network is of increasing interest in cell biology for their implications in plant growth and productivity. Redox regulation is an important point of control in the protein structure, interactions, cellular location and function, thioredoxins (TRXs) and glutaredoxins (GRXs) being key players in the maintenance of cellular redox homeostasis. The crosstalk between second messengers, ROS, thiol redox signalling and redox homeostasis-related genes, controls almost every aspect of plant development and stress response. We review the emerging roles of TRXs and GRXs in redox-regulated processes interacting with other cell signalling systems such as organellar retrograde communication and gene expression, especially in plants during their development and under stressful environments. This approach will throw light on the specific role of these proteins as redox signalling components, and their importance in different developmental processes during abiotic stress.

Perturbation of protein homeostasis brings plastids at the crossroad between repair and dismantling.

The chloroplast proteome is a dynamic mosaic of plastid- and nuclear-encoded proteins. Plastid protein homeostasis is maintained through the balance between de novo synthesis and proteolysis. Intracellular communication pathways, including the plastid-to-nucleus signalling and the protein homeostasis machinery, made of stromal chaperones and proteases, shape chloroplast proteome based on developmental and physiological needs. However, the maintenance of fully functional chloroplasts is costly and under specific stress conditions the degradation of damaged chloroplasts is essential to the maintenance of a healthy population of photosynthesising organelles while promoting nutrient redistribution to sink tissues. In this work, we have addressed this complex regulatory chloroplast-quality-control pathway by modulating the expression of two nuclear genes encoding plastid ribosomal proteins PRPS1 and PRPL4. By transcriptomics, proteomics and transmission electron microscopy analyses, we show that the increased expression of PRPS1 gene leads to chloroplast degradation and early flowering, as an escape strategy from stress. On the contrary, the overaccumulation of PRPL4 protein is kept under control by increasing the amount of plastid chaperones and components of the unfolded protein response (cpUPR) regulatory mechanism. This study advances our understanding of molecular mechanisms underlying chloroplast retrograde communication and provides new insights into cellular responses to impaired plastid protein homeostasis.

RETRACTED: UPL5 modulates WHY2 protein distribution in a Kub-site dependent ubiquitination in response to [Ca2+]cyt-induced leaf senescence

The translocation of proteins between various compartments of cells is the simplest and most direct way of an/retrograde communication. However, the mechanism of protein trafficking is far understood. In this study, we showed that the alteration of WHY2 protein abundance in various compartments of cells was dependent on a HECT-type ubiquitin E3 ligase UPL5 interacting with WHY2 in the cytoplasm, plastid, and nucleus, as well as mitochondrion to selectively ubiquitinate various Kub-sites (Kub 45 and Kub 227) of WHY2. Plastid genome stability can be maintained by the UPL5-WHY2 module, accompany by the alteration of photosystem activity and senescence-associated gene expression. In addition, the specificity of UPL5 ubiquitinating various Kub-sites of WHY2 was responded to cold or CaCl2 stress, in a dose [Ca2+]cyt-dependent manner. This demonstrates the integration of the UPL5 ubiquitination with the regulation of WHY2 distribution and retrograde communication between organelle and nuclear events of leaf senescence.

OLA1 Phosphorylation Governs the Mitochondrial Bioenergetic Function of Pulmonary Vascular Cells.

Mitochondrial function and metabolic homeostasis are integral to cardiovascular function and influence how vascular cells respond to stress. However, little is known regarding how mitochondrial redox control mechanisms and metabolic regulation interact in the developing lungs. Here we show that human OLA1 (Obg-like ATPase-1) couples redox signals to the metabolic response pathway by activating metabolic gene transcription in the nucleus. OLA1 phosphorylation at Ser232/Tyr236 triggers its translocation from the cytoplasm and mitochondria into the nucleus. Subsequent phosphorylation of OLA1 at Thr325 effectively changes its biochemical function from ATPase to GTPase, promoting the expression of genes involved in the mitochondrial bioenergetic function. This process is regulated by ERK1/2 (extracellular-regulated kinases 1 and 2), which were restrained by PP1A (protein phosphatase 1A) when stress abated. Knockdown of ERK1 or OLA1 mutated to a phosphoresistant T325A mutant blocked its nuclear translocation, compromised the expression of nuclear-encoded mitochondrial genes, and consequently led to cellular energy depletion. Moreover, the lungs of OLA1 knockout mice have fewer mitochondria, lower cellular ATP concentrations, and higher lactate concentrations. The ensuing mitochondrial metabolic dysfunction resulted in abnormal behaviors of pulmonary vascular cells and significant vascular remodeling. Our findings demonstrate that OLA1 is an important component of the mitochondrial retrograde communication pathways that couple stress signals with metabolic genes in the nucleus. Thus, phosphorylation-dependent nuclear OLA1 localization that governs cellular energy metabolism is critical to cardiovascular function.

Brain derived neurotrophic factor/tropomyosin related kinase B signaling impacts diaphragm neuromuscular transmission in a novel rat chemogenetic model.

The neuromuscular junction (NMJ) mediates neural control of skeletal muscle fibers. Neurotrophic signaling, specifically brain derived neurotrophic factor (BDNF) acting through its high-affinity tropomyosin related kinase B (TrkB) receptor is known to improve neuromuscular transmission. BDNF/TrkB signaling also maintains the integrity of antero- and retrograde communication between the motor neuron soma, its distal axons and pre-synaptic terminals and influences neuromuscular transmission. In this study, we employed a novel rat chemogenetic mutation (TrkB F616), in which a 1-naphthylmethyl phosphoprotein phosphatase 1 (1NMPP1) sensitive knock-in allele allowed specific, rapid and sustained inhibition of TrkB kinase activity. In adult female and male TrkB F616 rats, treatment with either 1NMPP1 (TrkB kinase inhibition) or DMSO (vehicle) was administered in drinking water for 14 days. To assess the extent of neuromuscular transmission failure (NMTF), diaphragm muscle isometric force evoked by nerve stimulation at 40 Hz (330 ms duration trains repeated each s) was compared to isometric forces evoked by superimposed direct muscle stimulation (every 15 s). Chronic TrkB kinase inhibition (1NMPP1 group) markedly worsened NMTF compared to vehicle controls. Acute BDNF treatment did not rescue NMTF in the 1NMPP1 group. Chronic TrkB kinase inhibition did not affect the apposition of pre-synaptic terminals (labeled with synaptophysin) and post-synaptic endplates (labeled with α-Bungarotoxin) at diaphragm NMJs. We conclude that inhibition of BDNF/TrkB signaling in TrkB F616 rats disrupts diaphragm neuromuscular transmission in a similar manner to TrkB F616A mice, likely via a pre-synaptic mechanism independent of axonal branch point failure.

Mitochondrial DNA polymorphisms in COX1 affect the lifespan of Caenorhabditis elegans through nuclear gene dct-15

Mutations in the mitochondrial DNA (mtDNA) are closely related to age and age-related complex diseases, but the exact regulatory mechanism of mtDNA natural variation or polymorphism and ageing remains unclear. Recently, nuclear genes that regulate mitochondrial functions and thereby influence ageing have been widely studied. In this study, the relationship between the retrograde communication from the mitochondria to the nucleus and its ultimate effect on ageing has been elucidated. This study found that the natural variations in COX1 of the mitochondria in the Caenorhabditis elegans population do not correlate with multiple phenotypes, except for a mild correlation with lifespan. After excluding the differences in the nuclear genome, the correlation between natural mitochondrial variation and lifespan increased significantly. Moreover, mtDNA variation downregulated the nuclear dct-15 gene expression, which consequently reduced the lifespan, development rate and motility of C. elegans. dct-15 mutations decreased mitochondria copy number but increased ATP content and mitochondrial ultrastructure. Thus, the results indicated that dct-15 interacted with the mitochondrial DNA polymorphisms in COX1 and is associated with ageing. Finally, bioinformatic analyses revealed that mtDNA variation regulated the structural constituent of the cuticle via dct-15 and suggested that the structural constituent of the cuticle could have an important role in the development and ageing processes. These results provide insights into the mtDNA mechanism that can alter the nuclear gene and thereby regulate ageing and ageing-related diseases.

Plant photoreceptors and their signalling components in chloroplastic anterograde and retrograde communication.

The red phytochrome and blue cryptochrome plant photoreceptors play essential roles in promoting genome-wide changes in nuclear and chloroplastic gene expression for photomorphogenesis, plastid development, and greening. While their importance in anterograde signalling has been long recognized, the molecular mechanisms involved remain under active investigation. More recently, the intertwining of the light signalling cascades with the retrograde signals for the optimization of chloroplast functions has been acknowledged. Advances in the field support the participation of phytochromes, cryptochromes, and key light-modulated transcription factors, including HY5 and the PIFs, in the regulation of chloroplastic biochemical pathways that produce retrograde signals, including the tetrapyrroles and the chloroplastic MEP-isoprenoids. Interestingly, in a feedback loop, the photoreceptors and their signalling components are targets themselves of these retrograde signals, aimed at optimizing photomorphogenesis to the status of the chloroplasts, with GUN proteins functioning at the convergence points. High light and shade are also conditions where the photoreceptors tune growth responses to chloroplast functions. Interestingly, photoreceptors and retrograde signals also converge in the modulation of dual-localized proteins (chloroplastic/nuclear) including WHIRLY and HEMERA/pTAC12, whose functions are required for the optimization of photosynthetic activities in changing environments and are proposed to act themselves as retrograde signals.

Mitochondria ensure immune surveillance by retro-communication with the nucleus

Mitochondria cover several functions within the cell, including an influence on the transcription of nuclear genes. Recent work by Tigano etal. (2021) in Nature has identified a pathway of mitochondrial retrograde communication in which the nucleus senses aberrations in the mtDNA to drive the innate immune response.

GUN1 and Plastid RNA Metabolism: Learning from Genetics.

GUN1 (genomes uncoupled 1), a chloroplast-localized pentatricopeptide repeat (PPR) protein with a C-terminal small mutS-related (SMR) domain, plays a central role in the retrograde communication of chloroplasts with the nucleus. This flow of information is required for the coordinated expression of plastid and nuclear genes, and it is essential for the correct development and functioning of chloroplasts. Multiple genetic and biochemical findings indicate that GUN1 is important for protein homeostasis in the chloroplast; however, a clear and unified view of GUN1′s role in the chloroplast is still missing. Recently, GUN1 has been reported to modulate the activity of the nucleus-encoded plastid RNA polymerase (NEP) and modulate editing of plastid RNAs upon activation of retrograde communication, revealing a major role of GUN1 in plastid RNA metabolism. In this opinion article, we discuss the recently identified links between plastid RNA metabolism and retrograde signaling by providing a new and extended concept of GUN1 activity, which integrates the multitude of functional genetic interactions reported over the last decade with its primary role in plastid transcription and transcript editing.

The plastid transcription machinery and its coordination with the expression of nuclear genome: Plastid-Encoded Polymerase, Nuclear-Encoded Polymerase and the Genomes Uncoupled 1-mediated retrograde communication.

Plastid genes in higher plants are transcribed by at least two different RNA polymerases, the plastid-encoded RNA polymerase (PEP), a bacteria-like core enzyme whose subunits are encoded by plastid genes (rpoA, rpoB, rpoC1 and rpoC2), and the nuclear-encoded plastid RNA polymerase (NEP), a monomeric bacteriophage-type RNA polymerase. Both PEP and NEP enzymes are active in non-green plastids and in chloroplasts at all developmental stages. Their transcriptional activity is affected by endogenous and exogenous factors and requires a strict coordination within the plastid and with the nuclear gene expression machinery. This review focuses on the different molecular mechanisms underlying chloroplast transcription regulation and its coordination with the photosynthesis-associated nuclear genes (PhANGs) expression. Particular attention is given to the link between NEP and PEP activity and the GUN1- (Genomes Uncoupled 1) mediated chloroplast-to-nucleus retrograde communication with respect to the Δrpo adaptive response, i.e. the increased accumulation of NEP-dependent transcripts upon depletion of PEP activity, and the editing-level changes observed in NEP-dependent transcripts, including rpoB and rpoC1, in gun1 cotyledons after norflurazon or lincomycin treatment. The role of cytosolic preproteins and HSP90 chaperone as components of the GUN1-retrograde signalling pathway, when chloroplast biogenesis is inhibited in Arabidopsis cotyledons, is also discussed. This article is part of the theme issue 'Retrograde signalling from endosymbiotic organelles'.

Adipose Tissue and FoxO1: Bridging Physiology and Mechanisms.

Forkhead box O class proteins (FoxOs) are expressed nearly in all tissues and are involved in different functions such as energy metabolism, redox homeostasis, differentiation, and cell cycle arrest. The plasticity of FoxOs is demonstrated by post-translational modifications that determine diverse levels of transcriptional regulations also controlled by their subcellular localization. Among the different members of the FoxO family, we will focus on FoxO1 in adipose tissue, where it is abundantly expressed and is involved in differentiation and transdifferentiation processes. The capability of FoxO1 to respond differently in dependence of adipose tissue subtype underlines the specific involvement of the transcription factor in energy metabolism and the “browning” process of adipocytes. FoxO1 can localize to nuclear, cytoplasm, and mitochondrial compartments of adipocytes responding to different availability of nutrients and source of reactive oxygen species (ROS). Specifically, fasted state produced-ROS enhance the nuclear activity of FoxO1, triggering the transcription of lipid catabolism and antioxidant response genes. The enhancement of lipid catabolism, in combination with ROS buffering, allows systemic energetic homeostasis and metabolic adaptation of white/beige adipocytes. On the contrary, a fed state induces FoxO1 to accumulate in the cytoplasm, but also in the mitochondria where it affects mitochondrial DNA gene expression. The importance of ROS-mediated signaling in FoxO1 subcellular localization and retrograde communication will be discussed, highlighting key aspects of FoxO1 multifaceted regulation in adipocytes.

Spatiotemporal regulation of GSK3β levels by miRNA-26a controls axon development in cortical neurons.

ABSTRACTBoth the establishment of neuronal polarity and axonal growth are crucial steps in the development of the nervous system. The local translation of mRNAs in the axon provides precise regulation of protein expression, and is now known to participate in axon development, pathfinding and synaptic formation and function. We have investigated the role of miR-26a in early stage mouse primary cortical neuron development. We show that micro-RNA-26a-5p (miR-26a) is highly expressed in neuronal cultures, and regulates both neuronal polarity and axon growth. Using compartmentalised microfluidic neuronal cultures, we identified a local role for miR-26a in the axon, where the repression of local synthesis of GSK3β controls axon development and growth. Removal of this repression in the axon triggers local translation of GSK3β protein and subsequent transport to the soma, where it can impact axonal growth. These results demonstrate how the axonal miR-26a can regulate local protein translation in the axon to facilitate retrograde communication to the soma and amplify neuronal responses, in a mechanism that influences axon development.

Control of retrograde signalling by protein import and cytosolic folding stress.

Communication between organelles and the nucleus is essential for fitness and survival. Retrograde signals are cues emitted from the organelles to regulate nuclear gene expression. GENOMES UNCOUPLED1 (GUN1), a protein of unknown function, has emerged as a central integrator, participating in multiple retrograde signalling pathways that collectively regulate the nuclear transcriptome. Here, we show that GUN1 regulates chloroplast protein import through interaction with the import-related chaperone cpHSC70-1. We demonstrated that overaccumulation of unimported precursor proteins (preproteins) in the cytosol causes a GUN phenotype in the wild-type background and enhances the GUN phenotype of the gun1 mutant. Furthermore, we identified the cytosolic HSP90 chaperone complex, induced by overaccumulated preproteins, as a central regulator of photosynthetic gene expression that determines the expression of the GUN phenotype. Taken together, our results suggest a model in which protein import capacity, folding stress and the cytosolic HSP90 complex control retrograde communication.

Current understanding of GUN1: a key mediator involved in biogenic retrograde signaling.

Chloroplast-nucleus communication takes place via processes called anterograde and retrograde signaling pathways. Discovery of the retrograde signaling pathways from the chloroplasts to the nucleus also raised an intriguing proposition that chloroplasts may serve as environmental sensors since multitudes of environmental factors disturb chloroplastic homeostasis. Certain chloroplastic perturbations, mostly impairing transcription/translation, are coupled to the repression of photosynthesis-associated nuclear genes (PhANGs), thus finely coordinating photosynthetic and chloroplastic homeostasis. The unbiased forward genetic screen in Arabidopsis leads to the identification of six independent loci called GENOMES UNCOUPLED (GUN), whose inactivation was found to de-repress the expression of PhANGs under certain conditions promoting retrograde signaling. Of the six GUNs, five encode proteins associated with tetrapyrrole biosynthesis and one, namely GUN1, encodes a member of the pentatricopeptide repeat protein family. Despite the fact that GUN1 plays a role as a central signaling mediator for retrograde communication, the molecular details of GUN1 protein still remain to be elucidated. Here, we recapitulate our current understanding of the GUN1-mediated retrograde signaling pathway and propose a possible mode of action of GUN1 in the chloroplasts together with different aspects of GUN1 protein activity that deserve further investigation.

A role of GUNs-Involved retrograde signaling in regulating Acetyl-CoA carboxylase 2 in Arabidopsis

In Arabidopsis thaliana (Arabidopsis), Acetyl-CoA Carboxylase 2 (ACC2) is a nuclear DNA-encoded and plastid-targeted enzyme that catalyzes the conversion of acetyl-CoA to malonyl-CoA. ACC2 improves plant growth and development when chloroplast translation is impaired. However, little is known about the upstream signals that regulate ACC2. Here, through analyzing the transcriptome changes in brz-insensitive-pale green (bpg) 2-2, a pale-green mutant with impaired chloroplast gene expression resulting from loss of the BPG2 function, we found that the level of ACC2 was significantly up-regulated. Through performing genetic analysis, we further demonstrated that loss of the GENOMES UNCOUPLED 1 (GUN1) or GUN5 function partly perturbed the up-regulation of ACC2 in the bpg2-2 mutant, whereas ABA INSENSITIVE 4 (ABI4)-function-loss had no clear effect on the ACC2 expression. Furthermore, when plants were treated with plastid translation inhibitors, such as lincomycin and spectinomycin, the ACC2 transcriptional level was also markedly increased in a GUN-dependent manner. In conclusion, our results suggested that the GUN-involved plastid-to-nucleus retrograde communication played a role in regulating ACC2 in Arabidopsis.

Cardiolipin Synthesis in Brown and Beige Fat Mitochondria Is Essential for Systemic Energy Homeostasis

SummaryActivation of energy expenditure in thermogenic fat is a promising strategy to improve metabolic health, yet the dynamic processes that evoke this response are poorly understood. Here we show that synthesis of the mitochondrial phospholipid cardiolipin is indispensable for stimulating and sustaining thermogenic fat function. Cardiolipin biosynthesis is robustly induced in brown and beige adipose upon cold exposure. Mimicking this response through overexpression of cardiolipin synthase (Crls1) enhances energy consumption in mouse and human adipocytes. Crls1 deficiency in thermogenic adipocytes diminishes inducible mitochondrial uncoupling and elicits a nuclear transcriptional response through endoplasmic reticulum stress-mediated retrograde communication. Cardiolipin depletion in brown and beige fat abolishes adipose thermogenesis and glucose uptake, which renders animals insulin resistant. We further identify a rare human CRLS1 variant associated with insulin resistance and show that adipose CRLS1 levels positively correlate with insulin sensitivity. Thus, adipose cardiolipin has a powerful impact on organismal energy homeostasis through thermogenic fat bioenergetics.

Development of a tissue-specific ribosome profiling approach in Drosophila enables genome-wide evaluation of translational adaptations.

Recent advances in next-generation sequencing approaches have revolutionized our understanding of transcriptional expression in diverse systems. However, measurements of transcription do not necessarily reflect gene translation, the process of ultimate importance in understanding cellular function. To circumvent this limitation, biochemical tagging of ribosome subunits to isolate ribosome-associated mRNA has been developed. However, this approach, called TRAP, lacks quantitative resolution compared to a superior technology, ribosome profiling. Here, we report the development of an optimized ribosome profiling approach in Drosophila. We first demonstrate successful ribosome profiling from a specific tissue, larval muscle, with enhanced resolution compared to conventional TRAP approaches. We next validate the ability of this technology to define genome-wide translational regulation. This technology is leveraged to test the relative contributions of transcriptional and translational mechanisms in the postsynaptic muscle that orchestrate the retrograde control of presynaptic function at the neuromuscular junction. Surprisingly, we find no evidence that significant changes in the transcription or translation of specific genes are necessary to enable retrograde homeostatic signaling, implying that post-translational mechanisms ultimately gate instructive retrograde communication. Finally, we show that a global increase in translation induces adaptive responses in both transcription and translation of protein chaperones and degradation factors to promote cellular proteostasis. Together, this development and validation of tissue-specific ribosome profiling enables sensitive and specific analysis of translation in Drosophila.

Disparate Postsynaptic Induction Mechanisms Ultimately Converge to Drive the Retrograde Enhancement of Presynaptic Efficacy

Retrograde signaling systems are fundamental modes of communication synapses utilize to dynamically and adaptively modulate activity. However, the inductive mechanisms that gate retrograde communication in the postsynaptic compartment remain enigmatic. We have investigated retrograde signaling at the Drosophila neuromuscular junction, where three seemingly disparate perturbations to the postsynaptic cell trigger a similar enhancement in presynaptic neurotransmitter release. We show that the same presynaptic genetic machinery and enhancements in active zone structure are utilized by each inductive pathway. However, all three induction mechanisms differ in temporal, translational, and CamKII activity requirements to initiate retrograde signaling in the postsynaptic cell. Intriguingly, pharmacological blockade of postsynaptic glutamate receptors, and not calcium influx through these receptors, is necessary and sufficient to induce rapid retrograde homeostatic signaling through CamKII. Thus, three distinct induction mechanisms converge on the same retrograde signaling system to drive the homeostatic strengthening of presynaptic neurotransmitter release.