Plant and Cell Physiology

BackgroundRAB Guanine Nucleotide Dissociation Inhibitors (RAB GDIs) are important vesicle transport regulators in eukaryotes, participating in the functional cycle of RAB GTPases by stabilizing their non-active GDP-conformation. AimsWe address the importance of the three Arabidopsis thaliana RAB GDI paralogs by genetic and developmental analyses and put these results into the seed plants evolution context. MethodsWe use methods of genetics, microscopy and phylogenetics. ResultsOur genetic analyses of Arabidopsis T-DNA insertional mutants confirm recent CRISPR alleles data indicating lethality of double gdi1 gdi2 mutants, and our microscopic data point to embryo development arrest in double mutant seeds. We also confirm the involvement of GDI2 and GDI3 in pollen tube growth. Moreover, our data show that GDI1 also contributes to proper pollen function. Our phylogenetic analysis reveals independent diversification of RAB GDIs in Gymnosperms and Angiosperms, with early specialization of an Angiosperm reproduction-and gametophyte-related clade. ConclusionsIn Arabidopsis, RAB GDI1 and 2 are important for the vegetative growth while RAB GDI2 and 3 are vital for reproduction. Evolution of the RAB GDI family reflects the evolution of seed plants. HighlightsRAB GDIs are vital for plant growth and reproduction and act redundantly. Even the low-transcribed RAB GDI1 isoform contributes to the proper pollen function. Two RAB GDI clades evolved in early Angiosperms.

Photosynthetic electron transport is mediated by several protein supercomplexes that are spatially arranged in the thylakoid membranes of chloroplasts. The chloroplast NADH dehydrogenase-like (NDH) complex is part of the photosynthetic alternative electron transport (AET) chain, which reduces the plastoquinone (PQ) pool using reduced ferredoxin as a substrate. This NDH complex is associated with photosystem I (PSI) and mediates a portion of AET in stroma lamellae, whereas photosystem II (PSII) is concentrated in grana stacks. This study presents the findings regarding post-illumination chlorophyll fluorescence increase (PIFI), a protein crucial for regulating AET via the NDH pathway. A marked increase in NDH activity and a reduction in the PQ pool in the dark were observed in PIFI-deficient mutant strains (g-pifi) generated by genome editing. Blue native PAGE analysis indicated that PIFI was associated with the NDH-PSI supercomplex in the wild type, and the NDH complex was dissociated from PSI in the g-pifi mutants. Additionally, the g-pifi mutants exhibited a decrease in the maximum quantum yield of PSII (Fv/Fm). Notably, Fv/Fm was restored in a double mutant harboring both g-pifi and NDH-deficient pnsl1 mutations, demonstrating that deregulated NDH activity in g-pifi causes downregulation of PSII efficiency. However, the lower Fv/Fm was not observed in a mutant lacking thioredoxin m4 (trxm4), which showed deregulated NDH activity but maintained the NDH-PSI supercomplex. These data suggest that PIFI stabilizes the NDH-PSI supercomplex and maintains the spatial localization of PQ reduction via AET in thylakoid membranes, which is essential for the proper functioning of PSII.

The RETINOBLASTOMA-RELATED (RBR) protein in plants functions as a cell-cycle inhibitor, regulating cell numbers in developing organs and establishing cellular quiescence during growth. Although the role of RBR counterparts in animals also involves regulating cell size, this potential function remains unexplored in plants. We investigated transgenic Arabidopsis plants with altered RBR levels and observed corresponding changes in cell size from embryogenesis through organ development. In addition, stomatal meristemoid cells with reduced RBR levels divided beyond the size threshold, whereas elevated RBR levels increased their size. RBR stimulated terminal differentiation in the stomatal lineage by inducing MUTE and CYCLIN D5;1 expression, whereas reduced RBR levels maintained asymmetric divisions through high SPEECHLESS and CYCLIN D3;1 expression. Interestingly, the cell proliferation-dependent phosphorylation of RBR at the conserved 911Ser site positively correlated with RBR protein levels in the transgenic lines and aligned with the effect of RBR on cell size. This study discusses the potential link between RBRs control of cell proliferation and cell size, providing new insights into the coordinated regulation of plant development.

The establishment of aphid-plant interaction involves the secretion of a salivary MIF protein. Morphological analyses revealed that aphid MpMIF1 prevents plant cell death, protects organelles from stress, and may promote plant cellular recovery. Co-expression of aphid MpMIF1 and the cell death inducer Npp1 revealed that MpMIF1 modulates autophagy-related genes ATG7/BECLIN1, impair plant senescence regulator ATAF1 and regulate apoptosis-like via Caspase-3-like activity. This effect on multiple-cell death pathways helps to maintain cellular homeostasis during aphid infection. Investigations on DNA Damage Response (DDR) signaling pathways demonstrated that aphid MpMIF1 reduces {gamma}H2A.X phosphorylation, maintains activity of the DNA repair protein RAD51 and stabilizes cell cycle checkpoint expression WEE1 under genotoxic stress. Therefore, MpMIF1 actively participates to the maintenance of a functional DDR. Finally, we showed that aphid MpMIF1 physically interacts with SOG1, a functional analog of animal p53 and central regulator of DDR, cell cycle arrest and programmed cell death in plants. These findings establish MpMIF1 as a key regulator of plant cell death during aphid-plant interactions and highlight its potential as a biotechnological tool for protecting major crops against aphid infection.

In flowering plants, pollen tubes communicate with ovular cells to achieve precise one-to-one pollen tube reception. The final step of this communication between the pollen tube and synergid cells has been extensively investigated and visualized by calcium imaging. Synergid cells exhibit characteristic cytoplasmic calcium concentration oscillations, which are thought to play a critical role in pollen tube reception. However, their significance and relationship with calcium dynamics in the entire ovule remain unclear. Here, we show, using the calcium sensor GCaMP6s, that proteins involved in asparagine-linked glycosylation (N-linked glycosylation) are required for normal calcium oscillations in synergid cells but are not essential for pollen tube reception. Using a semi-in vivo assay in Arabidopsis thaliana, we found that the amplitude of these oscillations prior to rapid pollen tube growth across the filiform apparatus was reduced in mutants lacking the oligosaccharyltransferase (OST) 3/6 subunit or alpha1,2-glucosyltransferase (ALG) 10, both of which are involved in N-linked glycosylation. Notably, these mutants did not exhibit reduced fertility attributable to defects in the female gametophyte but instead showed a polytubey phenotype due to a sporophytic defect. These findings suggest that N-linked glycans mediate communication between synergid cells and the pollen tube and indicate that the typical pattern of calcium oscillations in synergid cells is not essential for triggering pollen tube rupture. Furthermore, we show that sporophytic tissues of the ovule exhibit calcium waves that propagate toward the funiculus in correlation with pollen tube contact and rupture, implying that ovular tissues can potentially transmit these signals distantly beyond the ovule. Together, these findings reveal previously unrecognized intercellular calcium signaling and its significance in pollen tube reception by the ovule.

Salinity and sodicity stresses adversely affect rice growth and yield. To overcome yield losses, suitable tolerant rice cultivars can be developed through a marker-assisted breeding (MAB) program. In the present study, genomic regions associated with sodicity stress tolerance at the reproductive stage were identified using a high-density 50kSNP array in a recombinant inbred line (RIL) population derived from the contrasting rice genotypes CSR11 and MI48. A total of 50 QTLs were detected for various yield-related traits; further, 19 QTLs with [≥]15% of phenotypic variance were selected for integrated (omics) analysis. RNA sequencing of leaves and panicles at the reproductive stage under sodic stress conditions was employed to find differentially expressed genes. A total of 1368 and 1410 SNPs; 104 and 144 indels were found for MI48 and CSR11, respectively, within the QTL regions from resequencing. At chromosomes 1 and 6, colocalized QTLs (qPH1-1/qGP1-1 and qGP6-2/qSSI6-2) were discovered. Differentially expressed genes (DEGs) were mapped over the QTL regions selected, and SNP variations and indels were screened for colocalized QTLs. Potential candidate genes, namely Os-pGlcT1 (Os01g0133400), OsHKT2;1 (Os06g0701600) and OsHKT2;4 (Os06g0701700), OsANTH12 (Os06g0699800), and OsPTR2 (Os06g0706400), were identified as being responsible for glucose transport, ion homeostasis, pollen germination, and nitrogen use efficiency, respectively, under salt stress. Finally, our study provides important insights into the genes and potential mechanisms affecting grain yield under sodic stress in rice, which will contribute to the development of molecular markers for rice breeding programs.

The chloroplast NADH dehydrogenase-like (NDH) complex mediates cyclic electron transport (CET) around photosystem I (PSI) and contributes to photosynthetic regulation and photoprotection under various environmental stresses. Although NDH function has been extensively characterized under controlled conditions, NDH-deficient mutants often show only subtle phenotypes in such environments, leaving its physiological importance under naturally fluctuating field conditions poorly understood. Here, we evaluated growth, yield, and photosynthetic performance of NDH-deficient rice cultivated under field conditions. Mutant plants exhibited reduced biomass accumulation and grain yield compared with wild type. Detailed physiological analyses revealed that NDH deficiency markedly decreased PSI electron transport and CO2 assimilation, particularly under low temperature and sub-saturating irradiance. At moderate and high temperatures, reductions in carbon fixation were largely confined to low-light conditions, whereas at low temperatures, impairment extended across nearly the entire light response range. Under repetitive fluctuating light regimes, NDH-deficient plants showed progressive declines in photosynthesis accompanied by a selective decrease in PSI photochemical capacity without changes in PSII maximum efficiency, indicating PSI-specific photoinhibition. These findings demonstrate that NDH-dependent CET plays a crucial role in sustaining photosynthetic efficiency and crop productivity in dynamic field environments by stabilizing PSI redox balance and maintaining long-term carbon gain. Summary StatementNDH-dependent cyclic electron transport supports photosynthesis and yield in field-grown rice by maintaining PSI function under fluctuating light, low temperature, and sub-saturating irradiance.

Salt stress alters plant development, including the floral transition, but regulation of timing of flowering by salt is poorly understood at the molecular level. To identify genetic loci regulating the floral transition under high soil salinity, we performed a genome-wide association study (GWAS) in Arabidopsis thaliana and identified natural variation at the UGT74E1-UGT74E2-BT3 (UUB) locus that correlates with bolting time specifically in response to salt stress. Genetic analysis revealed BT3 as a novel repressor of the floral transition in control conditions. Similarly, the putative IBA glycosylases UGT74E1 & UGT74E2 delay the floral transition in control conditions. Furthermore, we identified that IBA homeostasis regulators TOB1 and ECH2/IBR10 play a key role in the floral transition, and that ECH2/IBR10 are required for the early flowering phenotype of the ugt74e1/ugt74e2 double mutant, indicating that UGT74E1 & UGT74E2 delay flowering by altering IBA homeostasis. A pangenome analysis of the UUB locus revealed variation in the occurrence of the DNA transposon SAUERKRAUT (SKRT). CRISPR-mediated SKRT deletion in Col-0 affected gene expression both within and outside the UUB locus and caused a salt-dependent delayed floral transition. The delayed bolting phenotype of the skrt-2 mutant also depends on ECH2/IBR10 function, indicating that SKRT accelerates the floral transition by altering IBA homeostasis. Finally, targeted demethylation of SKRT resulted in delayed floral transition under salt stress. Taken together, our data show a role for SKRT and its DNA methylation levels in the salt-dependent bolting time response in Arabidopsis, revealing a novel molecular mechanism to control flowering in adverse conditions.

The unicellular green alga Chlamydomonas reinhardtii provides a tractable model for investigating how carbon availability influences metabolic organization and cell-cycle control in photosynthetic eukaryotes. Its capacity for autotrophic (light, CO2), mixotrophic (light, CO2, acetate), and heterotrophic (acetate, dark) growth enables systematic analysis of trophic-state-dependent regulation. We performed comparative transcriptomic analyses of strain 21gr grown under these three regimes at 30 {degrees}C. Mixotrophy resulted in the highest biomass accumulation and was associated with earlier cell-cycle commitment compared with autotrophy, whereas heterotrophy displayed delayed commitment and reduced growth. Transcriptomic profiling revealed coordinated upregulation of central carbon metabolic pathways under mixotrophy, including photorespiration, glycolysis, the oxidative pentose phosphate pathway, and tricarboxylic acid cycle functions, consistent with enhanced carbon flux and biosynthetic capacity. In contrast, heterotrophy preferentially induced acetate assimilation and glyoxylate cycle genes and was accompanied by elevated expression of cell-cycle regulators, including the CDK-inhibitory kinase WEE1. Together, these findings indicate that trophic mode modulates the coupling between carbon metabolism and cell-cycle progression, with mixotrophy supporting integrated metabolic and proliferative activity, whereas heterotrophy is associated with delayed cell-cycle timing and transcriptional signatures of metabolic adjustment.

Photosynthesis accounts for most of the final grain yield in rice, making improvements in radiation use efficiency (RUE) a key strategy for enhancing productivity. Agronomically, RUE is defined as the biomass produced per unit of total solar radiation or photosynthetically active radiation intercepted by the canopy. However, the interaction between carbon and nitrogen metabolism plays a critical role in determining plant growth and grain yield. Assimilated nitrogen is required for the synthesis of photosynthetic pigments and enzymes, while the reduction of nitrate (NOLL) and nitrite (NOLL), as well as the assimilation of ammonium (NHLL), depend on the reducing power and carbon skeletons generated by photosynthesis. In this study, two high-yielding rice (Oryza sativa) cultivars--an indica-type (El Paso 144) and a japonica-type (INIA Parao) were subjected to two nitrogen treatments (3 mM and 9 mM NOLL/NHLL) and two light intensities (850 and 1500 mol mL{superscript 2} sL{superscript 1}). A strong interaction between light intensity and nitrogen metabolism was observed, with contrasting responses between subspecies. These differences reflect a coordinated regulation of carbon assimilation and primary nitrogen metabolism. The results provide new insights into the metabolic strategies underlying nitrogen compound accumulation under variable irradiance. Such knowledge is essential for improving nitrogen fertilizer use efficiency and yield performance in elite rice genotypes cultivated under commercial field conditions.

AO_SCPLOWBSTRACTC_SCPLOWNitrogen fertilizers are essential for crop productivity but cause environmental harm, necessitating the development of cultivars that thrive under limited nitrogen. This study investigates the transcriptomic response to nitrate in Arabidopsis thaliana (a model dicot), Brachypodium distachyon (a model Pooideae), and Hordeum vulgare (barley, a domesticated Pooideae) to identify conserved and species-specific molecular mechanisms. Using RNA-seq after 1.5 and 3 hours of nitrate treatment, we found that core nitrate-responsive biological processes - such as nitrate transport, assimilation, carbon metabolism, and hormone signaling - are largely conserved across species. However, comparative analysis at gene level based on orthology revealed specificities between the species. For instance, rRNA processing was uniquely stimulated in Arabidopsis, while cysteine biosynthesis from serine and gibberellin biosynthesis were specifically regulated in Brachypodium and barley. Orthologs of key nitrate-responsive genes (e.g., NRT, NLP, TCP20) exhibited variable regulation, reflecting potential adaptations linked to domestication or nutrient acquisition strategies. These findings highlight the importance of integrating model and crop species to uncover targets for improving nitrogen use efficiency in cereals. The study provides a pipeline integrating gene ontology and orthology analyses to compare transcriptomic responses between species.

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are essential regulators of plant growth, development, and stress adaptation. In this study, we performed a comprehensive genome-wide identification of SNARE genes in cucumber (Cucumis sativus L.), uncovering 51 putative members designated as CsSNAREs. Phylogenetic analysis confirmed that these genes cluster into five major clades: Qa-CsSNARE (14), Qb-CsSNARE (9), Qc-CsSNARE (10), Qb+c-CsSNARE (3), and R-CsSNARE (15). Bioinformatic analysis of their promoter regions, coupled with expression profiling under diverse abiotic stress conditions, highlighted a heightened responsiveness within the Qa-CsSNARE subfamily. To validate this, we selected representative Qa-CsSNARE genes for quantitative real-time PCR analysis under drought and salt stress. Among these, CsSYP121 was notably induced by salt treatment. We subsequently generated transgenic cucumber lines overexpressing CsSYP121 and challenged them with salinity. Phenotypic assessment, combined with measurements of reactive oxygen species (ROS) accumulation and K+/Na+ ratios, demonstrated that CsSYP121 overexpression (OE) confers enhanced salt tolerance and boosts antioxidant capacity. We propose a model wherein CsSYP121 mitigates ROS-induced cellular damage under salt stress, potentially through promoting K+/Na+ homeostasis, thereby improving plant performance under saline conditions. Our findings identify CsSYP121 as a promising candidate gene for breeding salt-tolerant crops.

Malate and fumarate constitute a significant transient carbon stock that is dynamically synthesized during the photoperiod. These organic acids are diurnally stored and remobilised from the vacuole, and they have a key role in the cellular metabolic regulation. This function is well known in C4 and CAM plants. However, in C3 species that are the majority of terrestrial plants, the importance of the vacuolar accumulation/release and its influence on plant growth is still an open question. In Here we addressed this issue generating multiple knockout mutants in Arabidopsis thaliana lacking vacuolar anion channels of the Aluminium-Activated Malate Transporter (ALMT) family, to impair malate and fumarate transport to the vacuole. We show that in these mutants reducing vacuolar transport of malate and fumarate in mesophyll cells leads to a dramatic growth impairment. Metabolic and fluxomic analysis revealed that vacuolar malate and fumarate transport influences plant carbon and nitrogen metabolism as well as cellular pH and ionic homeostasis. In conclusion, our results show that the transport organic acids like malate and fumarate across the vacuolar membrane is essential for plant growth in a C3 plant too. These results establish the importance of the vacuolar pools of malate and fumarate in plant metabolism.

Identifying loci in the genome that allow a population to respond to selection pressure is essential to understand evolution and improve crops. Temporally consecutive generations under selection offer the opportunity to identify signatures of selection. Maize, as one of the most important crops worldwide is rich in genetic diversity and a model for breeding advances. Therefore, it is an ideal system to study genetic changes in response to selection. Here, we study the genetic changes in two replicates of a selection experiment in a European maize landrace, which showed rapid trait improvement over three cycles of selection. We identified an increase in genetic divergence across successive doubled-haploid populations derived from each selection cycle, consistent with the effect of strong directional selection. The genetic divergence observed between the replicates was greater than that between generations. In addition to the genome-wide signal, we identified multiple candidate loci under selection through temporal FST outlier analysis comparing the original landrace population to subsequent cycles. These loci showed a significant overlap with genomic regions, controlling intentionally selected traits and other traits. The significant overlap of selected loci between the two replicates shows the importance of major loci in response to directional selection, while the large number of non-overlapping loci demonstrates the polygenic response. Our work shows that the temporal dimension in plant breeding time-series enables the identification of candidate loci under selection and the genome-wide dynamics of change in response to selection.

Hundreds of proteins in the cell require iron (Fe) or Fe-containing cofactors to function. However, how Fe2+ or Fe3+ are specifically allocated to each of these proteins in plant cells remains largely unknown. It has been proposed that Fe metalation could be driven by specific interactions with Fe-shuttling proteins known as Fe-chaperones. Here, we present the first family of plant Fe2+-chaperones (ICHAPs) with orthologues in dicots and monocots. The role of these proteins in Fe distribution to Fe-dependent metabolic processes has been illustrated using symbiotic nitrogen fixation in Medicago truncatula root nodules. ICHAP1 is a soluble Fe2+-binding protein that interacts with plasma membrane Fe2+ transporter NRAMP1, but not with symbiosome Fe2+-transporters. ICHAP1 mutants present altered Fe distribution in cells and they cannot fix nitrogen. A second family member, ICHAP2 is required to target Fe2+ to symbiosomes, as it accepts Fe2+ from ICHAP1 and interacts with symbiosome Fe2+-importer VTL8, but not with NRAMP1. These results indicate a path for Fe2+ allocation from the plasma membrane to the symbiosome through specific protein-protein interactions and Fe2+ exchange from NRAMP1 to ICHAP1, to ICHAP2, and to VTL8.

Inositol phosphates (InsPs) and inositol pyrophosphates (PP-InsPs) are conserved signalling molecules, but their evolutionary origin and diversification in the green lineage remain poorly understood. Here we investigated the InsP network in the streptophyte alga Chara braunii, a key lineage lose to the origin of land plants. Using capillary electrophoresis-electrospray ionization mass spectrometry, we detected a broad spectrum of InsP and PP-InsP species from InsP3 to InsP8, including multiple positional isomers. Developmental profiling across dormant oospores, young thalli and mature thalli revealed extensive metabolic remodeling, with InsP6 as the dominant metabolite and distinct stage-dependent changes in lower InsPs and pyrophosphorylated species. Multiple PP-InsP5 and (PP)2-InsP4 isomers were identified, together with an unassigned additional InsP8-like signal, indicating further pathway complexity. Bioinformatic analyses identified candidate homologs of major InsP metabolic enzymes, supporting the presence of an enzymatic framework for InsP synthesis and turnover similar to land plants. Environmental perturbation revealed isomer-selective effects: prolonged light and dark phases strongly affected the accumulation of specific InsP5 and PP-InsP5 isomers, with 1-PP-InsP5 emerging as the most stimulus-responsive pyrophosphate species, whereas heat stress preferentially reduced 4-PP-InsP5. Together, these findings show that a structurally complex and environmentally responsive InsP network was already established in streptophyte algae before the emergence of land plants.

Plant survival and growth depend partly on the ability to manage energy resources in response to changing environmental conditions. SnRK1 plays a central role in this process by restricting growth under energy-limiting conditions while promoting stress adaptation and survival. When activated, SnRK1 triggers transcriptional reprogramming that prioritizes energy-producing pathways. A key mediator of this response is the transcription factor bZIP63, whose activity is regulated by SnRK1-dependent phosphorylation. Given its roles in energy homeostasis and its interaction with the circadian clock, bZIP63 influences growth and is therefore a candidate component of the Metabolic Daylength Measurement (MDLM) system, which integrates starch and sucrose metabolism with circadian timing and photosynthetic duration to regulate vegetative growth under contrasting photoperiods. We show that 39 bZIP63 direct targets regulated by SnRK1 correspond to a subset of short-day-induced genes associated with the MDLM system and are downregulated in a bZIP63 T-DNA mutant (bzip63-2) and/or in an RNAi-induced silencing line (RNAiWs_L9). Downregulation of these genes was more extensive in RNAiWs_L9 than in bzip63-2, possibly due to the unexplained silencing of BAM4, a {beta}-amylase that promotes starch degradation. Under short-day conditions, the frameshift mutant bzip63-5 (Col-0), bzip63-2 (Ws), and the bzip1-1/bzip53-1/bzip63-5 (Col-0) triple mutant, which disrupts bZIP63 heterodimerization partners, showed similar deregulation of a subset of these genes and comparable growth inhibition, whereas both growth and gene deregulation were more strongly affected in RNAiWs_L9. We further show in two partially complemented bzip63-2 lines that bZIP63 protein levels increase toward the end of the night and decline toward the end of the day, in synchrony with the diel oscillation of its transcript. Additional analyses of these lines, together with bzip63-2 line overexpressing bZIP63, suggest that the timing and amplitude of bZIP63 accumulation contribute to shaping the expression profiles of a subset of the 39 MDLM-associated genes. Together, these findings indicate that bZIP63 participates in a regulatory network linking SnRK1 signaling, photoperiod-changes, and growth within the MDLM system.

Understanding the regulation of enzyme activity involved in photosynthesis is essential for engineering enhanced carbon fixation in crops. In C4 plants, the enzyme phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) is one of the most abundant leaf enzymes and plays an essential role in photosynthetic carbon dioxide (CO2) fixation. The enzyme also plays a key role in central metabolism (e.g., providing intermediates to the citric acid cycle) and therefore must be highly regulated to coordinate its activity. The regulation of PEPC activity can occur allosterically by glucose 6-phosphate activation and malate inhibition, which is in part influenced by reversible phosphorylation. A specific light-dependent phosphorylation of PEPC at an N-terminal serine residue by the PEPC-protein kinase (PEPC-PK) can regulate its sensitivity to this allosteric regulation. However, the impact of this PEPC phosphorylation has not been tested in a C4 crop. Therefore, we created PEPC-PK mutant lines in Zea mays to assess the impact of PEPC phosphorylation on its allosteric regulation, photosynthesis, and growth. While the maximum PEPC activity was unchanged, PEPC in the PEPC-PK mutant plants was not phosphorylated under light and was more sensitive to malate inhibition. However, gas exchange, electron transport, and field biomass analyses showed no differences in the PEPC-PK mutant plants. These results demonstrate that in Z. mays PEPC phosphorylation affects enzyme sensitivity to malate in vitro but does not substantially alert photosynthetic performance or growth under field conditions suggesting additional regulation of PEPC activity in planta.

Poplar seed fibers cause environmental and health concerns, yet their developmental mechanisms remain poorly understood. Here, we constructed a high-resolution spatiotemporal transcriptomic atlas of female poplar capsules by integrating single-nucleus and spatial transcriptomics. We delineated the developmental trajectory of seed fibers, confirming their origin from placenta cells, and identified three functionally distinct fiber cell subtypes involved in initiation, metabolic support, and elongation. Weighted gene co-expression network analysis (WGCNA) identified several hub transcription factors, including PtoMYB, PtoHDT1, PtoEIF6 and PtoPDF2, that may serve as key regulators of fiber development. Our study provides a cellular-resolution framework for understanding trichome development in woody perennials and offers candidate targets for functional characterization toward breeding low-fluff poplar cultivars. HighlightsO_LIA spatiotemporal transcriptomic atlas of poplar capsule development is constructed at single-cell resolution C_LIO_LIFiber cells originate from placenta cells and comprise three functionally distinct subtypes C_LIO_LIProvides molecular targets for breeding low-fluff poplar cultivars to mitigate environmental pollution C_LI

Chloroplast HSP70 is an essential component of the plastid proteostasis network, supporting protein folding, complex assembly and disassembly, and stress acclimation. Despite extensive genetic evidence for its essentiality, the cellular consequences of reduced chloroplast HSP70 activity remain poorly defined. Here, we investigated the function of the sole chloroplast HSP70 in Chlamydomonas reinhardtii, HSP70B, using an inducible artificial microRNA approach that reduced HSP70B abundance to below 30% of wild-type levels. HSP70B depletion resulted in cell division arrest and extensive proteome remodeling, characterized by strong upregulation of proteins involved in chloroplast protein quality control and membrane remodeling. Notably, this response was accompanied by increased abundance of protein quality control components in the endoplasmic reticulum, cytosol, and mitochondria, indicating pronounced proteostasis cross-talk between cellular compartments. In contrast, chloroplast and cytosolic ribosomes, photosynthetic and respiratory protein complexes, and central metabolic enzymes were broadly depleted, consistent with a collapse of cellular proteostasis. At the ultrastructural level, HSP70B-depleted cells exhibited lesions at thylakoid membrane conversion zones previously described in VIPP1-depleted cells. Accordingly, higher-order oligomeric forms of VIPP1 accumulated, and cells displayed extreme sensitivity to high-light stress. These findings confirm HSP70B as a key regulator of VIPP1 oligomer dynamics and highlight its central role in coordinating chloroplast membrane remodeling with cellular proteostasis in Chlamydomonas. One-sentence summaryDepletion of chloroplast HSP70B causes cell division arrest, proteostasis collapse, impaired VIPP1 oligomer dynamics with aberrant thylakoid structures, and increased light sensitivity.