Chilling stress severely restricts plant growth and productivity by impairing photosynthetic performance. However, the molecular mechanisms underlying chilling-induced photoinhibition and its mitigation remain incompletely understood. In this study, we functionally characterized two basic leucine zipper (bZIP) transcription factors, SlbZIP30 and SlbZIP51, from tomato (Solanum lycopersicum), and demonstrated that they are important regulators involved in photosynthetic protection in response to chilling stress. SlbZIP30 is localized in both chloroplasts and the nucleus with distinct compartment-specific functions. In chloroplasts, SlbZIP30 interacts with and stabilizes triose phosphate isomerase 1 (SlTPI1) and light-harvesting complex protein SlLHCA1 to preserve photosynthetic machinery integrity. In the nucleus, chilling induces SlbZIP51 translocation from the cytoplasm, where it forms a heterodimer with SlbZIP30 and activates transcription of SlCPN60α1, SlTPI2, and SlRBCS1, which encode chaperonin 60 subunit α1, triose phosphate isomerase isoform, and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) small subunit 1, respectively. These proteins support carbon assimilation, sustaining photosynthetic capacity under chilling stress. Collectively, our findings uncover a photoprotective mechanism mediated by dual-localized SlbZIP30, which coordinates chloroplastic protein stabilization and nuclear transcriptional regulation to mitigate chilling-induced photodamage, providing a practical framework for engineering chilling-resilient crops.

Nucleus-encoded proteins are involved in intron splicing in plant chloroplasts. Although many splicing factors were discovered, the mechanisms for chloroplast intron splicing remain unknown. Here, we identified a P-class pentatricopeptide repeat (PPR) protein, PHOTOSYSTEM ONE BIOGENESIS FACTOR6 (PBF6), that is essential for the accumulation of photosystem I complex. PBF6 bound to ycf3 intron 1 and clpP1 intron 2, and the introns of petB, ndhA, and ndhB transcripts, and was required for their splicing. Truncated PBF6 containing the 10 N-terminal PPR motifs bound to specific sequences to ycf3 intron 1, clpP1 intron 2, and to the introns of petB, ndhA, and ndhB. PBF6 formed two multi-subunit splicing complexes with other known splicing factors from the CRM, peptidyl-tRNA hydrolase, RNase III, PORR, APO, DEAD-box RNA helicase, and mTERF families. PBF6-containing complex I was about 600 kDa, comprising nine known splicing factors: CAF1, CAF2, CRS2, CFM3a, RNC1, WTF1, APO2, RH3, and mTERF2. PBF6-containing complex II was about 300 kDa and contained three known splicing factors: CAF1, CAF2, and CFM2. Furthermore, two known PPR-type splicing factors, PBF2 and ECD2, both of which are required for splicing the ycf3 intron 1, also formed two multi-subunit splicing complexes with other known splicing factors. Importantly, these three PPR-type splicing factors formed their splicing complexes independently. Our data suggest that a PPR-type splicing factor forms splicing complexes with other known splicing factors under transient expression conditions to facilitate intron splicing and that several PPR-type splicing factors work together to promote the splicing of the same intron through forming respective splicing complexes.

Red light promotes PsbS transcription through the PHYs-COP1 module, blue light through the CRY1/CRY2-COP1 pathway, and UV-B via the UVR8-COP1-HY5 cascade. ABA negatively regulates PsbS through the PYLs-SnRK2s-ABI5 signaling pathway. Non-photochemical quenching (NPQ) is a central photoprotective mechanism that enables plants to dissipate excess excitation energy under high light conditions, with PsbS functioning as a key regulatory protein. However, the environmental cues controlling PsbS transcription and the underlying regulatory mechanisms remain largely unclear. Analysis of the core promoter region of PsbS identified light-responsive element G-box and abscisic acid-responsive element (ABRE) cis-elements; dual-luciferase reporter assays confirmed that the core PsbS promoter can respond to light and abscisic acid (ABA) signals. Expression analyses showed that red, blue, and ultraviolet-B (UV-B) light significantly induced PsbS transcription, whereas ABA treatment repressed it. Genetic and signaling analyses revealed that red light promotes PsbS transcription through the PHYs-COP1 module, blue light through the CRY1/CRY2-COP1 pathway, and UV-B via the UVR8-COP1-HY5 cascade. In contrast, ABA negatively regulates PsbS through the PYLs-SnRK2s-ABI5 signaling pathway. Yeast one-hybrid (Y1H) screening identified 36 candidate transcription factors targeting PsbS. Dual-luciferase assays and electrophoretic mobility shift assays (EMSA) further confirmed that bZIP2, bZIP16, bZIP43, CCA1, and IDD1 act as candidate transcriptional repressors of PsbS. This study systematically integrates known light and ABA signaling pathways that regulate PsbS expression, providing potential molecular targets for improving crop photosynthetic efficiency.

B-cell receptor-associated protein 31 (BAP31) is an endoplasmic reticulum (ER) transmembrane protein in humans that regulates protein trafficking, apoptosis, and inflammatory responses to maintain cellular homeostasis. However, its role in plants remains largely unknown. Here, we identified two tomato (Solanum lycopersicum) homologs, SlBAP31 and SlBAP31-LIKE, which are strongly induced by heat stress and function redundantly to stabilize photosystem II-light-harvesting complex II (PSII-LHCII) supercomplexes. Although localization to the ER or Golgi apparatus has been detected, both proteins are also present in chloroplasts, where they interact with SlTOC75, a core component of the chloroplast TOC-TIC translocon complex. Loss of either protein reduces the abundance of PSBW under heat stress. Among the two tomato PSBW isoforms, SlPSBW2 plays a predominant role in PSII-LHCII stability, and its accumulation depends in part on glycine 121 at the C terminus and interaction with SlBAP31/SlBAP31-LIKE. Collectively, our results reveal that ER/Golgi- and chloroplast periphery-localized SlBAP31 and SlBAP31-LIKE positively regulate tomato thermotolerance by mediating SlPSBW2 accumulation and stabilizing the PSII-LHCII supercomplex.

Soil salinization threatens global food security, necessitating the development of saline-alkaline-tolerant crops. This study investigated the molecular mechanisms of alkali stress tolerance in maize. Screening 369 inbred lines identified two alkali-resistant and two alkali-sensitive varieties. Systematic analysis revealed that resistant varieties rapidly lowered rhizosphere pH and maintained root architecture, whereas sensitive varieties suffered reduced lateral roots and severe biomass loss. Metabolomic profiling showed that all varieties secreted malonic acid via the pyrimidine pathway to modulate rhizosphere pH, with resistant varieties exhibiting stronger accumulation. Transcriptome and RT-qPCR analysis identified two key genes: Zm00001eb396990 (asparagine synthetase), upregulated in resistant varieties and linked to organic acid synthesis, and Zm00001eb370000 (cytokinin dehydrogenase), downregulated in resistant varieties, potentially aiding root maintenance. Multi-omics correlation confirmed the association between Zm00001eb396990 expression and malonic acid content. This study demonstrates that maize roots can alleviate alkali stress through the secretion of malonic acid and the regulation of related genes, providing potential genetic targets and a theoretical basis for cultivating alkali-tolerant maize.

Photosystem I (PSI) is one of two large pigment–protein complexes responsible for converting solar energy into chemical energy in all oxygenic photosynthetic organisms. The PSI supercomplex consists of the PSI core complex and peripheral light-harvesting complex I (LHCI) in eukaryotic photosynthetic organisms. However, how the PSI complex assembles in land plants is unknown. Here we describe PHOTOSYSTEM I BIOGENESIS FACTOR 8 (PBF8), a thylakoid-anchored protein in Arabidopsis thaliana that is required for PSI assembly. PBF8 regulates two key consecutive steps in this process, the building of two assembly intermediates comprising eight or nine subunits, by interacting with PSI core subunits. We identified putative PBF8 orthologues in charophytic algae and land plants but not in Cyanobacteria or Chlorophyta. Our data reveal the major PSI assembly pathway in land plants. Our findings suggest that novel assembly mechanisms evolved during plant terrestrialization to regulate PSI assembly, perhaps as a means to cope with terrestrial environments.

The plastid-encoded RNA polymerase serves as the principal transcription machinery within chloroplasts, transcribing over 80% of all primary plastid transcripts. This polymerase consists of a prokaryotic-like core enzyme known as the plastid-encoded RNA polymerase core, and is supplemented by newly evolved associated proteins known as PAPs. However, the architecture of the plastid-encoded RNA polymerase and the possible functions of PAPs remain unknown. Here, we present the cryo-electron microscopy structure of a 19-subunit plastid-encoded RNA polymerase complex derived from spinach (Spinacia oleracea). The structure shows that the plastid-encoded RNA polymerase core resembles bacterial RNA polymerase. Twelve PAPs and two additional proteins (FLN2 and pTAC18) bind at the periphery of the plastid-encoded RNA polymerase core, forming extensive interactions that may facilitate complex assembly and stability. PAPs may also protect the complex against oxidative damage and has potential functions in transcriptional regulation. This research offers a structural basis for future investigations into the functions and regulatory mechanisms governing the transcription of plastid genes. The plastid-encoded RNA polymerase serves as the principal transcription machinery within chloroplasts. Here, the authors present the cryo-electron microscopy structure of a 19-subunit plastid-encoded RNA polymerase complex derived from spinach.

Carbon reserve remobilization in stems is closely related to rice grain filling. Sucrose phosphate synthase (SPS) is highly associated with carbon reserve remobilization. In this study, we investigated the expression pattern of SPS genes in various rice tissues, and found that SPS8 is the major SPS isoform in rice stems during the grain-filling stage. We then constructed sps8 mutants using the CRISPR/Cas9 system. The SPS activity of the sps8 mutants was markedly reduced in the stems. In addition, the sps8 mutants exhibited significant starch accumulation in stems. 14C-labelling experiments revealed that the remobilization of non-structural carbohydrates from rice stems to grains was impaired in the sps8 mutants. In the sps8 mutants, grain filling was delayed and yield decreased by 15% due to a reduced percentage of ripened grains. RNA sequencing and quantitative PCR analyses indicated that the genes involved in starch synthesis and degradation were up-regulated in the sps8 mutant stems. In addition, the activity of the enzymes involved in starch synthesis and degradation was increased in the sps8 stems. These results demonstrate that SPS8 is required for carbon reserve remobilization from rice stems to grains, and that its absence may enhance 'futile cycles' of starch synthesis and degradation in rice stems.

Photosystem I (PSI) and PSII are two large pigment-protein complexes that are responsible for converting solar energy into chemical energy. We identify the PSI assembly factor PBF8 and show that it mediates two key consecutive steps in PSI assembly, revealing major aspects of the PSI assembly pathway in land plants.

Plants frequently evade extreme environmental stress by initiating early flowering, yet the underlying mechanisms remain incompletely understood. Here, through extensive mutant screening, we identify a vegetative growth to reproductive growth transition factor (vrf1) mutant, which exhibits a deficiency in drought escape. Alternative splicing of VRF1 generates four isoforms, of which two encode functional proteins, VRF1-AS1 and VRF1-AS3. The proportions of VRF1-AS1 and VRF1-AS3 are modulated by environmental factors, serving as a molecular switch mediating the transition from tolerance to early flowering, irrespective of their absolute abundance. VRF1-AS1 and VRF1-AS3 competitively bind to MKK1, resulting in MKK1 phosphorylation at different sites, which opens two distinct regulatory pathways. The role of VRF1 is conserved across various conditions, and 66 Arabidopsis ecotypes suggest its central function in stress avoidance through premature flowering. In summary, our findings show that plants respond precisely and effectively to dynamic environmental changes by modulating their isoform ratios.

Chloroplasts evolved from an ancient cyanobacterial endosymbiont more than 1.5 billion years ago. During subsequent coevolution with the nuclear genome, the chloroplast genome has remained independent, albeit strongly reduced, with its own transcriptional machinery and distinct features, such as chloroplast-specific innovations in gene expression and complicated post-transcriptional processing. Light activates the expression of chloroplast genes via mechanisms that optimize photosynthesis, mini -mize photodamage, and prioritize energy investments. Over the past few years, studies have moved from describing phases of chloroplast gene expression to exploring the underlying mechanisms. In this re-view, we focus on recent advances and emerging principles that govern chloroplast gene expression in land plants. We discuss engineering of pentatricopeptide repeat proteins and its biotechnological effects on chloroplast RNA research; new techniques for characterizing the molecular mechanisms of chloroplast gene expression; and important aspects of chloroplast gene expression for improving crop yield and stress tolerance. We also discuss biological and mechanistic questions that remain to be answered in the future.

Flowering time (heading date) is a critical agronomic trait that determines the yield and regional adaptability of crops. Heading date 1 (Hd1) is a central regulator of photoperiodic flowering in rice (Oryza sativa). However, how the homeostasis of Hd1 protein is achieved is poorly understood. Here, we report that the nuclear autophagy pathway mediates Hd1 degradation in the dark to regulate flowering. Loss of autophagy function results in an accumulation of Hd1 and delays flowering under both short-day and long-day conditions. In the dark, nucleus-localized Hd1 is recognized as a substrate for autophagy and is subjected to vacuolar degradation via the autophagy protein OsATG8. The Hd1-OsATG8 interaction is required for autophagic degradation of Hd1 in the dark. Our study reveals a new mechanism by which Hd1 protein homeostasis is regulated by autophagy to control rice flowering. Our study also indicates that the regulation of flowering by autophagic degradation of Hd1 orthologs may have arisen over the course of mesangiosperm evolution, which would have increased their flexibility and adaptability to the environment by modulating flowering time.

OPINION article Front. Plant Sci., 30 September 2022Sec. Photosynthesis and Photobiology Volume 13 - 2022 | https://doi.org/10.3389/fpls.2022.967203

Light is essential for photosynthesis but light levels that exceed an organism's assimilation capacity can cause serious damage or even cell death. Plants and microalgae have developed photoprotective mechanisms collectively referred to as non-photochemical quenching to minimize such potential damage. One such mechanism is energy-dependent quenching (qE), which dissipates excess light energy as heat. Over the last 30 years, much has been learned about the molecular mechanism of qE in green algae and plants. However, the steps between light perception and qE represented a gap in our knowledge until the recent identification of light-signaling pathways that function in these processes in the green alga Chlamydomonas reinhardtii. In this review, we summarize the high light and UV-mediated signaling pathways for qE in Chlamydomonas. We discuss key questions remaining about the pathway from light perception to photoprotective gene expression in Chlamydomonas. We detail possible differences between green algae and plants in light-signaling mechanisms for qE and emphasize the importance of research on light-signaling mechanisms for qE in plants.

Root hydrotropism refers to root directional growth toward soil moisture. Cortical microtubule arrays are essential for determining the growth axis of the elongating cells in plants. However, the role of microtubule reorganization in root hydrotropism remains elusive. Here, we demonstrate that the well-ordered microtubule arrays and the microtubule-severing protein KATANIN (KTN) play important roles in regulating root hydrotropism in Arabidopsis. We found that the root hydrotropic bending of the ktn1 mutant was severely attenuated but not root gravitropism. After hydrostimulation, cortical microtubule arrays in cells of the elongation zone of wild-type (WT) Col-0 roots were reoriented from transverse into an oblique array along the axis of cell elongation, whereas the microtubule arrays in the ktn1 mutant remained in disorder. Moreover, we revealed that abscisic acid (ABA) signaling enhanced the root hydrotropism of WT and partially rescued the oryzalin (a microtubule destabilizer) alterative root hydrotropism of WT but not ktn1 mutants. These results suggest that katanin-dependent microtubule ordering is required for root hydrotropism, which might work downstream of ABA signaling pathways for plant roots to search for water.

RNA splicing refers to a process by which introns of a pre-mRNA are excised and the exons at both ends are joined together. Chloroplast introns are inherently self-splicing ribozymes, but over time, they have lost self-splicing ability due to the degeneration of intronic elements. Thus, the splicing of chloroplast introns relies heavily on nuclear-encoded splicing factors, which belong to diverse protein families. Different splicing factors and their shared intron targets are supposed to form ribonucleoprotein particles (RNPs) to facilitate intron splicing. As characterized in a previous review, around 14 chloroplast intron splicing factors were identified until 2010. However, only a few genetic and biochemical evidence has shown that these splicing factors are required for the splicing of one or several introns. The roles of splicing factors are generally believed to facilitate intron folding; however, the precise role of each protein in RNA splicing remains ambiguous. This may be because the precise binding site of most of these splicing factors remains unexplored. In the last decade, several new splicing factors have been identified. Also, several splicing factors were found to bind to specific sequences within introns, which enhanced the understanding of splicing factors. Here, we summarize recent progress on the splicing factors in land plant chloroplasts and discuss their possible roles in chloroplast RNA splicing based on previous studies.

The grain-filling process is crucial for cereal crop yields, but how the caryopsis of such plants is supplied with sugars, which are produced by photosynthesis in leaves and then transported long distance, is largely unknown. In rice (Oryza sativa), various SWEET family sucrose transporters are thought to have important roles in grain filling. Here, we report that OsSWEET14 plays a crucial part in this process in rice. ossweet14 knockout mutants did not show any detectable phenotypic differences from the wild type, whereas ossweet14;ossweet11 double-knockout mutants had much more severe phenotypes than ossweet11 single-knockout mutants, including strongly reduced grain weight and yield, reduced grain-filling rate, and increased starch accumulation in the pericarp. Both OsSWEET14 and OsSWEET11 exhibited distinct spatiotemporal expression patterns between the early stage of caryopsis development and the rapid grain-filling stage. During the rapid grain-filling stage, OsSWEET14 and OsSWEET11 localized to four key sites: vascular parenchyma cells, the nucellar projection, the nucellar epidermis, and cross cells. These results demonstrate that OsSWEET14 plays an important role in grain filling, and they suggest that four major apoplasmic pathways supply sucrose to the endosperm during the rapid grain-filling stage via the sucrose effluxers SWEET14 and SWEET11.

In past 30 years, the wheat yield per unit area of China has increased by 79%. The super-high-yield (SH) cultivation played an important role in improving the wheat photosynthesis and yield. In order to find the ecophysiological mechanism underneath the high photosynthesis of SH cultivation, in situ diurnal changes in the photosynthetic gas exchange and chlorophyll (Chl) a fluorescence of field-grown wheat plants during the grain-filling stage and environmental factors were investigated. During the late grain-filling stage at 24 days after anthesis (DAA), the diurnal changes in net CO2 assimilation rate were higher under SH treatment than under high-yield (H) treatment. From 8 to 24 DAA, the actual quantum yield of photosystem II (PSII) electron transport in the light-adapted state (ΦPSII) in the flag leaves at noon under SH treatment were significantly higher than those under H treatment. The leaf temperature, soil temperature and soil moisture were better suited for higher rates of leaf photosynthesis under SH treatment than those under H treatment at noon. Such diurnal changes in environmental factors in wheat fields could be one of the mechanisms for the higher biomass and yield under SH cultivation than those under H cultivation. ΦPSII and CO2 exchange rate in wheat flag leaves under SH and H treatments had a linear correlation which could provide new insight to evaluate the wheat photosynthesis performance under different conditions.

Deg1 protease functions in protease and chaperone of PSII complex components, but few works were performed to study the effects of Deg1 on electron transport activities on the donor and acceptor side of PSII and its correlation with the photoprotection of PSII during photoinhibition. Therefore, we performed systematic and comprehensive investigations of electron transfers on the donor and acceptor sides of photosystem II (PSII) in the Deg1-reduced transgenic lines deg1-2 and deg1-4. Both the maximal quantum efficiency of PSII photochemistry (Fv/Fm) and the actual PSII efficiency (ΦPSII) decreased significantly in the transgenic plants. Increases in nonphotochemical quenching (NPQ) and the dissipated energy flux per reaction center (DI0/RC) were also shown in the transgenic plants. Along with the decreased D1, CP47, and CP43 content, these results suggested photoinhibition under growth light conditions in transgenic plants. Decreased Deg1 caused inhibition of electron transfer on the PSII reducing side, leading to a decline in the number of QB-reducing centers and accumulation of QB-nonreducing centers. The Tm of the Q band shifted from 5.7 °C in the wild-type plant to 10.4 °C and 14.2 °C in the deg1-2 and deg1-4 plants, respectively, indicating an increase in the stability of S2QA¯ in transgenic plants. PSIIα in the transgenic plants largely reduced, while PSIIβ and PSIIγ increased with the decline in the Deg1 levels in transgenic plants suggesting PSIIα centers gradually converted into PSIIβ and PSIIγ centers in the transgenic plants. Besides, the connectivity of PSIIα and PSIIβ was downregulated in transgenic plants. Our results reveal that downregulation of Deg1 protein levels induced photoinhibition in transgenic plants, leading to loss of PSII activities on both the donor and acceptor sides in transgenic plants. These results give a new insight into the regulation role of Deg1 in PSII electron transport.

Plastid-encoded RNA polymerase (PEP)-dependent transcription is an essential process for chloroplast development and plant growth. It is a complex event that is regulated by numerous nuclear-encoded proteins. In order to elucidate the complex regulation mechanism of PEP activity, identification and characterization of PEP activity regulation factors are needed. Here, we characterize Plastid Deficient 1 (PD1) as a novel regulator for PEP-dependent gene expression and chloroplast development in Arabidopsis. The PD1 gene encodes a protein that is conserved in photoautotrophic organisms. The Arabidopsis pd1 mutant showed albino and seedling-lethal phenotypes. The plastid development in the pd1 mutant was arrested. The PD1 protein localized in the chloroplasts, and it colocalized with nucleoid protein TRXz. RT-quantitative real-time PCR, northern blot, and run-on analyses indicated that the PEP-dependent transcription in the pd1 mutant was dramatically impaired, whereas the nuclear-encoded RNA polymerase-dependent transcription was up-regulated. The yeast two-hybrid assays and coimmunoprecipitation experiments showed that the PD1 protein interacts with PEP core subunit β (PEP-β), which has been verified to be essential for chloroplast development. The immunoblot analysis indicated that the accumulation of PEP-β was barely detected in the pd1 mutant, whereas the accumulation of the other essential components of the PEP complex, such as core subunits α and β′, were not affected in the pd1 mutant. These observations suggested that the PD1 protein is essential for the accumulation of PEP-β and chloroplast development in Arabidopsis, potentially by direct interaction with PEP-β.