Length-Dependent Translation Efficiency of ER-Destined Proteins.

Gene expression is a fundamental process that enables cells to produce specific proteins in a timely and spatially dependent manner. In eukaryotic cells, the complex organization of the cell body requires precise control of protein synthesis and localization. Certain mRNAs encode proteins with an N-terminal signal sequences that direct the translation apparatus toward a specific organelle. Here, we focus on the mechanisms governing the translation of mRNAs, which encode proteins with an endoplasmic reticulum (ER) signal in human cells. The binding of a signal-recognition particle (SRP) to the translation machinery halts protein synthesis until the mRNA-ribosome complex reaches the ER membrane. The commonly accepted model suggests that mRNA that encodes a protein that contains an ER signal peptide continuously repeats the cycle of SRP binding followed by association and dissociation with the ER. In contrast to the current view, we show that the long mRNAs remain on the ER while being translated. On the other hand, due to low ribosome occupancy, the short mRNAs continue the cycle, always facing a translation pause. Ultimately, this leads to a significant drop in the translation efficiency of small, ER-targeted proteins. The proposed mechanism advances our understanding of selective protein synthesis in eukaryotic cells and provides new avenues to enhance protein production in biotechnological settings.

Light Drives and Temperature Modulates: Variation of Phenolic Compounds Profile in Relation to Photosynthesis in Spring Barley.

Accumulation and metabolic profile of phenolic compounds (PheCs; serving as UV-screening pigments and antioxidants) as well as carbon fixation rate (An) and plant growth are sensitive to irradiance and temperature. Since these factors are naturally co-acting in the environment, it is worthy to study the combined effects of these environmental factors to assess their possible physiological consequences. We investigated how low and high irradiance in combination with different temperatures modify the metabolic profile of PheCs and expression of genes involved in the antioxidative enzyme and PheCs biosynthesis, in relation to photosynthetic activity and availability of non-structural carbohydrates (NSC) in spring barley seedlings. High irradiance positively affected An, NSC, PheCs content, and antioxidant activity (AOX). High temperature led to decreased An, NSC, and increased dark respiration, whilst low temperature was accompanied by reduction of UV-A shielding but increase of PheCs content and AOX. Besides that, irradiance and temperature caused changes in the metabolic profile of PheCs, particularly alteration in homoorientin/isovitexin derivatives ratio, possibly related to demands on AOX-based protection. Moreover, we also observed changes in the ratio of sinapoyl-/feruloyl- acylated flavonoids, the function of which is not yet known. The data also strongly suggested that the NSC content may support the PheCs production.

Regulation of Phenolic Compound Production by Light Varying in Spectral Quality and Total Irradiance.

Photosynthetically active radiation (PAR) is an important environmental cue inducing the production of many secondary metabolites involved in plant oxidative stress avoidance and tolerance. To examine the complex role of PAR irradiance and specific spectral components on the accumulation of phenolic compounds (PheCs), we acclimated spring barley (Hordeum vulgare) to different spectral qualities (white, blue, green, red) at three irradiances (100, 200, 400 µmol m-2 s-1). We confirmed that blue light irradiance is essential for the accumulation of PheCs in secondary barley leaves (in UV-lacking conditions), which underpins the importance of photoreceptor signals (especially cryptochrome). Increasing blue light irradiance most effectively induced the accumulation of B-dihydroxylated flavonoids, probably due to the significantly enhanced expression of the F3'H gene. These changes in PheC metabolism led to a steeper increase in antioxidant activity than epidermal UV-A shielding in leaf extracts containing PheCs. In addition, we examined the possible role of miRNAs in the complex regulation of gene expression related to PheC biosynthesis.

Accumulation of geranylgeranylated chlorophylls in the pigment-protein complexes of Arabidopsis thaliana acclimated to green light: effects on the organization of light-harvesting complex II and photosystem II functions.

Light quality significantly influences plant metabolism, growth and development. Recently, we have demonstrated that leaves of barley and other plant species grown under monochromatic green light (500-590 nm) accumulated a large pool of chlorophyll a (Chl a) intermediates with incomplete hydrogenation of their phytyl chains. In this work, we studied accumulation of these geranylgeranylated Chls a and b in pigment-protein complexes (PPCs) of Arabidopsis plants acclimated to green light and their structural-functional consequences on the photosynthetic apparatus. We found that geranylgeranylated Chls are present in all major PPCs, although their presence was more pronounced in light-harvesting complex II (LHCII) and less prominent in supercomplexes of photosystem II (PSII). Accumulation of geranylgeranylated Chls hampered the formation of PSII and PSI super- and megacomplexes in the thylakoid membranes as well as their assembly into chiral macrodomains; it also lowered the temperature stability of the PPCs, especially that of LHCII trimers, which led to their monomerization and an anomaly in the photoprotective mechanism of non-photochemical quenching. Role of geranylgeranylated Chls in adverse effects on photosynthetic apparatus of plants acclimated to green light is discussed.

G-Quadruplex in Gene Encoding Large Subunit of Plant RNA Polymerase II: A Billion-Year-Old Story.

G-quadruplexes have long been perceived as rare and physiologically unimportant nucleic acid structures. However, several studies have revealed their importance in molecular processes, suggesting their possible role in replication and gene expression regulation. Pathways involving G-quadruplexes are intensively studied, especially in the context of human diseases, while their involvement in gene expression regulation in plants remains largely unexplored. Here, we conducted a bioinformatic study and performed a complex circular dichroism measurement to identify a stable G-quadruplex in the gene RPB1, coding for the RNA polymerase II large subunit. We found that this G-quadruplex-forming locus is highly evolutionarily conserved amongst plants sensu lato (Archaeplastida) that share a common ancestor more than one billion years old. Finally, we discussed a new hypothesis regarding G-quadruplexes interacting with UV light in plants to potentially form an additional layer of the regulatory network.

Do UV-A radiation and blue light during growth prime leaves to cope with acute high light in photoreceptor mutants of Arabidopsis thaliana?

We studied how plants acclimated to growing conditions that included combinations of blue light (BL) and ultraviolet (UV)-A radiation, and whether their growing environment affected their photosynthetic capacity during and after a brief period of acute high light (as might happen during an under-canopy sunfleck). Arabidopsis thaliana Landsberg erecta wild-type were compared with mutants lacking functional blue light and UV photoreceptors: phototropin 1, cryptochromes (CRY1 and CRY2) and UV RESISTANT LOCUS 8 (uvr8). This was achieved using light-emitting-diode (LED) lamps in a controlled environment to create treatments with or without BL, in a split-plot design with or without UV-A radiation. We compared the accumulation of phenolic compounds under growth conditions and after exposure to 30 min of high light at the end of the experiment (46 days), and likewise measured the operational efficiency of photosystem II (ϕPSII, a proxy for photosynthetic performance) and dark-adapted maximum quantum yield (Fv /Fm to assess PSII damage). Our results indicate that cryptochromes are the main photoreceptors regulating phenolic compound accumulation in response to BL and UV-A radiation, and a lack of functional cryptochromes impairs photosynthetic performance under high light. Our findings also reveal a role for UVR8 in accumulating flavonoids in response to a low UV-A dose. Interestingly, phototropin 1 partially mediated constitutive accumulation of phenolic compounds in the absence of BL. Low-irradiance BL and UV-A did not improve ϕPSII and Fv /Fm upon our acute high-light treatment; however, CRYs played an important role in ameliorating high-light stress.

Beyond the Primary Structure of Nucleic Acids: Potential Roles of Epigenetics and Noncanonical Structures in the Regulations of Plant Growth and Stress Responses.

Epigenetics deals with changes in gene expression that are not caused by modifications in the primary sequence of nucleic acids. These changes beyond primary structures of nucleic acids not only include DNA/RNA methylation, but also other reversible conversions, together with histone modifications or RNA interference. In addition, under particular conditions (such as specific ion concentrations or protein-induced stabilization), the right-handed double-stranded DNA helix (B-DNA) can form noncanonical structures commonly described as "non-B DNA" structures. These structures comprise, for example, cruciforms, i-motifs, triplexes, and G-quadruplexes. Their formation often leads to significant differences in replication and transcription rates. Noncanonical RNA structures have also been documented to play important roles in translation regulation and the biology of noncoding RNAs. In human and animal studies, the frequency and dynamics of noncanonical DNA and RNA structures are intensively investigated, especially in the field of cancer research and neurodegenerative diseases. In contrast, noncanonical DNA and RNA structures in plants have been on the fringes of interest for a long time and only a few studies deal with their formation, regulation, and physiological importance for plant stress responses. Herein, we present a review focused on the main fields of epigenetics in plants and their possible roles in stress responses and signaling, with special attention dedicated to noncanonical DNA and RNA structures.

Deubiquitinase OTUD1 Resolves Stalled Translation on polyA and Rare Codon Rich mRNAs.

OTUD1 is a deubiquitinating enzyme involved in many cellular processes including cancer and innate, immune signaling pathways. Here, we perform a proximity labeling-based interactome study that identifies OTUD1 largely present in the translation and RNA metabolism protein complexes. Biochemical analysis validates OTUD1 association with ribosome subunits, elongation factors and the E3 ubiquitin ligase ZNF598 but not with the translation initiation machinery. OTUD1 catalytic activity suppresses polyA triggered ribosome stalling through inhibition of ZNF598-mediated RPS10 ubiquitination and stimulates formation of polysomes. Finally, analysis of gene expression suggests that OTUD1 regulates the stability of rare codon rich mRNAs by antagonizing ZNF598.

Searching for G-Quadruplex-Binding Proteins in Plants: New Insight into Possible G-Quadruplex Regulation.

G-quadruplexes are four-stranded nucleic acid structures occurring in the genomes of all living organisms and viruses. It is increasingly evident that these structures play important molecular roles; generally, by modulating gene expression and overall genome integrity. For a long period, G-quadruplexes have been studied specifically in the context of human promoters, telomeres, and associated diseases (cancers, neurological disorders). Several of the proteins for binding G-quadruplexes are known, providing promising targets for influencing G-quadruplex-related processes in organisms. Nonetheless, in plants, only a small number of G-quadruplex binding proteins have been described to date. Thus, we aimed to bioinformatically inspect the available protein sequences to find the best protein candidates with the potential to bind G-quadruplexes. Two similar glycine and arginine-rich G-quadruplex-binding motifs were described in humans. The first is the so-called "RGG motif"-RRGDGRRRGGGGRGQGGRGRGGGFKG, and the second (which has been recently described) is known as the "NIQI motif"-RGRGRGRGGGSGGSGGRGRG. Using this general knowledge, we searched for plant proteins containing the above mentioned motifs, using two independent approaches (BLASTp and FIMO scanning), and revealed many proteins containing the G4-binding motif(s). Our research also revealed the core proteins involved in G4 folding and resolving in green plants, algae, and the key plant model organism, Arabidopsis thaliana. The discovered protein candidates were annotated using STRINGdb and sorted by their molecular and physiological roles in simple schemes. Our results point to the significant role of G4-binding proteins in the regulation of gene expression in plants.

Induction of phenolic compounds by UV and PAR is modulated by leaf ontogeny and barley genotype.

We investigated the effect of leaf ontogeny and barley genotype on the accumulation of phenolic compounds (PhCs) induced by ultraviolet (UV) and photosynthetically active radiation (PAR). We hypothesized that different groups of PhCs are induced in leaves differing in ontogeny, and that this has consequences for protective functions and the need for other protection mechanisms. Generally, lower constitutive contents of PhCs (under conditions of UV exclusion and reduced PAR) were found in a UV-sensitive genotype (Barke) compared to a tolerant genotype (Bonus). However, UV and PAR induced accumulation of PhCs exceeded the constitutive amounts several fold. Specifically, lutonarin, 3-feruloylquinic acid, unidentified hydroxycinnamic acid and luteolin derivatives were markedly enhanced by high PAR and UV irradiances. Leaves developed during UV and PAR treatments had higher PhCs contents than mature leaves already fully developed at the onset of the UV and PAR treatment. UV and PAR treatments had, however, a minor effect on saponarin and unidentified apigenin derivatives which occur particularly in mature leaves of the tolerant genotype Bonus. In addition, high UV and PAR intensities increased the total content of xanthophylls (VAZ), while chlorophyll content was reduced, particularly in developing leaves. A redundancy analysis revealed positive associations between most of PhCs and VAZ and a negative association between total chlorophylls and carotenoids. Non-linear relationships between VAZ and lutonarin and other PhCs indicate that VAZ accumulation can compensate for the insufficient efficiency of anti-oxidative protection mediated by PhCs. Accordingly, we conclude that UV and PAR-induced accumulation of PhCs is affected by leaf ontogeny, however, this effect is compound-specific.

Monochromatic green light induces an aberrant accumulation of geranylgeranyled chlorophylls in plants.

Light quality is an important environmental factor affecting the biosynthesis of photosynthetic pigments whose production seems to be affected not only quantitatively but also qualitatively. In this work, we set out to identify unusual pigment detected in leaves of barley (Hordeum vulgare L.) and explain its presence in plants grown under monochromatic green light (GL; 500-590 nm). The chromatographic analysis (HPLC-DAD) revealed that a peak belonging to this unknown pigment is eluted between chlorophyll (Chl) a and b. This pigment exhibited the same absorption spectrum and fluorescence excitation and emission spectra as Chl a. It was negligible in control plants cultivated under white light of the same irradiance (photosynthetic photon flux density of 240 µmol m-2 s-1). Mass spectrometry analysis of this pigment (ions m/z = 889 [M-H]-; m/z = 949 [M+acetic acid-H]-) indicates that it is Chl a with a tetrahydrogengeranylgeraniol side chain (containing two double bonds in a phytyl side chain; Chl aTHGG), which is an intermediate in Chl a synthesis. In plants grown under GL, the proportion of Chl aTHGG to total Chl content rose to approximately 8% and 16% after 7 and 14 days of cultivation, respectively. Surprisingly, plants cultivated under GL exhibited drastically increased concentration of the enzyme geranylgeranyl reductase, which is responsible for the reduction of phytyl chain double bonds in the Chl synthesis pathway. This indicates impaired activity of this enzyme in GL-grown plants. A similar effect of GL on Chl synthesis was observed for distinct higher plant species.

Protective effect of UV-A radiation during acclimation of the photosynthetic apparatus to UV-B treatment.

We examined the acclimation response of the photosynthetic apparatus of barley (Hordeum vulgare L.) to a combination of UV-A and UV-B radiation (UVAB) and to UV-B radiation alone. Our aim was to evaluate whether UV-A radiation prevents UV-B-induced damage to the photosynthetic apparatus and whether UV-A pre-acclimation is required to mitigate the negative influence of UV-B radiation. Barley plants were grown from seeds under low photosynthetically active radiation (50 µmol m(-2) s(-1)) either in the absence or presence of UV-A radiation (UVA- and UVA+ plants, respectively). After 8 days of development, plants were exposed simultaneously to UV-A and UV-B radiation for the next 6 days. Additionally, UVA- plants were exposed to UV-B radiation alone. The UVA+ plants had a higher CO2 assimilation rate near the light-saturation region (A(N)) and a higher content of both total chlorophylls (Chls) and total carotenoids than the UVA- plants. Chls content, A(N), the potential quantum yield of photosystem II (PSII) photochemistry (F(V)/F(M)), the capacity of light-induced thermal energy dissipation and the efficiency of excitation energy transfer within PSII remained the same or even increased in both UVA+ and UVA- plants after UVAB treatment. On the contrary, exposure of UVA- plants to UV-B radiation itself led to a reduction in all these characteristics. We revealed that the presence of UV-A radiation during UVAB treatment not only mitigated but completely eliminated the negative effect of UV-B radiation on the functioning of the photosynthetic apparatus and that UV-A pre-acclimation was not crucial for development of this UV-A-induced resistance against UV-B irradiation.

Ultraviolet and photosynthetically active radiation can both induce photoprotective capacity allowing barley to overcome high radiation stress.

The main objective of this study was to determine the effects of acclimation to ultraviolet (UV) and photosynthetically active radiation (PAR) on photoprotective mechanisms in barley leaves. Barley plants were acclimated for 7 days under three combinations of high or low UV and PAR treatments ([UV-PAR-], [UV-PAR+], [UV+PAR+]). Subsequently, plants were exposed to short-term high radiation stress (HRS; defined by high intensities of PAR - 1000 µmol m(-2) s(-1), UV-A - 10 W m(-2) and UV-B 2 W m(-2) for 4 h), to test their photoprotective capacity. The barley variety sensitive to photooxidative stress (Barke) had low constitutive flavonoid content compared to the resistant variety (Bonus) under low UV and PAR intensities. The accumulation of lutonarin and 3-feruloylquinic acid, but not of saponarin, was greatly enhanced by high PAR and further increased by UV exposure. Acclimation of plants to both high UV and PAR intensities also increased the total pool of xanthophyll-cycle pigments (VAZ). Subsequent exposure to HRS revealed that prior acclimation to UV and PAR was able to ameliorate the negative consequences of HRS on photosynthesis. Both total contents of epidermal flavonols and the total pool of VAZ were closely correlated with small reductions in light-saturated CO2 assimilation rate and maximum quantum yield of photosystem II photochemistry caused by HRS. Based on these results, we conclude that growth under high PAR can substantially increase the photoprotective capacity of barley plants compared with plants grown under low PAR. However, additional UV radiation is necessary to fully induce photoprotective mechanisms in the variety Barke. This study demonstrates that UV-exposure can lead to enhanced photoprotective capacity and can contribute to the induction of tolerance to high radiation stress in barley.