Transcription Factors of the GLRs Family Are Involved in Cytokinin-Dependent Regulation of Plastid RNA Polymerase SCA3 Gene Expression during Deetiolation of Arabidopsis thaliana.

Light-dependent transcription factors GLKs of Arabidopsis thaliana are involved in the anterograde regulation of chloroplast biogenesis during deetiolation: they regulate the expression of photosynthetic nuclear-encoded genes and also mediate the transcription of plastid genes. Chloroplast biogenesis is determined at the same time by light and by endogenous factors (phytohormones), among which cytokinins significantly accelerate the formation of photosynthetically active chloroplasts. In this work, it was shown that trans-factors GLKs function as cytokinin-dependent regulators, mediating the positive cytokinin effect on the plastome expression through the activation of transcription of the SCA3 nuclear gene encoding the plastid RNA polymerase RPOTp.

The Melatonin Receptor CAND2/PMTR1 Is Involved in the Regulation of Mitochondrial Gene Expression under Photooxidative Stress.

Melatonin is a signaling molecule that mediates multiple stress-dependent reactions. Under photooxidative stress conditions generating intensive ROS production, exogenous melatonin (50 µM) contributed to maintaining the expression of mitochondrial encoded genes and up-regulation of RNA-polymerase genes RPOTm and RPOTmp, operating through the CAND2 receptor and α-subunit of the heterotrimeric G protein GPA1 coupled with CAND2. Unlike wild-type plants, mutants with defective CAND2 and GPA1 genes exhibited no decrease in the alternative pathway of leaf respiration, as well as the activity of an alternative oxidase, and the expression of the AOX1a gene. At the same time, the protective effect of exogenous melatonin on some physiological indicators did not depend on the receptor and was associated with the direct antioxidant function of the regulator. Thus, melatonin under photooxidative stress conditions can act as an antioxidant and as a hormone capable of regulating the expression of nuclear and organelle genes through the components of melatonin signal perception.

Ontogenetic, Light, and Circadian Regulation of PAP Protein Genes during Seed Germination of Arabidopsis thaliana.

The expression profiles of the PAP genes, encoding proteins associated with plastid multisubunit RNA polymerase, were studied in dry seeds, during germination, and at the early stages of Arabidopsis thaliana seedling formation. A detailed analysis of the PAP transcript levels by RT-PCR showed that the transition of seeds from dormancy to active growth is accompanied by a drastic increase in the transcript accumulation of all studied genes on the first day of germination, both in the light and in the dark. Further changes in transcript levels differed among PAP genes and were apparently determined by their functional specificity. It was established for the first time that the expression of individual PAP genes is regulated by circadian rhythms, in addition to factors of ontogenetic and light nature.

Cold Stress Activates the Expression of Genes of the Chloroplast Transcription Apparatus in Arabidopsis thaliana Plants.

The physiological and molecular responses of Arabidopsis thaliana plants to cold stress were studied. Exposure to a low non-freezing temperature (4°C, 5 days) caused a decrease in the physiological functions and activity of a number of photosynthetic genes and elevation in expression of the cold stress gene COR15a, the product of which protects chloroplasts. It was shown for the first time that in parallel to a general inhibition of physiological functions under hypothermia, an increase in the expression of most genes for the chloroplast transcription apparatus was observed. This is obviously one of the compensatory mechanisms of adaptation aimed to maintain cellular homeostasis and physiological functions under hypothermia.

The Transcription Factor HY5 Is Involved in the Cytokinin-Dependent Regulation of the Expression of Genes Encoding Proteins Associated with Bacterial Plastid RNA-Polymerase during De-Etiolation of Arabidopsis thaliana.

HY5 (ELONGATED HYPOCOTYL5), a bZIP transcription factor, is one of the main regulators of light and hormonal signaling. Among the targets of this gene, the genes for the transcriptional complex of chloroplasts whose coordinated expression ensures the initial stages of photomorphogenesis are particularly significant. In this study, we showed that, during de-etiolation, HY5 functions as a positive CK-dependent regulator of the expression of genes encoding proteins associated with plastid RNA polymerase (PAP), which functions below the primary chain of sensing the cytokinin signal. The absence of blocking effect of mutations of the CRY1, CRY2, PHYA, and PHYB photoreceptor genes on the CK-dependent content of PAP gene transcripts indicates the parallel action of the hormone and light in their regulation.

Cytokinins and Abscisic Acid Regulate the Expression of the Genes for Plastid Transcription Apparatus during Heat Shock.

The treatment of Arabidopsis thaliana plants with exogenous cytokinin (CK) followed by heat shock (HS) activated the expression of the genes for the plastid transcription machinery but adversely affected the plant viability. Abscisic acid (ABA), conversely, promoted maintaining the resistance to HS and had differentially affected different components of the plastid transcriptional complex. This hormone suppressed the accumulation of transcripts of PEP genes and the genes encoding PAP proteins, which are involved in DNA-RNA metabolism. However, it had no effect or activated the expression of NEP genes and PAP genes, which are involved in the redox regulation, as well as the genes encoding the stress-inducible trans-factor (SIG5) and the plastid transcription Ser/Thr protein kinase (cpCK2). Thus, for the adaptation of plants to elevated temperatures, both increase and decrease in the expression of the genes for the plastid transcriptional machinery with the involvement of various regulatory systems, including phytohormones, are equally significant.

Phytohormones Regulate the Expression of Nuclear Genes Encoding the Components of the Plastid Transcription Apparatus.

As multifunctional regulators of physiological processes, phytohormones play an important role in the regulation of expression of the plastid genome and chloroplast biogenesis. Hormones can directly regulate the expression of genes localized in the chloroplast genome. However, many components of the plastid transcription apparatus are encoded by nuclear rather than plastid genes. It remains obscure whether these nuclear genes are subject to hormonal regulation. This is the first study to show that phytohormones exert differential effects on the expression of nuclear genes of the transcription machinery of the Arabidopsis thaliana plastome. RT-PCR analysis showed that the level of transcripts of the majority of studied genes was activated by trans-zeatin but decreased under the influence of ABA, methyl jasmonate, and salicylic acid, whereas ethylene had no significant effect, and the effects of brassinolide depended on the illumination conditions. The results of this study indicate that the hormonal regulation of the plastome expression can be mediated by differential regulation of the nuclear genes encoding plastid transcription machinery components.

Cytokinin membrane receptors participate in regulation of plastid genome expression in the skotomorphogenesis.

Analysis by real-time PCR of single and double insertion mutants of A. thaliana with inactivated cytokinin receptor genes showed that the level of transcripts of some of plastid genes during skotomorphogenesis depended on the state of functionally active receptor AHK3. The cytokinin-regulated plastid encoded genes involved the housekeeping genes (rpoB and accD) and the genes for photosynthetic proteins (ndhA, psbA, atpB, and psaA). However, the absence of hormone activation of plastid encoded genes engaged in the translation of plastid proteins, rRNA (rrn16), and tRNA (trnE), indirectly indicate the disruption of the synthesis of chloroplast proteins in the dark.

The spinach AtpC and AtpD genes contain elements for light-regulated, plastid-dependent and organ-specific expression in the vicinity of the transcription start sites.

Run-on assays with isolated nuclei demonstrate that the transcription rates of AtpC and AtpD (gene products: the CF1 subunits gamma and delta of the chloroplast ATP synthase) are comparable in spinach seedlings. However, chimeric GUS gene fusions with 5'-flanking regions of the AtpC gene direct an approximately 10-fold lower GUS level in transgenic tobacco compared with equivalent fragments from the AtpD gene. Both promoters contain sequences in the vicinity of the respective TATA boxes, which are sufficient to direct light-regulated, plastid-dependent and organ-specific expression of the GUS gene. In contrast, the upstream regions of both promoters differ the higher GUS level directed by the AtpD promoter is caused by enhancer-like elements located upstream of the region involved in the regulated expression, while nucleotides upstream of -73 in the AtpC promoter contribute relatively little to the promoter activity. 5'-Deletion analyses and site-directed mutagenesis studies indicated that the -73/-48 bp AtpC region contains cis-elements crucial for this regulated expression. If five nucleotides within this region (-59/-55) are exchanged, the GUS gene is constitutively expressed and the activity in etiolated seedlings, in seedlings with photobleached plastids and in roots increases to the level detectable in green cotyledons. It is concluded that signal transduction pathways from different regulators converge prior to gene regulation and that these five nucleotides are part of a cis-element which functions as a repressor in darkness, in tissues with impaired plastids and in roots.

Changes in the level of chloroplast transcripts in pumpkin cotyledons during heat shock.

The levels of plastid gene transcripts are shown to be controlled by temperature in isolated pumpkin cotyledons. The temperature at which maximum transcript accumulation occurs varies between 38 and 42 degrees C for the genes studied (rbcL, psaA, psbA, psbB, psbC, psbD, psbE, atpA). Heat shock-induced transcript accumulation is transitory with a maximum after an approximately 3 h exposure at high temperatures. On the other hand, the accumulation of rbcL transcript was only moderately thermosensitive. A temperature increase to 46-48 degrees C induces a sharp decrease of transcript levels which correlates with damage to the plant. Relatively little correlation has been noted between RNA and protein patterns. However, there is a remarkable coincidence between temperature dependence of the accumulation of transcripts and the temperature dependence (28-46 degrees C) of the synthesis of chloroplast-located heat shock proteins indicating that both processes may be related.