Ritesh Ghosh

Differential acetylation of histone H3 at the regulatory region of OsDREB1b promoter facilitates chromatin remodelling and transcription activation during cold stress.

The rice ortholog of DREB1, OsDREB1b, is transcriptionally induced by cold stress and over-expression of OsDREB1b results in increase tolerance towards high salt and freezing stress. This spatio-temporal expression of OsDREB1b is preceded by the change in chromatin structure at the promoter and the upstream region for gene activation. The promoter and the upstream region of OsDREB1b genes appear to be arranged into a nucleosome array. Nucleosome mapping of ∼700bp upstream region of OsDREB1b shows two positioned nucleosomes between −610 to −258 and a weakly positioned nucleosome at the core promoter and the TSS. Upon cold stress, there is a significant change in the nucleosome arrangement at the upstream region with increase in DNaseI hypersensitivity or MNase digestion in the vicinity of cis elements and TATA box at the core promoter. ChIP assays shows hyper-acetylation of histone H3K9 throughout the locus whereas region specific increase was observed in H3K14ac and H3K27ac. Moreover, there is an enrichment of RNA PolII occupancy at the promoter region during transcription activation. There is no significant change in the H3 occupancy in OsDREB1b locus negating the possibility of nucleosome loss during cold stress. Interestingly, cold induced enhanced transcript level of OsDREB1b as well as histone H3 acetylation at the upstream region was found to diminish when stressed plants were returned to normal temperature. The result indicates absolute necessity of changes in chromatin conformation for the transcription up-regulation of OsDREB1b gene in response to cold stress. The combined results show the existence of closed chromatin conformation at the upstream and promoter region of OsDREB1b in the transcription “off” state. During cold stress, changes in region specific histone modification marks promote the alteration of chromatin structure to facilitate the binding of transcription machinery for proper gene expression.

Plant acoustics: in the search of a sound mechanism for sound signaling in plants.

Being sessile, plants continuously deal with their dynamic and complex surroundings, identifying important cues and reacting with appropriate responses. Consequently, the sensitivity of plants has evolved to perceive a myriad of external stimuli, which ultimately ensures their successful survival. Research over past centuries has established that plants respond to environmental factors such as light, temperature, moisture, and mechanical perturbations (e.g. wind, rain, touch, etc.) by suitably modulating their growth and development. However, sound vibrations (SVs) as a stimulus have only started receiving attention relatively recently. SVs have been shown to increase the yields of several crops and strengthen plant immunity against pathogens. These vibrations can also prime the plants so as to make them more tolerant to impending drought. Plants can recognize the chewing sounds of insect larvae and the buzz of a pollinating bee, and respond accordingly. It is thus plausible that SVs may serve as a long-range stimulus that evokes ecologically relevant signaling mechanisms in plants. Studies have suggested that SVs increase the transcription of certain genes, soluble protein content, and support enhanced growth and development in plants. At the cellular level, SVs can change the secondary structure of plasma membrane proteins, affect microfilament rearrangements, produce Ca(2+) signatures, cause increases in protein kinases, protective enzymes, peroxidases, antioxidant enzymes, amylase, H(+)-ATPase / K(+) channel activities, and enhance levels of polyamines, soluble sugars and auxin. In this paper, we propose a signaling model to account for the molecular episodes that SVs induce within the cell, and in so doing we uncover a number of interesting questions that need to be addressed by future research in plant acoustics.

Exposure to Sound Vibrations Lead to Transcriptomic, Proteomic and Hormonal Changes in Arabidopsis

Sound vibration (SV) is considered as an external mechanical force that modulates plant growth and development like other mechanical stimuli (e.g., wind, rain, touch and vibration). A number of previous and recent studies reported developmental responses in plants tailored against SV of varied frequencies. This strongly suggests the existence of sophisticated molecular mechanisms for SV perception and signal transduction. Despite this there exists a huge gap in our understanding regarding the SV-mediated molecular alterations, which is a prerequisite to gain insight into SV-mediated plant development. Herein, we investigated the global gene expression changes in Arabidopsis thaliana upon treatment with five different single frequencies of SV at constant amplitude for 1 h. As a next step, we also studied the SV-mediated proteomic changes in Arabidopsis. Data suggested that like other stimuli, SV also activated signature cellular events, for example, scavenging of reactive oxygen species (ROS), alteration of primary metabolism, and hormonal signaling. Phytohormonal analysis indicated that SV-mediated responses were, in part, modulated by specific alterations in phytohormone levels; especially salicylic acid (SA). Notably, several touch regulated genes were also up-regulated by SV treatment suggesting a possible molecular crosstalk among the two mechanical stimuli, sound and touch. Overall, these results provide a molecular basis to SV triggered global transcriptomic, proteomic and hormonal changes in plant.

Multiple Reaction Monitoring Mode Based Liquid Chromatography-Mass Spectrometry Method for Simultaneous Quantification of Brassinolide and Other Plant Hormones Involved in Abiotic Stresses

Plant hormones are the key regulators of adaptive stress response. Abiotic stresses such as drought and salt are known to affect the growth and productivity of plants. It is well known that the levels of plant hormones such as zeatin (ZA), abscisic acid (ABA), salicylic acid (SA), jasmonic acid (JA), and brassinolide (BR) fluctuate upon abiotic stress exposure. At present, there is not any single suitable liquid chromatography-mass spectrometry (LC-MS) method for simultaneous analysis of BR and other plant hormones involved in abiotic stresses. In the present study, we developed a simple, sensitive, and rapid method for simultaneous analysis of five major plant hormones, ZA, ABA, JA, SA, and BR, which are directly or indirectly involved in drought and salt stresses. The optimized extraction procedure was simple and easy to use for simultaneous measurement of these plant hormones in Arabidopsis thaliana. The developed method is highly reproducible and can be adapted for simultaneous measurement of changes in plant hormones (ZA, ABA, JA, SA, and BR) in response to abiotic stresses in plants like A. thaliana and tomato.

Expression Analysis of Sound Vibration-Regulated Genes by Touch Treatment in Arabidopsis

Sound vibration (SV) is considered to be a mechanical stimulus which gives rise to various physiological and molecular changes in plants. Previously, we identified 17 SV-regulated genes (SRGs) which were up-regulated by SV treatments in Arabidopsis. Here, we analyzed the expression pattern of similar genes after an exposure of 500 Hertz at 80 decibels, for various time periods. Simultaneously, we confirmed the SV-mediated expression of these genes under lighted condition as many of them were reported to be dark-induced. For this, we designed an improved SV treatment chamber. Additionally, we checked the electrolyte leakage (EL), photosynthetic performance and expression of mechanosensitive (MS) ion channel genes after 5 days of SV treatment in the illuminated chamber. EL was higher, and the photosynthetic performance index was lower in the SV-treated plants compared to control. Seven out of the 13 MS ion channel genes were differentially expressed after the SV treatment. Simultaneously, we checked the touch-mediated expression pattern of 17 SRGs and 13 MS ion channel genes. The distinct expression pattern of 6 SRGs and 1 MS ion channel gene generate an idea that SV as a stimulus is different from touch. Developmental stage-specific expression profiling suggested that the majority of the SRGs were expressed spatiotemporally in different developmental stages of Arabidopsis, especially in imbibed seed, seedlings and leaves.

Characterization of Developmental- and Stress-Mediated Expression of Cinnamoyl-CoA Reductase in Kenaf (Hibiscus cannabinus L.)

Cinnamoyl-CoA reductase (CCR) is an important enzyme for lignin biosynthesis as it catalyzes the first specific committed step in monolignol biosynthesis. We have cloned a full length coding sequence of CCR from kenaf (Hibiscus cannabinus L.), which contains a 1,020-bp open reading frame (ORF), encoding 339 amino acids of 37.37 kDa, with an isoelectric point (pI) of 6.27 (JX524276, HcCCR2). BLAST result found that it has high homology with other plant CCR orthologs. Multiple alignment with other plant CCR sequences showed that it contains two highly conserved motifs: NAD(P) binding domain (VTGAGGFIASWMVKLLLEKGY) at N-terminal and probable catalytic domain (NWYCYGK). According to phylogenetic analysis, it was closely related to CCR sequences of Gossypium hirsutum (ACQ59094) and Populus trichocarpa (CAC07424). HcCCR2 showed ubiquitous expression in various kenaf tissues and the highest expression was detected in mature flower. HcCCR2 was expressed differentially in response to various stresses, and the highest expression was observed by drought and NaCl treatments.

Positive regulatory role of sound vibration treatment in Arabidopsis thaliana against Botrytis cinerea infection

Sound vibration (SV), a mechanical stimulus, can trigger various molecular and physiological changes in plants like gene expression, hormonal modulation, induced antioxidant activity and calcium spiking. It also alters the seed germination and growth of plants. In this study, we investigated the effects of SV on the resistance of Arabidopsis thaliana against Botrytis cinerea infection. The microarray analysis was performed on infected Arabidopsis plants pre-exposed to SV of 1000 Hertz with 100 decibels. Broadly, the transcriptomic analysis revealed up-regulation of several defense and SA-responsive and/or signaling genes. Quantitative real-time PCR (qRT-PCR) analysis of selected genes also validated the induction of SA-mediated response in the infected Arabidopsis plants pre-exposed to SV. Corroboratively, hormonal analysis identified the increased concentration of salicylic acid (SA) in the SV-treated plants after pathogen inoculation. In contrast, jasmonic acid (JA) level in the SV-treated plants remained stable but lower than control plants during the infection. Based on these findings, we propose that SV treatment invigorates the plant defense system by regulating the SA-mediated priming effect, consequently promoting the SV-induced resistance in Arabidopsis against B. cinerea.

Erratum to: Deciphering the role of the AT-rich interaction domain and the HMG-box domain of ARID-HMG proteins of Arabidopsis thaliana

1 Division of Plant Biology, Bose Institute, Kolkata 700054, India 2 School of Biotechnology, Yeungnam University, Gyeongsan 712-749, South Korea 3 Department of Biophysics, Bose Institute, Kolkata 700054, India 4 Department of Chemistry, Bose Institute, Kolkata 700054, India Published online: 5 September 2016

Developmental- and stress-mediated expression analysis of cinnamoyl-CoA reductase 1 (CCR1) from Hibiscus cannabinus

Cinnamoyl-CoA reductase (CCR, EC 1.2.1.44) is an important enzyme responsible for lignin biosynthesis in plants that belongs to the family of oxidoreductases. We analyzed developmental, tissue specific, and stress-mediated expression of the HcCCR1 (HM151381) gene from Hibiscus cannabinus. Gene expression analysis revealed that HcCCR1 was highly upregulated in mature leaves of 16-week-old plants. The maximum downregulation and upregulation of HcCCR1 was caused by cold and MeJA treatment, respectively. Sequence analysis demonstrated that HcCCR1 protein (ADK24219) contains a conserved NWYCYGK catalytic domain, while bioinformatics prediction indicated the presence of a palmitoylation site in the HcCCR1 protein. Phylogenetic analysis showed that HcCCR1 is more closely related to HcCCR2 (AGJ84130) and AtCCR proteins than CCR-like proteins. Comparative sequence analysis showed presence of significant differences between HcCCR1 and HcCCR2, which are homologs of H. cannabinus. Expression analysis demonstrated that the HcCCR1 gene is modulated by different external stresses.

Chlorophyll-a fluorescence evaluation of PEG-induced osmotic stress on PSII activity in Arabidopsis plants expressing SIP1

Abstract In this study, we evaluated the effect of osmotic stress on photosynthetic machinery of Arabidopsis plants expressing a gene encoding small basic intrinsic protein (SIP1) isolated from Solanum tuberosum. Intact leaves of SIP Arabidopsis plants were exposed to 15% polyethylene glycol (PEG) solution and fast Chlorophyll-a (Chl-a) fluorescence induction kinetics was measured. Photosynthetic parameters like ratio of variable and maximum fluorescence (FV/FM), absorbance of photons per active reaction center (ABS/RC), trapping of photons per active reaction center (TRo/RC), electron transport per active reaction center (ETo/RC), and performance index (PI) were measured. Furthermore, the energy pipeline model was deduced in response to PEG stress. The membrane model includes a visualization of the average “antenna size”, which follows the value of the ABS/RC. Analysis of SIP Arabidopsis plants under PEG stress through fast Chl-a fluorescence transient showed that the damage caused due to PEG is more prominent at the donor side rather than the acceptor side of PSII. Higher PI in SIP plants under PEG stress indicated a better vitality than control plants. Overall, these results indicate that constitutive expression of SIP1 in Arabidopsis plants induces significant changes in the photosynthetic machinery under PEG-induced osmotic stress.