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Oxygen deprivation by waterlogging reduces the productivity of several crop species, including the oil-producing crop Brassica napus L., which is highly sensitive to excess moisture. Among factors induced by oxygen deficiency are phytoglobins (Pgbs), heme-containing proteins known to ameliorate the response of plants to the stress. This study examined the early responses to waterlogging in B. napus plants over-expressing or down-regulating the class 1 (BnPgb1) and class 2 (BnPgb2) Pgbs. The depression of gas exchange parameters and plant biomass was exacerbated by the suppression of BnPgb1, while suppression of BnPgb2 did not evoke any changes. This suggests that natural occurring levels of BnPgb1 (but not BnPg2) are required for the response of the plants to waterlogging. Typical waterlogging symptoms, including the accumulation of reactive oxygen species (ROS) and the deterioration of the root apical meristem (RAM) were attenuated by over-expression of BnPgb1. These effects were associated with the activation of antioxidant system and the transcriptional induction of folic acid (FA). Pharmacological treatments revealed that high levels of FA were sufficient to revert the inhibitory effect of waterlogging, suggesting that the interplay between BnPgb1, antioxidant responses and FA might contribute to plant tolerance to waterlogging stress., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Combining physiological, molecular and biochemical approaches, this study investigated the transcriptional coordination and abscisic acid (ABA) mediated regulation of genes involved in sucrose import and its conversion to starch during grain filling in wheat. Sucrose import appears to be mediated by seed localized TaSUT1, mainly TaSUT1D, while sucrose cleavage by TaSuSy2. Temporal overlapping of the transcriptional activation of AGPL1 and AGPS1a that encode AGPase with that of the above genes suggests their significance in the synthesis of ADP-glucose; TaAGPL1A and TaAGPL1D contributing the majority of AGPL1 transcripts. ABA induced repressions of TaSUT1, TaSuSy2, TaAGPL1 and TaAGPS1a imply that ABA negatively regulates sucrose import into the endosperm and its subsequent metabolism to ADP-glucose, the substrate for starch synthesis. The formations of amyloses and amylopectin from ADP-glucose appear to be mediated by specific members of GBSS, and SS, SBE and DBE gene families, and the ABA-induced transcriptional change in most of these genes implies that ABA regulates amylose and amylopectin synthesis. The findings provide insights into the molecular mechanisms underlying the coordination and ABA mediated regulation of sucrose transport into the developing endosperm and its subsequent metabolism to starch during grain filling in wheat., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

We have identified an abscisic acid (ABA) and stress-responsive GRAM (Glucosyltransferases, Rab-like GTPase activators and Myotubularins) domain protein GER5 (GEm-Related 5) closely related to GEM (GLABRA2 Expression Modulator), a novel regulator of cell division and cell fate determination in epidermal cells. A loss-of-function T-DNA line (ger5-2) and transgenic lines silencing (GER5(RNAi)) or overexpressing (GER5(OE)) GER5 displayed several defects in reproductive development affecting seed and embryo development. RNA in situ studies revealed GER5 and related GRAM genes (GEM and GEm-Related 1 (GER1)) have both overlapping and unique expression domains in male and female reproductive organs. Hormone immunolocalization experiments further indicate GER5 transcripts preferentially localize to reproductive tissues which accumulate ABA. Expression analysis revealed members of the GRAM family (GER5, GER1, GEM) display tissue-specific expression patterns and are responsive to phytohormones and abiotic stress, in addition to genetic lesions (aba1, aba2, ctr1) affecting ABA biosynthesis or ethylene signalling. Mature seeds of ger5-2 mutants also exhibit reduced sensitivity to ABA during seed germination assays. Microarray analysis of aborting and developing seeds isolated from ger5-2 mutants revealed underlying transcriptional changes in carbohydrate metabolism, hormone signalling and catabolic processes (e.g. protein degradation, autophagy). Taken together, our results indicate ABA-responsive GRAM genes play a novel role in regulating the reproductive development of plants, and raise intriguing questions regarding the functional relationship between members of the GRAM gene family., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Programmed cell death (PCD) is a universal process in all multicellular organisms. It is a critical component in a diverse number of processes ranging from growth and differentiation to response to stress. Somatic embryogenesis is one such process where PCD is significantly involved. Nitric oxide is increasingly being recognized as playing a significant role in regulating PCD in both mammalian and plant systems. Plant hemoglobins scavenge NO, and evidence is accumulating that events that modify NO levels in plants also affect hemoglobin expression. Here, we review the process of PCD, describing the involvement of NO and plant hemoglobins in the process. NO is an effector of cell death in both plants and vertebrates, triggering the cascade of events leading to targeted cell death that is a part of an organism's response to stress or to tissue differentiation and development. Expression of specific hemoglobins can alter this response in plants by scavenging the NO, thus, interrupting the death process. Somatic embryogenesis is used as a model system to demonstrate how cell-specific expression of different classes of hemoglobins can alter the embryogenic process, affecting hormone synthesis, cell metabolite levels and genes associated with PCD and embryogenic competence. We propose that plant hemoglobins influence somatic embryogenesis and PCD through cell-specific expression of a distinct plant hemoglobin. It is based on the premise that both embryogenic competence and PCD are strongly influenced by cellular NO levels. Increases in cellular NO levels result in elevated Zn(2+) and reactive-oxygen species associated with PCD, but they also result in decreased expression of MYC2, a transcription factor that is a negative effector of indoleacetic acid synthesis, a hormone that positively influences embryogenic competence. Cell-specific hemoglobin expression reduces NO levels as a result of NO scavenging, resulting in cell survival., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Arabidopsis shoot meristem activity is regulated by a molecular network involving the participation of several components, including SHOOTMERISTEMLESS (STM), CLAVATA1 (CLV1), and ZWILLE (ZLL). In an effort to identify the role of these genes during in vitro shoot formation Brassica and Arabidopsis plants were transformed with the Brassica napus (Bn) STM, CLV1, ZLL1 and ZLL2 identified in previous work [1]. In both systems shoot organogenesis was promoted by the over-expression of BnSTM, BnZLL1, and BnZLL2, and repressed by the over-expression of BnCLV1. This distinct regulation, analogous to that occurring during in vivo meristem formation where STM and ZLL encourage stem cell formation while CLV1 accelerates transition to differentiation, suggests similar regulatory mechanisms governing shoot formation in vivo and in vitro. While the BnZLL1 and BnZLL2 induction of shoot organogenesis correlated only to changes in auxin signaling, BnSTM and BnCLV1 evoked major transcriptional alterations in cytokinin response. Besides increasing the transcript levels of two cytokinin receptors, ARABIDOPSIS HISTIDINE KINASE4 (AHK4) and CYTOKININ INDEPENDENT KINASE (CKI1), ectopic expression of BnSTM induced Type-B ARABIDOPSIS RESPONSE REGULATORS (ARRs) and repressed Type-A ARRs. Opposite transcriptional patterns occurred in explants over-expressing BnCLV1, characterized by a decreased ability to produce shoots. The role played by Type-A and Type-B ARRs during shoot organogenesis was further examined using a genetic approach which revealed the requirement of ARR12 for the BnSTM positive regulation of shoot organogenesis. Collectively these results expand our knowledge on the function of meristem genes, and provide new tools for enhancing in vitro propagation systems., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

The phytohormone abscisic acid (ABA) plays an important role in developmental processes in addition to mediating plant adaptation to stress. In the current study, transcriptional response of 17 genes involved in ABA metabolism and transport has been examined in vegetative and reproductive organs exposed to cold and heat stress. Temperature stress activated numerous genes involved in ABA biosynthesis, catabolism and transport; however, several ABA biosynthesis genes (ABA1, ABA2, ABA4, AAO3, NCED3) were differentially expressed (up- or down-regulated) in an organ-specific manner. Key genes (CYP707As) involved in ABA catabolism responded differentially to temperature stress. Cold stress strongly activated ABA catabolism in all organs examined, whereas heat stress triggered more subtle activation and repression of select CYP707A genes. Genes involved in conjugation (UGT71B6), hydrolysis (AtBG1), and transport (ABCG25, ABCG40) of ABA or ABA glucose ester responded to temperature stress and displayed unique organ-specific expression patterns. Comparing the transcriptional response of vegetative and reproductive organs revealed ABA homeostasis is differentially regulated at the whole plant level. Taken together our findings indicate organs in close physical proximity undergo vastly different transcriptional programs in response to abiotic stress and developmental cues., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

The auxin transport inhibitor 2,3,5-triiodobenzoic acid (TIBA) is a useful compound for investigating the role of auxin flow during plant growth and development. In Arabidopsis lines, applications of TIBA during the induction phase of somatic embryogenesis inhibit embryo development and induce the differentiation of the meristematic cells of the shoot apical meristem (SAM), leading to the fusion of the cotyledons. These abnormalities were associated to changes in the expression levels of auxin transporter genes (PINs) and endogenous distribution of IAA. Treatments of TIBA caused a rapid accumulation of IAA within the epidermal and cortical root cells of the explants (bent-cotyledon zygotic embryos), as well as in the apical and sub-apical cells of the callus generated by the surface of the cotyledons of the explants. Within the callus only a few cells acquired meristematic characteristics, and this was associated to low expression levels of genes involved in embryogenic cell fate acquisition, such as WUSCHEL (WUS), LEAFY COTYLEDON 1 and 2. All these deleterious effects were attenuated when TIBA was administered to lines over-expressing SHOOT MERISTEMLESS (STM) isolated from Brassica oleracea (Bo), B. napus (Bn), and B. rapa (Br). Of interest, TIBA-treated explants of Arabidopsis lines over-expressing the Brassica STM were able to produce a large number of embryogenic cells and somatic embryos which exhibited a normal morphology and two distinct cotyledons. A proposed reason for this behaviour was ascribed to the ability of the transformed tissue to retain a normal distribution of auxin in the presence of TIBA. Proper localization of auxin might be required for the normal expression of several genes needed for the acquisition of embryogenic competence and formation of somatic embryos., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

Nucleotide metabolism was investigated during white spruce organogenesis by following the metabolic fate of (14)C-labeled adenine, adenosine and inosine, as purine precursors, and orotic acid, uridine, and uracil, as pyrimidine intermediates. Key enzymes of purine and pyrimidine metabolism were also assayed during the organogenic process. White spruce epicotyl explants cultured on shoot-forming (SF) medium had a better ability to utilize adenine and adenosine for nucleotide and nucleic acid synthesis, compared to tissue cultured on non-shoot forming (NSF) medium. High levels of salvage products were observed in SF tissue after 10 days in culture, when shoot formation was initiated along the epicotyl axis of the explants. Such a differential utilization of purine precursors was mainly due to the higher specific activity of the two adenine and adenosine salvage enzymes, adenine phosphoribosyltransferase (APRT) and adenosine kinase (AK), measured in SF tissue. Similar catabolism of inosine was observed in both SF and NSF conditions during the 30 days of culture. For pyrimidines, the higher activities of the de novo, salvage, and degradation pathways observed in SF tissue, compared to NSF tissue throughout the course of the experiment, clearly denote a faster turnover of pyrimidine nucleotides in the former. Taken together, these results suggest that a better utilization of purine bases and nucleosides for nucleotide and nucleic acid synthesis, as well as a more rapid turnover of pyrimidine nucleotides, represent a physiological switch, which occurs during the initiation and continuation of the organogenic process in white spruce., (Copyright © 2006 Elsevier Ireland Ltd. All rights reserved.)

Contribution of the adenine, adenosine and inosine salvage to the purine nucleotide and nucleic acid biosynthesis during white spruce (Picea glauca) somatic embryo maturation was estimated by in situ assays using [8-(14)C]adenine, [8-(14)C]adenosine and [8-(14)C]inosine. The salvage of adenine and adenosine was high during the initial stages of embryo maturation, characterized by rapid cell proliferation, but it declined upon further embryo development. Inosine salvage activity was always much lower than that observed for adenine and adenosine. Consistent with these results, activities of adenine phosphoribosyltransferase (APRT) and adenosine kinase (AK) measured in the embryo extracts in vitro were much higher than the activity of inosine kinase (IK) during all stages of embryo development. Utilization of adenosine and inosine for nucleotide and nucleic acid synthesis was found to be regulated by the enzymes AK and IK, as the pattern of their activities was very similar to the activity of adenosine and inosine salvage, estimated with exogenously supplied precursors. However, little correlation between salvage of adenine and activity of APRT was found throughout somatic embryo maturation. As no adenosine nucleosidase activity was found in white spruce embryos, adenosine, but not adenine, seems to be the major end product of adenylate catabolism and becomes the predominant substrate for purine salvage in vivo. Thus, adenosine salvage appeared to have the most important role in white spruce embryos. Studies on the metabolic fate of [8-(14)C]adenine and [8-(14)C]adenosine suggest that turnover of adenine nucleotides is rapid, as some of them are utilized for nucleic acid synthesis. In contrast, most of [8-(14)C]inosine taken up by the embryos seems to be directly catabolized by the conventional purine catabolic pathway via ureides in all stages of embryo maturation.