Phytoglobin expression in hypoxic root apical meristems alleviates programmed cell death by removing NO and moderating ethylene and ROS in the maize meristemati

Maintenance of a functional root is fundamental to plant survival in response to some abiotic stresses, such as water deficit. In this study, we found that overexpression of Arabidopsis class 1 phytoglobin (AtPgb1) alleviated the growth retardation of polyethylene glycol (PEG)-induced water stress by reducing programmed cell death (PCD) associated with protein folding in the endoplasmic reticulum (ER). This was in contrast to PEG-stressed roots down-regulating AtPgb1 that exhibited extensive PCD and reduced expression of several attenuators of ER stress, including BAX Inhibitor-1 (BI-1). The death program experienced by the suppression of AtPgb1 in stressed roots was mediated by reactive oxygen species (ROS) and ethylene. Suppression of ROS synthesis or ethylene perception reduced PCD and partially restored root growth. The PEG-induced cessation of root growth was preceded by structural changes in the root apical meristem (RAM), including the loss of cell and tissue specification, possibly as a result of alterations in PIN1- and PIN4-mediated auxin accumulation at the root pole. These events were attenuated by the overexpression of AtPgb1 and aggravated when AtPgb1 was suppressed. Specifically, suppression of AtPgb1 compromised the functionality of the WOX5-expressing quiescent cells (QCs), leading to the early and premature differentiation of the adjacent columella stem cells and to a rapid reduction in meristem size. The expression and localization of other root domain markers, such as SCARECROW (SCR), which demarks the endodermis and QCs, and WEREWOLF (WER), which specifies the lateral root cap, were also most affected in PEG-treated roots with suppressed AtPgb1. Collectively, the results demonstrate that AtPgb1 exercises a protective role in roots exposed to lethal levels of PEG, and suggest a novel function of this gene in ensuring meristem functionality through the retention of cell fate specification.

Root growth in maize (Zea mays L.) is regulated by the activity of the quiescent center (QC) stem cells located within the root apical meristem. Here, we show that despite being highly hypoxic under normal oxygen tension, QC stem cells are vulnerable to hypoxic stress, which causes their degradation with subsequent inhibition of root growth. Under low oxygen, QC stem cells became depleted of starch and soluble sugars and exhibited reliance on glycolytic fermentation with the impairment of the TCA cycle through the depressed activity of several enzymes, including pyruvate dehydrogenase (PDH). This finding suggests that carbohydrate delivery from the shoot might be insufficient to meet the metabolic demand of QC stem cells during stress. Some metabolic changes characteristic of the hypoxic response in mature root cells were not observed in the QC. Hypoxia-responsive genes, such as PYRUVATE DECARBOXYLASE (PDC) and ALCOHOL DEHYDROGENASE (ADH), were not activated in response to hypoxia, despite an increase in ADH activity. Increases in phosphoenolpyruvate (PEP) with little change in steady-state levels of succinate were also atypical responses to low oxygen tensions. Over-expression of PHYTOGLOBIN 1 (ZmPgb1.1) preserved the functionality of the QC stem cells during stress. The QC stem cell preservation was underpinned by extensive metabolic rewiring centered around activation of the TCA cycle and retention of carbohydrate storage products, denoting a more efficient energy production and diminished demand for carbohydrates under conditions where nutrient transport may be limiting. Overall, this study provides an overview of metabolic responses occurring in plant stem cells during oxygen deficiency.

Soybean (Glycine max) is an economically important crop which is very susceptible to salt stress. Tolerance to Na₂SO₄ stress was evaluated in soybean plants overexpressing or suppressing the phytoglobin GmPgb1. Salt stress depressed several gas exchange parameters, including the photosynthetic rate, caused leaf damage, and reduced the water content and dry weights. Lower expression of respiratory burst oxidase homologs (RBOHB and D), as well as enhanced antioxidant activity, resulting from GmPgb1 overexpression, limited ROS-induced damage in salt-stressed leaf tissue. The leaves also exhibited higher activities of the H₂O₂-quenching enzymes, catalase (CAT) and ascorbate peroxidase (APX), as well as enhanced levels of ascorbic acid. Relative to WT and GmPgb1-suppressing plants, overexpression of GmPgb1 attenuated the accumulation of foliar Na⁺ and exhibited a lower Na⁺/K⁺ ratio. These changes were attributed to the induction of the Na⁺ efflux transporter SALT OVERLY SENSITIVE 1 (SOS1) limiting Na⁺ intake and transport and the inward rectifying K⁺ channel POTASSIUM TRANSPORTER 1 (AKT1) required for the maintenance of the Na⁺/K⁺ balance.

Somatic embryogenesis in Arabidopsis encompasses an induction phase requiring auxin as the inductive signal to promote cellular dedifferentiation and formation of the embryogenic tissue, and a developmental phase favoring the maturation of the embryos. Strigolactones (SLs) have been categorized as a novel group of plant hormones based on their ability to affect physiological phenomena in plants. The study analyzed the effects of synthetic strigolactone GR24, applied during the induction phase, on auxin response and formation of somatic embryos. The expression level of two SL biosynthetic genes, MOREAXILLARY GROWTH 3 and 4 (MAX3 and MAX4), which are responsible for the conversion of carotene to carotenal, increased during the induction phase of embryogenesis. Arabidopsis mutant studies indicated that the somatic embryo number was inhibited in max3 and max4 mutants, and this effect was reversed by applications of GR24, a synthetic strigolactone, and exacerbated by TIS108, a SL biosynthetic inhibitor. The transcriptional studies revealed that the regulation of GR24 and TIS108 on somatic embryogenesis correlated with changes in expression of AUXIN RESPONSIVE FACTORs 5, 8, 10, and 16, known to be required for the production of the embryogenic tissue, as well as the expression of WUSCHEL (WUS) and Somatic Embryogenesis Receptor-like Kinase 1 (SERK1), which are markers of cell dedifferentiation and embryogenic tissue formation. Collectively, this work demonstrated the novel role of SL in enhancing the embryogenic process in Arabidopsis and its requirement for inducing the expression of genes related to auxin signaling and production of embryogenic tissue.

Summary: With the intensification of unpredictable flooding events because of global warming, there is a need to understand how root cells perceive and respond to oxygen deprivation. The use of high‐throughput single‐nuclei RNA‐sequencing (snRNA‐Seq) allows the examination of gene expression profiles in discrete cell types. Root tip segments of Arabidopsis seedlings subjected to normoxic (21% oxygen) or hypoxic (4% oxygen) treatments for 4 h were analyzed by snRNA‐Seq. Hypoxia induces a major transcriptome rewiring, most prominent in the meristematic root cells, with the exclusion of the quiescent center (QC), which is vulnerable to low oxygen. Changes in carbon and nitrogen metabolism by low oxygen were centered around increasing demand for carbohydrate to drive glycolytic fermentation, reduction of nitrate to sustain energetic processes, and the bypass of the TCA cycle via aspartate aminotransferase upregulation. The reduction of nitrate and nitrite in hypoxic cells contributes to amino acid metabolism and the utilization of NO in the phytoglobin–nitric oxide cycle to decrease the redox level and sustain energy production. In conclusion, the use of snRNA‐Seq provides a high‐resolution atlas of gene expression events defining root cell‐specific responses to low oxygen.