Oxidative and osmotolerant effects in Salvator merianae (Squamata: Teiidae) red blood cells during hibernation

Hibernation is a natural condition of animals that lives in the temperate zone, although some tropical lizards also experience hibernation annually, such as the lizard native from South America, Salvator merianae, or “tegu” lizard. Even though physiological and metabolic characteristic associated with hibernation have been extensively studied, possible alterations in the red blood cells (RBC) integrity during this period remains unclear. Dehydration and fasting are natural consequences of hibernating for several months and it could be related to some cellular modifications. In this study, we investigated if the osmotic tolerance of RBCs of tegu lizard under hibernation is different from the cells obtained from animals while normal activity. Additionally, we indirectly investigated if the RBCs membrane of hibernating tegus could be associated with oxidation by quantifying oxidized biomolecules and the activity of antioxidant enzymes. Our findings suggest that RBCs are more fragile during the hibernation period, although we did not find evidence of an oxidative stress scenario associated with the accentuated fragility. Even though we did not exclude the possibility of oxidative damage during hibernation, we suggested that an increased RBCs volume as a consequence of hypoosmotic blood during hibernation could also affect RBCs integrity as noted.

The Functional Association of ACQOS/VICTR with Salt Stress Resistance in Arabidopsis thaliana Was Confirmed by CRISPR-Mediated Mutagenesis

Clustered regularly interspaced palindromic repeat (CRISPR)-mediated mutagenesis has become an important tool in plant research, enabling the characterization of genes via gene knock-out. CRISPR genome editing tools can be applied to generate multi-gene knockout lines. Typically, multiple single-stranded, single guide RNAs (gRNAs) must be expressed in an organism to target multiple genes simultaneously; however, a single gRNA can target multiple genes if the target genes share similar sequences. A gene cluster comprising ACQUIRED OSMOTOLERANCE (ACQOS; AT5G46520) and neighboring nucleotide-binding leucine-rich repeats (NLRs; AT5G46510) is associated with osmotic tolerance. To investigate the role of ACQOS and the tandemly arranged NLR in osmotic tolerance, we introduced small insertion/deletion mutations into two target genes using a single gRNA and obtained transformant plant lines with three different combinations of mutant alleles. We then tested our mutant lines for osmotic tolerance after a salt-stress acclimation period by determining the chlorophyll contents of the mutant seedlings. Our results strongly suggest that ACQOS is directly associated with salt resistance, while the neighboring NLR is not. Here, we confirmed previous findings suggesting the involvement of ACQOS in salt tolerance and demonstrated the usefulness of CRISPR-mediated mutagenesis in validating the functions of genes in a single genetic background.

Differential Accumulation of Metabolites in Suaeda Species Provides New Insights into Abiotic Stress Tolerance in C4-Halophytic Species in Elevated CO2 Conditions

Halophytic plants can adapt to grow and thrive in highly saline conditions. Suaeda species are annual halophytes with high salt tolerance and are most suitable in the restoration of salinized or contaminated saline land and as food, forage, medicine, and bioenergy. In this study, we comprehensively analyzed the different metabolic responses of Suaeda species under salt and drought stress at ambient and elevated CO2 conditions. Seedlings of Suaeda species were treated with 500 mM NaCl and 5% of polyethylene glycol under elevated CO2 stress conditions for 24 h. Then, widely untargeted metabolites were detected by gas chromatography–mass spectrometry. Different metabolites involved in amino acid metabolism, glycolysis, photorespiration, and tricarboxylic acid cycle were quantitatively determined after stress treatments. A total of 61 primary metabolites were annotated. Different treatments increased the contents of certain metabolites, such as amino acids, sugars, and organic acids, as well as some antioxidants, such as quininic acid, kaempferol, and melatonin. These substances may be correlated with osmotic tolerance, increased antioxidant activity, and medical and nutritional value in the species. This study suggests that various metabolites differentially accumulated in C4Suaeda species under varying stress conditions. Furthermore, this work provides new insights into the key secondary metabolite pathway involved in stress tolerance.

MsCIPK, a CBL-interacting Protein Kinase in Medicago sativa L., Confers Salt and Osmotic Stress Tolerance in Transgenic Tobacco

Alfalfa is an important perennial forage but suffers from salt and osmotic stresses worldwide. Calcineurin B-like proteins (CBLs) and CBL-interacting protein kinases (CIPKs) are reported to play important roles in response to diverse plant stresses, but are largely unvalidated in alfalfa. In this study, we cloned a MsCIPK gene, which contained 1530 bp, coding 509 amino acids, with typical CIPK functional domains. The expression pattern of MsCIPK was measured using qRT-PCR under salt, drought, heat, cold and ABA stresses. Under NaCl, heat and ABA treatment, the expression pattern of MsCIPK was generally similar, with a first steady decrease and then a gradual increase pattern. The highest expression of MsCIPK was all observed at the start point of all treatments, except in cold treatment. Using transgenic tobaccos of MsCIPK, we further measured the content of malondialdehyde (MDA), superoxide dismutase (SOD), soluble protein (SOP), and proline (Pro) under 21 days’ salt and 24 hours’ cold treatment. Under both salt and cold conditions, the content of MDA, SOP and Pro had a similar overall increase pattern with the time of treatment. These results indicated that the MsCIPK played an important role in improving alfalfa’s salt and osmotic tolerance.