Chemical Engineering of Plants for Salt Tolerance

Salinity is one of the serious threats to global agriculture that threatens human food security. To tackle the problem, there were and still are quite several scientific efforts in place. Hence, in this dissertation, we used chemical and genetic engineering approaches to unravel the sophisticated knot of the problem using a Trojan peptoid called plant PeptoQ and overexpression of OsOPR7 gene in non-transformed WT tobacco BY-2 cells. First, plant PeptoQ that can be used to target a functional cargo (a rhodamine-labelled semiquinone peptoid as mimetic of coenzyme Q10) into mitochondria of tobacco BY-2 cells was characterized with regard to its cellular uptake and potential cytotoxicity. We found that the uptake is specific for mitochondria, rapid, dose-dependent, and requires both clathrin-mediated and clathrin-independent endocytosis, as well as actin filaments, while microtubules seem to be dispensable. Viability of the treated cells was not affected, and they showed better survival under salt stress, a condition that perturbs oxidative homeostasis in mitochondria. Using double labelling with appropriate fluorescent markers, we showed that targeting of this Trojan Peptoid to the mitochondria is not based on a passage through the plasma membrane (as thought hitherto), but on import via endocytotic vesicles and subsequent accumulation of the positively charged side chains at the negatively charged inner mitochondrial membrane. Second, the effects of pretreatment with plant PeptoQ and OsOPR7 overexpression, on salt stress induced detrimental effects in WT BY-2 cells were investigated. In general, both pretreatment with plant PeptoQ and overexpression of OsOPR7 in WT BY-2 cells, mitigated salt stress induced deleterious effects more or less in a similar manner. Cell expansion and cell viability were fully and partially compensated at moderate (75 mM NaCl) and high (150 mM NaCl) salt stress respectively by peptoid treatment and OsOPR7 overexpression. However, even if, the detrimental effects of salt stress on cell division (proliferation) were mitigated by both approaches, it was more sensitive as compared to cell expansion and viability. Furthermore, they significantly ameliorated doubling time, and effectively suppressed salt stress induced increase in MDA and superoxide levels in WT BY-2 cells. However, both approaches had no effect on hydrogen peroxide level. Plant PeptoQ pretreatment and OsOPR7 overexpression lead to increased SOD activity but decreased Mn-SOD transcript induction. However, they had no effect on catalase (CAT) activity. Except SOS1, NAC and OPR7 genes , other salt-related genes such as ion channels (NHX1 and SKOR), regulators for ion channels (SOS3 and SLT1) and jasmonate related gene (JAZ3) did not show strong transcript modulation in response to salinity, plant PeptoQ treatment and OsOPR7 overexpression. Similarly, even if, ionic balance was strongly perturbed by salt stress, both plant PeptoQ treatment and OsOPR7 overexpression had no mitigatory role at all. On the other hand, pretreatment of salt stressed WT and OsOPR7 overexpressor (OE) BY-2 cells with plant PeptoQ, caused increased OPDA level; however, it had no significant effect on JA-Ile level. It lead to a significant shift of the biosynthetic pathway from JA-Ile to OPDA, and this channeling of the pathway towards OPDA was significantly more accentuated for moderate salt stress (75 mM NaCl) but it faded as it proceeds to high salt stress (150 mM NaCl). Both plant PeptoQ pretreatment and OsOPR7 overexpression conferred salt tolerance to the non-transformed WT BY-2 cells by mitigating the salt stress induced detrimental effects effectively and efficiently

Effects of Salinity on Yield and Yield Components of tef [Eragrostis tef (Zucc.) Trotter] Accessions and Varieties

Abstract: This study aimed to screen fifteen low land tef genotypes (10 accessions and 5 varieties) with respect to yield and yield components at 0dS/m (control), 2, 4, 8 and 16 dS/m salinity levels and data analysis was carried out using SAS package (SAS version 8. On the other hand, accession/variety*treatment interaction effect was significant for MPDW (p<0.001) and SP/MP (p<0.01). This implies that all the accessions and varieties respond to salinity stress differently with respect to these two characters. However, the accession/variety*treatment interaction for the rest characters was insignificant reflecting that the entire varieties and accessions react to salinity stress similarly. Accession 236514, varieties DZ-Cr-358 and DZ-01-1681 were salt sensitive genotypes whereas accession 237186 and variety DZ-cr-37 were salt tolerant genotypes of all. Generally, the study revealed the presence of broad intraspecific genetic variation in tef accessions and varieties for salt tolerance but more in the former. Irrespective of salinity being a growing problem in Ethiopia in general and the Awash valley in particular, only little has been done on crops salt tolerance. Therefore, to alleviate the already existing and the inevitable incoming salinity problem, there should be similar and profound studies on tef and other crops

A Peptoid Delivers CoQ-derivative to Plant Mitochondria via Endocytosis

Controlled delivery of molecules interfering specifically with target activities in a cell of interest can be a powerful tool for experimental manipulation, because it can be administered at a defined time point and does not require genetic transformation, which in some systems is difficult and time consuming. Peptides as versatile tools that can be tailored for binding numerous binding partners, are of special interest. However, their passage through membranes, their intracellular targeting, and their sensitivity to proteases is limiting. The use of peptoids, where cationic amino-acid side chains are linked to nitrogen (rather than to carbon) of the peptide bond, can circumvent these limitations, because they are not cleavable by proteases. In the current work, we provide a proof-of-concept that such Trojan Peptoids, the plant PeptoQ, can be used to target a functional cargo (i.e. a rhodamine-labelled peptoid and a coenzyme Q10 derivative) into mitochondria of tobacco BY-2 cells as experimental model. We show that the uptake is specific for mitochondria, rapid, dose-dependent, and requires clathrin-mediated endocytosis, as well as actin filaments, while microtubules seem to be dispensable. Viability of the treated cells is not affected, and they show better survival under salt stress, a condition that perturbs oxidative homeostasis in mitochondria. In congruence with improved homeostasis, we observe that the salt induced accumulation of superoxide is mitigated and even inverted by pretreatment with PeptoQ. Using double labelling with appropriate fluorescent markers, we show that targeting of this Trojan Peptoid to the mitochondria is not based on a passage through the plasma membrane (as thought hitherto), but on import via endocytotic vesicles and subsequent accumulation in the mitochondrial intermembrane space, from where it can enter the matrix, e.g. when the permeability of the inner membrane is increased under salt stress

Response of Dry Matter Production of tef [Eragrostis tef (Zucc.) Trotter] Accessions and Varieties to NaCl Salinity

Abstract: This study aimed to screen fifteen low land tef genotypes (10 accessions and 5 varieties) with respect to dry matter production at 0 dS/m (control), 2, 4, 8 and 16 dS/m salinity levels. Data analysis was carried out using SAS package (SAS version 8.2, 2001) and SPSS version 12. The two ways ANOVA showed significant variation with respect to Root Dry Weight per plant (RDW) at p<0.001 and Total Dry Weight per plant (TDW) at p<0.05 for accessions/varieties. Moreover, it was significant for treatment (p<0.001) with respect to all the above three parameters. On the other hand, the two ways ANOVA for accession/variety*treatment interaction was significant for above Ground Dry Weight per plant (AGDW) and Total Dry Weight per plant (TDW) at p<0.01. This implies that all the accessions and varieties respond to salinity stress differently with respect to these three dry matter production characters. However, the two ways ANOVA for the accession/variety*treatment interaction for Root Dry Weight per plant (RDW) was insignificant reflecting that the entire varieties and accessions react to salinity stress similarly. Accessions 212611, 55017, 231217 and varieties DZ-Cr-358 and DZ-01-1281 were salt sensitive genotypes whereas accessions 237131, 237186, 212928 and variety DZ-Cr-37 were salt tolerant genotypes of all. Varieties were more salt affected than accessions with respect to dry matter production

Investigation of the Reasons for the Unique Growth and Development of Agave Species (Agave sisalana and Agave americana) Crop Plants at the Southern, Central, North Western and Eastern Parts of Tigray, Ethiopia

Abstract: Composite topsoil samples from 16 study sites were taken and then their physical as well as chemical attributes were determined at the National Soil Testing Center (NSTC). Moreover, plant attributes such as Leaf Length (LL), Leaf Biomass (LB), Fiber Length (FL) and Tensile Strength (TS) were measured. Data analysis was carried out using JMP5 (version 5.0) and SPSS (version 12.0) statistical softwares. The one way analysis of variance (ANOVA) for Leaf Length (LL), Leaf Biomass (LB), and Tensile Strength (TS) showed statistical significance with respect to the study sites (p<0.0001). Furthermore, one way ANOVA for tensile strength (TS), Leaf Biomass (LB) and Leaf Length (LL) with respect to blocks found statistically significant at p<0.0001, p<0.001 and p<0.05 respectively. Nevertheless, one way ANOVA for Fiber Length (FL) appeared statistically insignificant (p>0.05) with respect of blocks. On the other hand, the one way ANOVA for both soil chemical and physical attributes with respect to blocks found statistically insignificant (p>0.05). The rainfall, sunshine, and temperature situations as well as the soil chemical and physical attributes being inline with the requirements of Agave species could be reasons for the unique growth and development of A. americana and A. sisalana crop plants in the study area