scholarly journals Exogenous abscisic acid impacts the development of isolated immature endosperm in bread wheat

2021 ◽  
Author(s):  
Iwona Chłosta ◽  
Małgorzata Kozieradzka-Kiszkurno ◽  
Dagmara Kwolek ◽  
Izabela Marcińska ◽  
Apolonia Sieprawska ◽  
...  
Keyword(s):  
Abscisic Acid ◽  
Bread Wheat ◽  
Positive Impact ◽  
Wheat Variety ◽  
Culture Conditions ◽  
Soluble Carbohydrates ◽  
In Planta ◽  
Specific Storage ◽  
The Media

AbstractEndosperm in cereals such as wheat, is a part of the mature seeds and a valuable source of key substances for humans and animals. For this reason, the development of immature endosperm tissues in planta was the focus of this research. However, it is commonly known that tissue culture conditions can alter the developmental pathway of plant cells and can expose their potency. There is scarce information about research on isolated endosperm in wheat. The development of isolated immature endosperm in the winter bread wheat variety ‘Kobra’, depending on the media composition, is presented in this study. Abscisic acid (ABA) is a key plant growth regulator for proper seed development. The addition of exogenous ABA had a positive impact on the size and ultrastructural features in isolated endosperm, especially of the outer aleurone-like cells. Furthermore, the content of starch in the endosperm cultured on a medium with ABA did not significantly differ from that of caryopsis at the same age, in contrast to soluble carbohydrates. Fluorescein diacetate (FDA) staining and confocal microscopy observation confirmed the viability of the cells from the outer layers. The analysis of internucleosomal fragmentation of DNA in the explants suggests the induction of programmed cell death (PCD) and DNA degradation typical of necrosis. We concluded that the development of isolated immature endosperm in bread wheat depends on the composition of the media. Thus, it could be a model for in vitro studies of this specific storage tissue and its response to culture conditions in bread wheat.

scholarly journals Expression of the ToxA and PtrPf2 genes of the phytopathogenic fungus Pyrenophora tritici-repentis at the beginning of the infection process

2019 ◽  
Author(s):  
Nina V. Mironenko ◽  
Alexandra S. Orina ◽  
Nadezhda M. Kovalenko
Keyword(s):  
Transcription Factor ◽  
Wheat Variety ◽  
Infection Process ◽  
Phytopathogenic Fungus ◽  
In Planta ◽  
Effector Gene ◽  
Pyrenophora Tritici Repentis ◽  
Wheat Leaves ◽  
Necrotrophic Effector

This study shows that the necrotrophic effector gene ToxA is differentially expressed in isolates of P. tritici-repentis fungus at different time periods after inoculation of the wheat variety Glenlea which has the gene Tsn1 controlling sensitivity to the necrosis inducing toxin Ptr ToxA. Two P. tritici-repentis isolates with different ability to cause necrosis on the leaves of Glenlea variety (nec + and nec-) and with different expression level of ToxA and gene of factor transcription PtrPf2 in vitro were used for analysis. Isolates of P. tritici-repentis are characterized by the differential expression of ToxA in planta. The expression of the ToxA gene in P. tritici-repentis ToxA+ isolates significantly increased when infected the wheat leaves compared to ToxA expression results obtained in vitro. The levels of ToxA expression in both isolates differed significantly after 24, 48 and 96 hours after inoculation, however, the dynamics of the trait change over time were similar. However, the highest ToxA expression in the virulent (nec+) isolate in contrast with the avirulent (nec-) isolate was observed at a point of 48 hours. Whereas the expression of regulating transcription factor PtrPf2 in planta differed imperceptibly from expression in vitro throughout the observation period. Obviously, the role of the fungal transcription factor in regulating the effector gene expression weakens in planta, and other mechanisms regulating the expression of pathogen genes at the biotrophic stage of the disease develop.

Isozyme assessment of the genetic stability of micropropagated hybrid larch (Larix × eurolepis Henry)

1995 ◽  
Vol 25 (7) ◽  
pp. 1103-1112 ◽  
Author(s):  
Sylvie Richard ◽  
Sylvie Gauthier ◽  
Sylvie Laliberté
Keyword(s):  
Tissue Culture ◽  
Growth Regulators ◽  
Genetic Stability ◽  
Physiological Response ◽  
Culture Conditions ◽  
Hybrid Larch ◽  
Short Shoot ◽  
In Vitro Shoots ◽  
The Media

The search for the occurrence of somaclonal variation of in vitro shoots and acclimatized plants of a hybrid larch (Larix × urolepis Henry) clone was performed by the analysis of eight isozyme systems. Cultures were established from short shoot buds of mature material. The effects of growth regulators in the media, subculture intervals, and periods in culture were analyzed for in vitro shoots. Variability was found in in vitro shoots but appeared to be related to a physiological response to culture conditions. Once acclimatized, most tissuecultured plants expressed the same enzymatic patterns as those of control plants (stecklings and the ortet). The variations observed for some acclimatized plants were also observed in control plants and were not related to ontogenic stage. Results from the isoenzymatic systems studied showed that hybrid larch plants regenerated from tissue culture were not significantly different from stecklings and the ortet.

scholarly journals A Nuclear and Cytoplasmic Characterization of Bovine Oocytes Reveals That Cysteamine Partially Rescues the Embryo Development in a Model of Low Ovarian Reserve

Animals ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1936
Author(s):  
Valentina Lodde ◽  
Alberto Maria Luciano ◽  
Giulia Musmeci ◽  
Ileana Miclea ◽  
Irene Tessaro ◽  
...  
Keyword(s):  
Embryo Development ◽  
Positive Impact ◽  
Antral Follicle ◽  
Oocyte Quality ◽  
Antral Follicle Count ◽  
Culture Conditions ◽  
Free Radical Scavenger ◽  
Radical Scavenger ◽  
Ovarian Aging

Decreased oocyte quality is a major determinant of age-associated fertility decline. Similarly, individuals affected by early ovarian aging carry low-quality oocytes. Using an established bovine model of early ovarian aging, we investigated key features of ‘quality’ oocyte maturation, associated with the onset of egg aneuploidy and reproductive aging, such as histone modifications, mitochondria distribution and activity, reduced glutathione (GSH) content, and gap junction functionality. Bovine ovaries were classified according to the antral follicle count (AFC), and the retrieved oocytes were processed immediately or matured in vitro. We observed alterations in several cellular processes, suggesting a multifactorial etiology of the reduced oocyte quality. Furthermore, we performed a rescue experiment for one of the parameters considered. By adding cysteamine to the maturation medium, we experimentally increased the free radical scavenger ability of the ‘low competence’ oocytes and obtained a higher embryo development. Our findings show that adopting culture conditions that counteract the free radicals has a positive impact on the quality of ‘compromised’ oocytes. Specifically, cysteamine treatment seems to be a promising option for treating aging-related deficiencies in embryo development.

scholarly journals Fine-tuning of ABA responses by the protein kinase WNK8

2018 ◽  
Author(s):  
Rainer Waadt ◽  
Kenji Hashimoto ◽  
Esther Jawurek ◽  
Melanie Krebs ◽  
Martin Scholz ◽  
...  
Keyword(s):  
Abscisic Acid ◽  
Protein Kinase ◽  
Signaling Pathway ◽  
Direct Interaction ◽  
Negative Regulator ◽  
Fine Tuning ◽  
Dependent Manner ◽  
In Planta ◽  
Young Seedling

SUMMARYThe phytohormone abscisic acid (ABA) regulates various growth- and developmental processes in response to limiting water conditions. ABA functions through an established signaling pathway consisting of PYR1/PYL/RCAR ABA receptors that inhibit group A type 2C protein phosphatases (PP2Cs) in an ABA-dependent manner. Inhibition of PP2Cs enables the activation of SnRK2-type protein kinases that phosphorylate downstream targets including transcription factors and ion channels. However, ABA-dependent signals have to be integrated into other growth- and developmental programs to ensure a successful life cycle. Here, we have characterized the role of the protein kinase WNK8 in the ABA signalling pathway. Two T-DNA insertion alleles wnk8-1 and wnk8-4 exhibited contrasting ABA responses during seed germination and young seedling growth. However, reciprocal crossings with wild type plants suggested that wnk8-1 that still expressed the WNK8 kinase domain functioned in a hypermorphic and dominant manner. WNK8 directly interacted with the PP2C PP2CA in planta and was negatively regulated by this phosphatase in vitro. WNK8 also phosphorylated the ABA receptor PYR1 in vitro. Double mutant analyses revealed that the dominant allele wnk8-1 suppressed the ABA- and glucose hypersensitivity of the pp2ca-1 T-DNA allele. In transient protoplast assays WNK8 suppressed ABA-induced reporter gene expression that was dependent on a functional kinase. In summary, we have identified the protein kinase WNK8 as a negative regulator of ABA responses during young seedling establishment through its direct interaction with core ABA signaling components.SIGNIFICANCE STATEMENTThe phytohormone abscisic acid regulates the water household of plants through a defined core signaling pathway. Here we have identified the protein kinase WNK8 as a direct interactor of core abscisic acid signalling components and as a negative modulator of abscisic acid responses during young seedling development in Arabidopsis.

scholarly journals 229 EFFECT OF CULTURE CONDITIONS ON AQUAPORIN mRNA ABUNDANCE IN MOUSE BLASTOCYSTS

2005 ◽  
Vol 17 (2) ◽  
pp. 265
Author(s):  
H. Offenberg ◽  
P.D. Thomsen
Keyword(s):  
Blastocyst Stage ◽  
Culture Conditions ◽  
Mrna Abundance ◽  
Control Group ◽  
Mrna Levels ◽  
Blastocyst Formation ◽  
Gapdh Mrna ◽  
The Media

It is known that culture conditions can alter gene expression of the pre-implantation embryo. We have previously shown that aquaporins (AQPs) are expressed in the mouse embryo and that they are involved in the passage of water across the trophoblast cells during blastocyst formation. This study was conducted to investigate whether AQP mRNA abundance is altered by culturing embryos in vitro compared to in vivo developed embryos. Furthermore we wanted to investigate if AQP mRNA abundance was influenced by the osmolality of the media. It is possible to compare the effect of hyperosmolality that the embryo may be able to compensate for by adding glycerol which can cross some AQPs, compared to the addition of sucrose which can not cross the membranes. Mouse embryos were obtained by superovulating B6D2F1 mice followed by culture of the flushed presumptive zygotes in KSOM to the blastocyst stage (in vitro) or by flushing blastocysts from the uterus (in vivo). For the study of the influence of osmolality on AQP mRNA abundance, zygotes were flushed and cultured to the compacted 8-cell stage and then placed in media of increasing osmolality, using either glycerol or sucrose. The osmolalities of the media were 243 (control), 300, 350, and 400 mOsm. Embryos were cultured to the blastocyst stage and frozen in liquid nitrogen. Embryonic RNA was extracted using a Dynabeads mRNA Capture kit (Dynal, Oslo, Norway). Real time PCR was performed on embryonic cDNA on a Lightcycler (Roche Diagnostics, 2650 Hvidovre, Denmark) using aquaporin-specific primers and primers for β-actin and GAPDH. The results of the quantitative RT-PCR analysis showed that in vitro-cultured embryos had a lower mRNA abundance for AQP 8, 9, and 11 compared to the in vivo-developed embryos but that the AQP 3 mRNA abundance was unaltered. Analysis of the housekeeping genes showed that GAPDH mRNA levels were unchanged in vitro, whereas β-actin was up-regulated in vitro. The osmotically challenged embryos showed the following blastocyst rates compared to the controls: glycerol 300: 100%; glycerol 350: 100%; glycerol 400: 100%; sucrose 300: 100%; sucrose 350: 78%; and sucrose 400: 0%. Thus, glycerol up to 400 mOsm had no effect on blastocyst rates, whereas addition of sucrose reduced blastocyst formation, with a total inhibition at 400 mOsm. Analysis of the mRNA abundance showed a reduction of AQP 8 in the glycerol solutions. The level was reduced to 30% of the control group at 300 mOsm, to 27% at 350 mOsm and to 8% at 400 mOsm. There was no corresponding reduction of AQP 8 mRNA abundance in sucrose solutions. Further, AQP 3, 7, 9, and 11 mRNA levels as well as β-actin and GAPDH mRNA levels were unaltered in the osmotically challenged embryos. In conclusion, this study shows that embryonic culture affects the abundance of several AQPs and that compensation of a glycerol-induced osmotical challenge induces down-regulation of AQP 8 expression. Embryos tolerate high glycerol concentrations better than high sucrose concentrations but the possible role of AQP 8 in this process is unclear at present.

scholarly journals The effect of blue and red light on physiological and biochemical characteristics of wheat plants

2020 ◽  
Vol 27 ◽  
pp. 19-22
Author(s):  
I. V. Azizov ◽  
F. I. Gasymova ◽  
U. F. Ibragimova ◽  
K. R. Tagiyeva ◽  
A. B. Abdullayeva
Keyword(s):  
Hydrogen Peroxide ◽  
Sodium Chloride ◽  
Durum Wheat ◽  
Bread Wheat ◽  
Blue Light ◽  
Ascorbate Peroxidase ◽  
Red Light ◽  
Wheat Variety ◽  
Soluble Carbohydrates ◽  
Physiological And Biochemical

Aim. The effect of blue and red light on the activity of catalase and ascorbate peroxidase, the content of proteins and soluble carbohydrates in the leaves of wheat varieties under the action of sodium chloride were investigated. Methods. The objects of research were bread wheat Saratovskaya-29 (Triticum aestivum L.) and durum wheat Barakatli-95 (Triticum durum Desf.). Plants were grown in laboratory conditions in an aqueous medium using a Knoop nutrient medium. Experimental plants were coated with transparent films transmitting light at wavelengths of 420-480 nm (blue light) and 640-680 nm (red light). During the growing season, samples of fully formed leaves were taken for physiological and biochemical studies every week at 11 a.m. Results. Under the action of sodium chloride, the accumulation of hydrogen peroxide in white light was higher than in blue and red light (table). Low catalase activity was also observed in blue and red light under the influence of sodium chloride in both varieties. In bread wheat variety Saratovskaya -29 level of hydrogen peroxide, the activity of catalase and ascorbate peroxidase enzymes was lower than that of the Barakatli-95 durum wheat variety. Conclusions. Blue light stimulated the synthesis of proteins, while red light stimulated the synthesis of carbohydrates. Blue and red light prevented the formation of H2O2 under the influence of NaCl. Keywords: blue light, red light, catalase, ascorbate peroxidase, proteins, carbohydrates.

scholarly journals Improving In Vitro Culture of Human Male Fetal Germ Cells

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 2033
Author(s):  
Myriam Martin-Inaraja ◽  
Monica Ferreira ◽  
Jasin Taelman ◽  
Cristina Eguizabal ◽  
Susana M. Chuva De Sousa Lopes
Keyword(s):  
Germ Cells ◽  
Culture Medium ◽  
Culture Media ◽  
Culture Conditions ◽  
Pronounced Difference ◽  
Coated Substrate ◽  
Male Gametes ◽  
The Media

Male human fetal germ cells (hFGCs) give rise to spermatogonial stem cells (SSCs), which are the adult precursors of the male gametes. Human SSCs are a promising (autologous) source of cells for male fertility preservation; however, in contrast to mouse SSCs, we are still unable to culture them in the long term. Here, we investigated the effect of two different culture media and four substrates (laminin, gelatin, vitronectin and matrigel) in the culture of dissociated second trimester testes, enriched for hFGCs. After 6 days in culture, we quantified the presence of POU5F1 and DDX4 expressing hFGCs. We observed a pronounced difference in hFGC number in different substrates. The combination of gelatin-coated substrate and medium containing GDNF, LIF, FGF2 and EGF resulted in the highest percentage of hFGCs (10% of the total gonadal cells) after 6 days of culture. However, the vitronectin-coated substrate resulted in a comparable percentage of hFGCs regardless of the media used (3.3% of total cells in Zhou-medium and 4.8% of total cells in Shinohara-medium). We provide evidence that not only the choices of culture medium but also choices of the adequate substrate are crucial for optimizing culture protocols for male hFGCs. Optimizing culture conditions in order to improve the expansion of hFGCs will benefit the development of gametogenesis assays in vitro.

scholarly journals Global Transcriptome Profiling of Xanthomonas oryzae pv. oryzae under in planta Growth and in vitro Culture Conditions

2017 ◽  
Vol 33 (5) ◽  
pp. 458-466 ◽  
Author(s):  
So Eui Lee ◽  
Ravi Gupta ◽  
Ramesha H. Jayaramaiah ◽  
Seo Hyun Lee ◽  
Yiming Wang ◽  
...  
Keyword(s):  
In Vitro Culture ◽  
Transcriptome Profiling ◽  
Culture Conditions ◽  
Xanthomonas Oryzae ◽  
In Planta ◽  
Global Transcriptome

scholarly journals Identification and characterization of interactions between abscisic acid and mitochondrial adenine nucleotide translocators

2011 ◽  
Vol 437 (1) ◽  
pp. 117-123 ◽  
Author(s):  
Olesya A. Kharenko ◽  
Jason Boyd ◽  
Ken M. Nelson ◽  
Suzanne R. Abrams ◽  
Michele C. Loewen
Keyword(s):  
Abscisic Acid ◽  
Stress Responses ◽  
Adenine Nucleotide ◽  
In Planta ◽  
Chemical Proteomics ◽  
Physiological Relevance ◽  
Functional Localization ◽  
Proteomics Approach

ABA (abscisic acid) is a plant hormone involved in important processes including development and stress responses. Recent reports have identified a number of plant ABA receptors and transporters, highlighting novel mechanisms of ABA action. In the present paper we describe application of a chemical proteomics approach leading to the identification of mitochondrial ANTs (adenine nucleotide translocators) as ABA-interacting proteins. Initial in vitro studies confirmed inhibition of ANT-dependent ATP translocation by ABA. Further analysis demonstrated ANT-dependent uptake of ABA into both recombinant Arabidopsis thaliana ANT2-containing proteoliposomes and native isolated spinach mitochondria; the latter with a Km of 3.5 μM and a Vmax of 2.5 nmol/min per g of protein. ATP was found to inhibit ANT-dependent ABA translocation. Specificity profiles highlight the possibility of mechanistic differences in translocation of ABA and ATP. Finally, ABA was shown to stimulate ATPase activity in spinach mitochondrial extracts. ABA concentrations in plant cells are estimated to reach the low micromolar range during stress responses, supporting potential physiological relevance of these in vitro findings. Overall, the present in vitro work suggests the possibility of as yet uncharacterized mechanisms of ABA action in planta related to inhibition of mitochondrial ATP translocation and functional localization of ABA in the mitochondrial matrix.

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