Manifestation of agronomically valuable traits in the progeny of a sorghum mutant carrying the genetic construct for RNA silencing of the γ-kafirin gene.

Improving the nutritional value of grain sorghum, a drought- and heat-tolerant grain crop, is an important task in the context of global warming. One of the reasons for the low nutritional value of sorghum grain is the resistance of its storage proteins (kafirins) to proteolytic digestion, which is due, among other things, to the structural organization of protein bodies, in which γ-kafirin, the most resistant to proteases, is located on the periphery, encapsulating more easily digested α-kafirins. The introduction of genetic constructs capable of inducing RNA silencing of the γ-kafirin (gKAF1) gene opens up prospects for solving this problem. Using Agrobacterium-mediated genetic transformation of immature embryos of the grain sorghum cv. Avans we have obtained a mutant with improved digestibility of endosperm proteins (up to 92 %) carrying a genetic construct for RNA silencing of the gKAF1 gene. The goal of this work was to study the stability of inheritance of the introduced genetic construct in T2-T4 generations, to identify the number of its copies, as well as to trace the manifestation of agronomically valuable traits in the offspring of the mutant. The mutant lines were grown in experimental plots in three randomized blocks. The studied lines were characterized by improved digestibility of kafirins, a modified type of endosperm, completely or partially devoid of the vitreous layer, an increased percentage of lysine (by 75 %), reduced plant height, peduncle length, 1000-grains weight, and grain yield from the panicle. In T2, a line with monogenic control of GA resistance was selected. qPCR analysis showed that in different T3 and T4 plants, the genetic construct was present in 2-4 copies. In T3, a line with a high digestibility of endosperm proteins (81 %) and a minimal decrease in agronomically valuable traits (by 5-7 %) was selected.

Fringe is a glycosyltransferase that modifies Notch.

Notch receptors function in highly conserved intercellular signalling pathways that direct cell-fate decisions, proliferation and apoptosis in metazoans. Fringe proteins can positively and negatively modulate the ability of Notch ligands to activate the Notch receptor. Here we establish the biochemical mechanism of Fringe action. Drosophila and mammalian Fringe proteins possess a fucose-specific beta1,3 N-acetylglucosaminyltransferase activity that initiates elongation of O-linked fucose residues attached to epidermal growth factor-like sequence repeats of Notch. We obtained biological evidence that Fringe-dependent elongation of O-linked fucose on Notch modulates Notch signalling by using co-culture assays in mammalian cells and by expression of an enzymatically inactive Fringe mutant in Drosophila. The post-translational modification of Notch by Fringe represents a striking example of modulation of a signalling event by differential receptor glycosylation and identifies a mechanism that is likely to be relevant to other signalling pathways.

Dorsal-ventral signaling in the Drosophila eye.

The development of the Drosophila eye has served as a model system for investigations of tissue patterning and cell-cell communication; however, early eye development has not been well understood. The results presented here indicate that specialized cells are established along the dorsal-ventral midline of the developing eye by Notch-mediated signaling between dorsal and ventral cells, and that Notch activation at the midline plays an essential role both in promoting the growth of the eye primordia and in regulating eye patterning. These observations imply that the developmental homology between Drosophila wings and vertebrate limbs extends to Drosophila eyes.

Modulators of Notch signaling.

In addition to the core components of the Notch pathway, a number of proteins have been identified that exert positive or negative influences on Notch signaling. These include extracellular modulators, which may influence binding or activation of Notch by its ligands, cytoplasmic modulators, which presumably influence signal transduction steps after receptor activation, and nuclear modulators, which may influence the transcriptional activity of a Notch-CSL protein complex. Many of the cytoplasmic and nuclear modulators appear to bind directly to discrete domains within the intracellular domain of Notch. Genetic studies indicate that distinct modulators are deployed during distinct modes of Notch signaling.

[Interconnection of hierarchical compactization levels as the basis for functional organization of chromatin]. / Vzaimosviaz' ierarkhichiskikh urovnei kompaktizatsii kak osnova funktsional'noi organizatsii khromatina.

Recent data on chromatin structural organization are presented. In the light of present concept of chromatin structure the problem of hierarchical levels of DNA packaging within chromatin is discussed. Particular emphasis has been placed on the correlation between the changes occurring at different DNA compactization levels within topological domains and chromatin functional activity.

Fringe modulates Notch-ligand interactions.

The Notch family of transmembrane receptor proteins mediate developmental cell-fate decisions, and mutations in mammalian Notch genes have been implicated in leukaemia, breast cancer, stroke and dementia. During wing development in Drosophila, the Notch receptor is activated along the border between dorsal and ventral cells, leading to the specification of specialized cells that express Wingless (Wg) and organize wing growth and patterning. Three genes, fringe (fng), Serrate (Ser) and Delta (Dl), are involved in the cellular interactions leading to Notch activation. Ser and Dl encode transmembrane ligands for Notch, whereas fng encodes a pioneer protein. We have investigated the relationship between these genes by a combination of expression and coexpression studies in the Drosophila wing. We found that Ser and Dl maintain each other's expression by a positive feedback loop. fng is expressed specifically by dorsal cells and functions to position and restrict this feedback loop to the developing dorsal-ventral boundary. This is achieved by fng through a cell-autonomous mechanism that inhibits a cell's ability to respond to Serrate protein and potentiates its ability to respond to Delta protein.

[Effect of specific nucleotide sequences on the transcription activity of long terminal repeats of avian retroviruses]. / Vliianie spetsificheskikh nukleotidnykh posledovatel'nostei na transkriptsionnuiu aktivnost' dlinnykh kontsevykh povtorov retrovirusov ptits.

Influence of specific nucleotide sequences on the constitutive transcriptional activity of the Rous sarcoma long terminal repeats (LTR) was studied. The possible function of different elements of the transcriptional control region of the avian retroviral LTR was discussed on the basis of a comparative analysis of the effect of specific nucleotide insertions on LTR activity in vivo and in vitro.

A relatively small 5' regulatory region of esterase S gene of Drosophila virilis determines the specific expression as revealed in transgenic experiments.

Expression of the esterase S gene of Drosophila virilis was studied in transgenic experiments. Truncated genomic copy of this gene including 400 bp of 5' regulatory region was integrated into the genome of Drosophila melanogaster. The products of the transferred gene were detected. It was found that strict temporal and tissue specificity of the esterase S gene expression is conserved in transformed flies. The results suggest that this specificity is evidently determined by the regulatory region of the esterase S gene and controlled by cis mechanism.

The expression of esterase S gene of Drosophila virilis in Drosophila melanogaster.

Drosophila melanogaster was transformed with the esterase S gene from Drosophila virilis. This gene is strongly activated in ejaculatory bulbs of mature males of Drosophila virilis. The closely related gene from Drosophila melanogaster is activated in ejaculatory ducts. The tissue- and stage-specific expression of incomplete genomic copy of the esterase S gene integrated into the Drosophila melanogaster genome is the same as in Drosophila virilis. These data show that tissue and stage specificity is determined by relatively small 5' regulatory region of the esterase S gene. The comparison between deduced amino-acid sequences of the esterase S of Drosophila virilis and esterase 6 of Drosophila melanogaster was performed. These sequences revealed 50% homology.

[Acetylation of histones in vitro causes chromatin decompactization]. / Atsetilirovanie gistonov in vitro vyzyvaet dekompaktizatsiiu khromatina.

It has been discovered that chemical acetylation of chromatin in vitro by acetyladenylate leads to decrease in the mobility of nucleosomal fiber fragments in agarose gel. Acetylation of HI-depleted chromatin does not produce this effect. Based on these results together with other available data, it is possible to conclude that acetylation causes decompactization of the chromatin fiber. A possible molecular mechanism of in vivo fiber decondensation and interaction of different chromatin compactization levels is discussed.

[Dependence of the chromatin condensation process on the temperature and ionic strength of the media: investigation using gel electrophoresis in low density agarose]. / Zavisimost' protsessa kondensatsii khromatina ot temperatury i ionnoi sredy: issledovanie s pomoshch'iu gel'-élektroforeza v agaroze nizkoi plotnosti.

Conformational mobility of 30 nm nucleosomal fiber under temperature variations at different ionic strength of the medium has been analyzed. A new property of chromatin organization has been characterized, namely, nucleosomal solenoid condensation in response to increase of temperature. This property is analogous to salt-induced condensation and becomes particularly prominent at low ionic strength. In this context previously obtained data on changes in the DNA linking number of topological domains are discussed. It is concluded that these changes probably arise from modification of supernucleosomal level of chromatin organization.

[Study of the process of chromatin decondensation using low density agar gel electrophoresis]. / Izuchenie protsessa dekondensatsii khromatina s pomoshch'iu élektroforeza v geliakh agarozy nizkoi plotnosti.

A method was proposed for analysis of conformational mobility of supranucleosomal chromatin organization at different ionic conditions with the help of electrophoresis in low-density agarose gels. This simple and highly reproducible method yields the results which are in good agreement with the data of other traditional approaches. This method offers an alternative to high-speed ultracentrifugation for chromatin condensation studies.

Acetylation of core histones causes the unfolding of 30 nm chromatin fiber: analysis by agarose gel electrophoresis.

In the present work it was directly demonstrated that increased acetylation level of histones causes the decompactization of 30 nm chromatin fiber, as revealed by low-percentage agarose gel electrophoresis. In the light of obtained results the possible molecular mechanism of the decompactization of acetylated fiber is discussed.

Flexibility of DNA within transcriptionally active nucleosomes: analysis by circular dichroism measurements.

The conformational flexibility of DNA in transcriptionally active chromatin fractions has been estimated by circular dichroism spectroscopy analysis and was found to be restricted in the same fashion as in bulk chromatin. The observation is discussed in the context of different models of active chromatin organization.