Overexpression of the cytosolic cytokinin oxidase/dehydrogenase (CKX7) from Arabidopsis causes specific changes in root growth and xylem differentiation

Degradation of the plant hormone cytokinin is catalyzed by cytokinin oxidase/dehydrogenase (CKX) enzymes. The Arabidopsis thaliana genome encodes seven CKX proteins which differ in subcellular localization and substrate specificity. Here we analyze the CKX7 gene, which to the best of our knowledge has not yet been studied. pCKX7:GUS expression was detected in the vasculature, the transmitting tissue and the mature embryo sac. A CKX7–GFP fusion protein localized to the cytosol, which is unique among all CKX family members. 35S:CKX7‐expressing plants developed short, early terminating primary roots with smaller apical meristems, contrasting with plants overexpressing other CKX genes. The vascular bundles of 35S:CKX7 primary roots contained only protoxylem elements, thus resembling the wol mutant of the CRE1/AHK4 receptor gene. We show that CRE1/AHK4 activity is required to establish the CKX7 overexpression phenotype. Several cytokinin metabolites, in particular cis‐zeatin (cZ) and N‐glucoside cytokinins, were depleted stronger in 35S:CKX7 plants compared with plants overexpressing other CKX genes. Interestingly, enhanced protoxylem formation together with reduced primary root growth was also found in the cZ‐deficient tRNA isopentenyltransferase mutant ipt2,9. However, different cytokinins were similarly efficient in suppressing 35S:CKX7 and ipt2,9 vascular phenotypes. Therefore, we hypothesize that the pool of cytosolic cytokinins is particularly relevant in the root procambium where it mediates the differentiation of vascular tissues through CRE1/AHK4. Taken together, the distinct consequences of CKX7 overexpression indicate that the cellular compartmentalization of cytokinin degradation and substrate preference of CKX isoforms are relevant parameters that define the activities of the hormone.     

Detection of self-incompatible oilseed rape plants (Brassica napus L.) based on molecular markers for identification of the class I S haplotype

The selection of desirable genotypes with recessive characteristics, such as self-incompatible plants, is often difficult or even impossible and represents a crucial barrier in accelerating the breeding process. Molecular approaches and selection based on molecular markers can allow breeders to overcome this limitation. The use of self-incompatibility is an alternative in hybrid breeding of oilseed rape. Unfortunately, stable self-incompatibility is recessive and phenotype-based selection is very difficult and time-consuming. The development of reliable molecular markers for detecting desirable plants with functional self-incompatible genes is of great importance for breeders and allows selection at early stages of plant growth. Because most of these reliable molecular markers are based on discrimination of class I S-locus genes that are present in self-compatible plants, there is a need to use an internal control in order to detect possible PCR inhibition that gives false results during genotyping. In this study, 269 double haploid F₂ oilseed rape plants obtained by microspore embryogenesis were used to verify the applicability of an improved PCR assay based on the detection of the class I SLG gene along with an internal control. Comparative analysis of the PCR genotyping results vs. S phenotype analysis confirmed the applicability of this molecular approach in hybrid breeding programs. This approach allows accurate detection of self-incompatible plants via a different amplification profile.     

PPARA Intron Polymorphism Associated with Power Performance in 30-s Anaerobic Wingate Test

To date, polymorphisms in several genes have been associated with a strength/power performance including alpha 3 actinin, ciliary neurotrophic factor, vitamin D receptor, or angiotensin I converting enzyme, underlining the importance of genetic component of the multifactorial strength/power-related phenotypes. The single nucleotide variation in peroxisome proliferator-activated receptor alpha gene (PPARA) intron 7 G/C (rs4253778; g.46630634G>C) has been repeatedly found to play a significant role in response to different types of physical activity. We investigated the effect of PPARA intron 7 G/C polymorphism specifically on anaerobic power output in a group of 77 elite male Czech ice hockey players (18–36 y). We determined the relative peak power per body weight (P_max.kg⁻¹) and relative peak power per fat free mass (W.kg⁻¹ _FFM) during the 30-second Wingate Test (WT30) on bicycle ergometer (Monark 894E Peak bike, MONARK, Sweden). All WT30s were performed during the hockey season. Overall genotype frequencies were 50.6% GG homozygotes, 40.3% CG heterozygotes, and 9.1% CC homozygotes. We found statistically significant differences in P_max.kg⁻¹ and marginally significant differences in P_max.kg⁻¹ _FFM values in WT30 between carriers and non-carriers for C allele (14.6±0.2 vs. 13.9±0.3 W.kg⁻¹ and 15.8±0.2 vs. 15.2±0.3 W.kg⁻¹ _FFM, P = 0.036 and 0.12, respectively). Furthermore, P_max.kg⁻¹ _FFM strongly positively correlated with the body weight only in individuals with GG genotypes (R = 0.55; p<0.001). Our results indicate that PPARA 7C carriers exhibited higher speed strength measures in WT30. We hypothesize that C allele carriers within the cohort of trained individuals may possess a metabolic advantage towards anaerobic metabolism.     

Pdsg1 and Pdsg2, Novel Proteins Involved in Developmental Genome Remodelling in Paramecium

The epigenetic influence of maternal cells on the development of their progeny has long been studied in various eukaryotes. Multicellular organisms usually provide their zygotes not only with nutrients but also with functional elements required for proper development, such as coding and non-coding RNAs. These maternally deposited RNAs exhibit a variety of functions, from regulating gene expression to assuring genome integrity. In ciliates, such as Paramecium these RNAs participate in the programming of large-scale genome reorganization during development, distinguishing germline-limited DNA, which is excised, from somatic-destined DNA. Only a handful of proteins playing roles in this process have been identified so far, including typical RNAi-derived factors such as Dicer-like and Piwi proteins. Here we report and characterize two novel proteins, Pdsg1 and Pdsg2 (Paramecium protein involved in Development of the Somatic Genome 1 and 2), involved in Paramecium genome reorganization. We show that these proteins are necessary for the excision of germline-limited DNA during development and the survival of sexual progeny. Knockdown of PDSG1 and PDSG2 genes affects the populations of small RNAs known to be involved in the programming of DNA elimination (scanRNAs and iesRNAs) and chromatin modification patterns during development. Our results suggest an association between RNA-mediated trans-generational epigenetic signal and chromatin modifications in the process of Paramecium genome reorganization.     

Salicylic acid regulates secondary metabolites content in leaves of Matricaria chamomilla

The influence of increasing doses of salicylic acid (SA) on selected physiological parameters and the content of coumarin-related compounds of diploid and tetraploid cultivars of Matricaria chamomilla plants were studied. Forty-eight hours after treatment SA showed growth-promoting effect with decrease in tissue water content, chlorophylls and soluble proteins. High doses of SA led to an increase of reactive oxygen species (hydrogen peroxide and superoxide radical) resulted in membrane damage (measured by accumulation of malondialdehyde). Changes in secondary metabolites accumulation in leaves were also observed. The pattern of quantitative changes of studied compounds was similar in tetraploid and diploid plants. The content of herniarin and its precursor (Z)- and (E)-2-β-d-glucopyranosyloxy-4-methoxycinnamic acid increased simultaneously. A considerable increase of umbelliferone and decrease in chlorogenic acid was registered. The rise of ene-yne-dicycloether in treated plant was also detected.     

Synthesis and antiproliferative activity of novel steroidal dendrimer conjugates

We describe the synthesis of steroidal dendrimer conjugates of first and second generation with tetramethylene core and 5-hydroxy-isophtalic acid dimethyl ester as branching unit modified to incorporate ethynylestradiol or 17α-estradiol as terminal units. The steroidal dendrimer conjugates, the free drug (steroids) and dendrimer were tested against a panel of cancer cell lines (CEM, MCF7, HeLa) and normal human fibroblast (BJ). The steroidal dendrimer conjugates of first generation exhibited cytotoxic activity and induced apoptosis in chronic leukemia (CEM) as resultant activation of caspase cascade which is mainly provoked in G2/M arrested cells.     

Evaluation of the strain identity between isolates from caries lesions and root canals in early childhood caries cases

Early childhood caries (ECC) has become a serious medical problem worldwide in the last decade. Bacterial microflora of the dental plaque and oral cavity is considered an important factor in the formation and progression of dental caries. The aim of this study was strain typing and comparison of bacterial isolates retrieved from caries lesions and root canal contents of the same teeth. In total, 18 pairs of presumptive streptococci and lactobacilli retrieved from dental caries and root canals isolated from ECC-affected children, were selected on the basis of biotyping results and rep-PCR fingerprinting with (GTG)₅ primer. Strain typing was further done using the RiboPrinter microbial characterization system (DuPont Qualicon). The automated ribotyping determined 14 pairs of the strains (77.8 %) to be identical. The results obtained confirmed that identical bacterial strains colonized both the decayed dental surface and the necrotic content of the dental pulp cavity during the cariogenesis. Our finding supports the assumption that bacteria could penetrate through the damaged dental surface to the inner parts of the teeth.     

Ability of mammalian fibroblasts to grow in synthetic medium containing neither serum nor exogenously added macromolecules

Summary Rous sarcoma virus transformed Chinese hamster fibroblasts, clone CHR1-3, were established at high temperature, then subcloned. Six subclones with round and flat morphology harboring undeleted and partially deleted RSV proviruses, respectively, were seeded into serum-free synthetic medium with no macromolecular additives, and maintained for 2 mo. One flat subclone no. 14, fully designatedsfCHR1-3.14 for itsserum-free phenotype, was further propagated in the same medium. The cells grew exponentially in loosely attached monolayers and dould be serially passaged on bare polystyrene with an average population doubling time of 46 h. Cell attachment could be improved by using collagen-coated polystyrene or by adding a methionine supplement to the culture medium. Furthermore, thesfCHR1-3.14 cells could be subcloned and further grown in nonselective medium. The reversion rate of thesf phenotype was estimated to be 1 to 2%/cell generation. Evidence for an autocrinal stimulation was obtained by cloning efficiency assays showing a requirement for a threshold cell density. Slight growth stimulation could also be detected in assays using conditioned medium fromsfCHR1-3.14 cells and serum-restrictedwild-type (wt)NIH3T3, but notwtCHR1-3.14, cells as indicator cells. Finally,wtNIH3T3 cells used in these assays were assayed for serum-free growth and found to be able to develop their ownsf phenotype; in this respect they resemble the previously establishedsfCHR1-3.14 cells. Supported by grants from CNRS, INSERM, contract 852012, Association pour la Recherche sur le Cancer, and Fondation pour la Recherche Médicale. V. K. was supported by the Association pour la Recherche sur le Cancer and by the Fédération Nationale des Centres de Lutte contre le Cancer.     

Growth and reproduction of monoxenic and polyxenic cultures of the slug-parasitic nematodes Phasmarhabditis spp. in naturally occurring organic substrates

BioControl - In the present study, we studied ecology of three species of mollusc-parasitic nematodes: Phasmarhabditis bohemica, P. bonaquaense and P. apuliae. We demonstrate that these facultative...     

Energetics in a solitary subterranean rodent, the silvery mole-rat, Heliophobius argenteocinereus, and allometry of RMR in African mole-rats (Bathyergidae)

Low resting metabolic rate (RMR) in subterranean rodents used to be considered as a physiological adaptation to cope with stresses of the belowground environment. In African mole-rats (Bathyergidae, Rodentia), RMR was reported to be independent of body mass. This deviation from a general mammalian pattern was considered a precondition for evolution of eusociality, occurring in some bathyergids. We measured metabolic rate and thermoregulation in the silvery mole-rat, Heliophobius argenteocinereus, the only bathyergid genus for which well-supported, comparable data were still missing. Low RMR (154.04 mL O(2) h(-1), which is 82% of the value predicted for a rodent) corresponds to the value expected in a subterranean rodent. Broad range of the thermoneutral zone (25-33 degrees C) and only slightly higher conductance (17.3 mL O(2) h(-1) degrees C(-1), i.e. 112.5% of that predicted for subterranean mammals) indicate that H. argenteocinereus is adapted to lower burrow temperatures rather than to high temperatures. Low RMR in this solitary species, as in other subterranean rodents in general, is probably associated particularly with high energetic cost of foraging. Our results combined with data on other mole-rats show clearly that RMR within the Bathyergidae is mass-dependent.