Isolation and characterisation of an antifolate insensitive (afi1) mutant of Arabidopsis thaliana.

Antifolates can impair the synthesis and/or function of folates in living organisms. Mechanisms of resistance or tolerance to antifolates have been mainly described in plants using the drug methotrexate. In this work, the antifolate trimethoprim (TMP) was used with the aim of revealing a novel mechanism of resistance. EMS mutagenised seeds from Arabidopsis were screened to isolate individuals insensitive to TMP. Genetic analysis revealed a homozygous recessive mutation that segregates with the phenotype of tolerance to 50 µm TMP. Mapping analysis localised the mutation at the end of the short arm of chromosome 3. Preliminary characterisation demonstrated up-regulation of several genes from the folate biosynthetic pathway in the TMP insensitive mutant, and a slight increase in total folate content in the mutant as compared with the Col-0 control. Moreover, sequence analysis of the DHFR (dihydrofolate reductase) genes, which encode a known target for resistance to antifolates, did not reveal any changes. This study is the first report of a stable mutant insensitive (afi1) to the antifolate trimethoprim in plants, and suggests the existence of a novel mechanism of resistance to antifolates.

[Kidney function in oncologic patients after cisplastin chemotherapy].

During examination of kidney function under cisplastin chemotherapy in oncologic patients the nephrotoxicity of the drug have been shown during 24 hours after administration. We found that in 24 hours after injection ofcisplastin in the dose 50 mg/m2 of the surface of the body, the excretion of creatinine, sodium, potassium, osmotic active compounds and chlorides is increased more than two times, indicating for disorder in osmolality, volume and ionic regulation of kidney function. The determined peculiarities are consequences of kidney injuries, which are confirmed by an increase in erythrocyte- and leucocyteuria. We showed that alterations in homeostatic kidney function is due to damage of renal tubules. Our study necessitates a need for renal protection during the whole period of cisplastin chemotherapy.

[Potentialities of medicinal correction of cerebrovascular disturbances in patients with arterial hypertension].

The use of up-to-date diagnostic methods for the examination of 72 patients presenting with grade II-III arterial hypertension and high risk of cardiovascular complications made it possible to estimate the state of brachiocephalic arteries before and after combined antihypertensive therapy that continued during 14 days. The following variables were measured: blood flow rate, peripheral vascular resistance indices at the extra- and intracranial levels, metabolic indices of cerebrovascular responsiveness, and coefficient of variability as an indicator of the cerebrovascular reserve capacity under conditions of antihypertensive treatment. Three antihypertensive therapeutic regimens were employed, viz. lisinopril plus indapamide, bisoprolol plus indapamide, and amlodipine plus indapamide. It was shown that all the three regimen resulted in positive changes in the parameters of blood flow and peripheral vascular resistance. At the same time, the use of amlodipine plus indapamide ensured a more gradual reduction of peripheral vascular resistance and the most pronounced increase of cerebrovascular reserve capacity compared with the two remaining regimens. It is concluded that the parameters of peripheral vascular resistance and metabolic indices of cerebrovascular responsiveness are the most sensitive and informative end points for the pharmacotherapeutic treatment of cerebral hemodynamics in the patients presenting with grade II-III arterial hypertension and high risk of development of cardiovascular complications.

Willingness-to-accept and purchase genetically modified rice with high folate content in Shanxi Province, China.

Neural-tube defects (NTDs) are considered to be the most common congenital malformations. As Shanxi Province, a poor region in the North of China, has one of the highest reported prevalence rates of NTDs in the world, folate fortification of rice is an excellent alternative to low intake of folate acid pills in this region. This paper investigates the relations between socio-demographic indicators, consumer characteristics (knowledge, consumer perceptions on benefits, risks, safety and price), willingness-to-accept and willingness-to-pay genetically modified (GM) rice. The consumer survey compromises 944 face-to-face interviews with rice consumers in Shanxi Province, China. Multivariate analyses consist of multinomial logistic regression and multiple regression. The results indicate that consumers generally are willing-to-accept GM rice, with an acceptance rate of 62.2%. Acceptance is influenced by objective knowledge and consumers' perceptions on benefits and risks. Willingness-to-pay GM rice is influenced by objective knowledge, risk perception and acceptance. Communication towards the use of GM rice should target mainly improving knowledge and consumers' perceptions on high-risk groups within Shanxi Province, in particular low educated women.

Arabidopsis coactivator ALY-like proteins, DIP1 and DIP2, interact physically with the DNA-binding domain of the Zn-finger poly(ADP-ribose) polymerase.

Two novel homologous Arabidopsis proteins, DIP1 and DIP2, interact with the DNA-binding domain of plant poly(ADP-ribose) polymerase (PARP) in the yeast two-hybrid system and in vitro. Their homology to the transcriptional coactivator ALY suggests that plant PARP may play a role in the regulation of transcription.

A mutation of the mitochondrial ABC transporter Sta1 leads to dwarfism and chlorosis in the Arabidopsis mutant starik.

A mutation in the Arabidopsis gene STARIK leads to dwarfism and chlorosis of plants with an altered morphology of leaf and cell nuclei. We show that the STARIK gene encodes the mitochondrial ABC transporter Sta1 that belongs to a subfamily of Arabidopsis half-ABC transporters. The severity of the starik phenotype is suppressed by the ectopic expression of the STA2 homolog; thus, Sta1 function is partially redundant. Sta1 supports the maturation of cytosolic Fe/S protein in Deltaatm1 yeast, substituting for the ABC transporter Atm1p. Similar to Atm1p-deficient yeast, mitochondria of the starik mutant accumulated more nonheme, nonprotein iron than did wild-type organelles. We further show that plant mitochondria contain a putative l-cysteine desulfurase. Taken together, our results suggest that plant mitochondria possess an evolutionarily conserved Fe/S cluster biosynthesis pathway, which is linked to the intracellular iron homeostasis by the function of Atm1p-like ABC transporters.

The heat-shock element is a functional component of the Arabidopsis APX1 gene promoter.

Ascorbate peroxidases are important enzymes that detoxify hydrogen peroxide within the cytosol and chloroplasts of plant cells. To better understand their role in oxidative stress tolerance, the transcriptional regulation of the apx1 gene from Arabidopsis was studied. The apx1 gene was expressed in all tested organs of Arabidopsis; mRNA levels were low in roots, leaves, and stems and high in flowers. Steady-state mRNA levels in leaves or cell suspensions increased after treatment with methyl viologen, ethephon, high temperature, and illumination of etiolated seedlings. A putative heat-shock cis element found in the apx1 promoter was shown to be recognized by the tomato (Lycopersicon esculentum) heat-shock factor in vitro and to be responsible for the in vivo heat-shock induction of the gene. The heat-shock cis element also contributed partially to the induction of the gene by oxidative stress. By using in vivo dimethyl sulfate footprinting, we showed that proteins interacted with a G/C-rich element found in the apx1 promoter.

Higher plants possess two structurally different poly(ADP-ribose) polymerases.

One of the immediate reactions of the mammalian cell to many environmental stresses is a massive synthesis of poly(ADP-ribose), catalyzed by poly(ADP-ribose) polymerase (PARP). Most of the biological functions attributed to PARP are inferred from experimentation with mammalian cells. In plants, the biology of PARP may be more complicated and diverse than was previously thought. Two poly(ADP-ribose) polymerase homologues were found in plants, the classical Zn-finger-containing polymerase (ZAP) and the structurally non-classical PARP proteins (APP and NAP), which lack the characteristic N-terminal Zn-finger domain. By enzymatic and cytological experiments the recombinant APP protein was shown to be located in the nucleus and to possess DNA-dependent poly(ADP-ribose) polymerase activity in yeast. The nuclear localization was further confirmed by the analysis of transgenic tobacco plants that expressed a translational gene fusion between APP and the bacterial beta-glucuronidase. The app promoter was transcriptionally up-regulated in cells pre-determined to die because of deficiency in a DNA ligase I.

Identification of an Arabidopsis thaliana cDNA encoding a HSP70-related protein belonging to the HSP110/SSE1 subfamily.

Heat-shock protein 70 (HSP70)-related proteins are classified in two main subfamilies: the DnaK subfamily and the HSP110/SSE1 subfamily. We have characterized the first plant member of the HSP110/SSE1 subfamily, HSP91. At least two, tightly linked genes encoding HSP91 are present per haploid Arabidopsis genome. HSP91 is constitutively expressed in non-stressed Arabidopsis plants and is transiently induced by heat shock.

[The import of DNA topoisomerase type II from Drosophila melanogaster into the chloroplasts of transgenic Nicotiana tabacum plants]. / Import DNK-topoizomerazy II tipa Drosophila melanogaster v khloroplasty transgennykh rastenii Nicotiana tabacum.

The construct containing the structural gene of the type II DNA topoisomerase of Drosophila melanogaster fused to the sequence of the chloroplast transit peptide directed by the 35 S promoter was produced on the basis of the binary vector Bin 19. The construct was introduced into Nicotiana tabacum plants by Agrobacterium-mediated transformation. The integration of the construct into genomic DNA was demonstrated by PCR and Southern blot hybridization. Expression of the chimeric gene at the transcription level was studied by Northern blotting. The protein produce of the expression was identified in chloroplasts by Western blot hybridization with polyclonal anti-type II Drosophila topoisomerase antibodies.