ClaR—a novel key regulator of cellobiose and lactose metabolism in Lactococcus lactis IL1403

In a number of previous studies, our group has discovered an alternative pathway for lactose utilization in Lactococcus lactis that, in addition to a sugar-hydrolyzing enzyme with both P-β-glucosidase and P-β-galactosidase activity (BglS), engages chromosomally encoded components of cellobiose-specific PTS (PTSCel-Lac), including PtcA, PtcB, and CelB. In this report, we show that this system undergoes regulation via ClaR, a novel activator protein from the RpiR family of transcriptional regulators. Although RpiR proteins are widely distributed among lactic acid bacteria, their roles have yet to be confirmed by functional assays. Here, we show that ClaR activity depends on intracellular cellobiose-6-phosphate availability, while other sugars such as glucose or galactose have no influence on it. We also show that ClaR is crucial for activation of the bglS and celB expression in the presence of cellobiose, with some limited effects on ptcA and ptcB activation. Among 190 of carbon sources tested, the deletion of claR reduces L. lactis growth only in lactose- and/or cellobiose-containing media, suggesting a narrow specificity of this regulator within the context of sugar metabolism

Genome-wide analysis and expression profiling of calcium-dependent protein kinases in potato (Solanum tuberosum)

Calcium-dependent protein kinases (CDPKs or CPKs), unique to plants and some protists, are involved in growth and developmental processes as well as in defence against diverse environmental stresses. CDPKs are encoded by multi-gene families. Despite extensive studies of the CDPKs in many species, information about the evolutionary history and expression patterns of the CDPK family in the staple crop potato (Solanum tuberosum) remains poorly known. In this study, we performed bioinformatics analysis of the potato whole genome sequence and identified 23 potential CDPK genes. These genes are located in eleven, of twelve, potato chromosomes. Based on the phylogenetic tree and gene structures, the CDPKs were divided into four subfamilies. To determine their expression, reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis was carried out for the CDPK genes in different organs of potato such as young and mature leaves, stems, young shoots, roots, stolons, swollen stolons, flowers and tubers. The CDPKs were expressed in all the organs analysed, but their expression patterns varied greatly. The expression of some CDPKs was strongly organ specific, for example StCPK13 and StCPK18 was found only/ mostly in flowers. In Solanum genotypes differing in resistance to Phytophthora infestans, the expression and activity of CDPKs increased in response to a P. infestans elicitor with different kinetics and intensity. The expression levels and activity of the CDPKs correlated positively with the level of the resistance. Our results support earlier suggestion that CDPKs are involved in potato organ development and defence against stresses. We provide new information about the CDPK gene family in the potato and a perspective on its evolutionary history and biological roles of the individual kinases

Antihypertensive Efficacy and Safety of Lacidipine in the Treatment of Patients with Mild to Moderate Essential Hypertension - Multi-Center Study POL-LACY

Wstęp Lacydypina jest długodziałającym antagonistą wapnia z grupy pochodnych dihydropirydynowych i charakteryzuje się łagodnym początkiem działania hipotensyjnego i trwającym 24 h efektem hipotensyjnym. W badaniach z użyciem tego leku potwierdzono jej skuteczność hipotensyjną, a także korzystny wpływ na układ sercowo-naczyniowy. Celem wieloośrodkowego badania POL-LACY była ocena skuteczności hipotensyjnej oraz tolerancji lacydypiny podawanej raz dziennie w zakresie dawek 2&#8211;6 mg chorym z łagodnym i umiarkowanym nadciśnieniem tętniczym pierwotnym. Materiał i metody Otwarte badanie o charakterze otwartym przeprowadzono w 37 ośrodkach w Polsce, włączając do niego 470 chorych z łagodnym i umiarkowanym nadciśnieniem tętniczym pierwotnym. Badanie składało się z dwóch faz. W trakcie 7-dniowego okresu kwalifikacyjnego u chorych odstawiano stosowane wcześniej leki hipotensyjne oraz wykonywano badania laboratoryjne. W trakcie 8-tygodniowego okresu aktywnego leczenia chorzy otrzymywali lacydypinę w początkowej dawce dobowej 4 mg lub 2 mg u chorych powyżej 65 rż. pacjenci podawanej raz dziennie z możliwością zwiększenia dawki do 6 lub 4 mg na dobę u osób, u których nie uzyskano normalizacji ciśnienia po 4 tygodniach leczenia. Normalizację ciśnienia definiowano jako skurczowe ciśnienie tętnicze (SBP) o wartości < 140 mm Hg i rozkurczowe ciśnienie tętnicze (DBP) wynoszące < 90 mm Hg. W czasie leczenia chorzy odbyli 2 wizyty kontrolne, po 4 i po 8 tygodniach, w trakcie których dokonywano oceny osiągniętego efektu hipotensyjnego oraz tolerancji leczenia. Wyniki Badanie ukończyło 446 chorych (255 M, 191 K; średnia wieku 52 &plusmn; 13 lat). Pod koniec badania 9,9% chorych leczono dawką 2 mg, 72,9% chorych - dawką 4 mg, a 17,3% chorych - dawką 6 mg lacydypiny. Po 8 tygodniach leczenia tym preparatem w badanej grupie uzyskano znamienne obniżenie SBP o 23 mm Hg (p < 0,0001) i DBP o 15 mm Hg (p < 0,0001) w pozycji siedzącej. Uzyskany efekty hipotensyjny nie zależał od płci, wieku i wskaźnika masy ciała (BMI). Pełną normalizację wartości ciśnienia uzyskano u 71% chorych. W czasie badania stwierdzono istotne obniżenie średnich wartości ciśnienia tętna o 8 mm Hg (p < 0,0001). Lacydypina była dobrze tolerowana przez chorych, działania niepożądane wystąpiły u 21% z nich. Wnioski Lacydypina (Lacipil) stosowana w monoterapii charakteryzuje się wysoką skutecznością hipotensyjną u chorych z łagodnym i umiarkowanym nadciśnieniem tętniczym pierwotnym. Efekt hipotensyjny nie zależy od wieku i płci pacjentów. Preparat cechuje korzystny wpływ na wartości ciśnienia tętna uważanego za niezależny czynnik ryzyka powikłań sercowo-naczyniowych. Uzyskane wyniki wskazują na dobrą tolerancję leku przez chorych z nadciśnieniem tętniczym. Badanie wskazuje na przydatność leku w monoterapii nadciśnienia tętniczego.Background Lacidipine is an orally administered calcium channel blocker of the dihydropyridine class, which shows selectivity for vascular smooth muscle and has a long duration of action. The aim of multi-center study POL-LACY was to establish antihypertensive effect and safety of lacidipine given once a day 2 to 6 mg in patient with mild to moderate hypertension. Material and methods The study was an open, multi-centre study in 37 Polish medical centers. 470 patients with mild to moderate hypertension were included into the study. 7 days qualification period was followed by 8 weeks of active treatment. During qualification period antihypertensive treatment was withdrawn and patients underwent clinical evaluation. In the phase of active treatment patients were treated with lacidipine (Lacipil&reg;, GlaxoSmithKline Pharmaceuticals) starting from dose of 4 mg or 2 mg in patients older than 65 years. The dose could be tritated up to 6 or 4 mg if there was no normalization of BP level after 4 weeks of treatment. Normalization of BP was defined as systolic BP < 140 mm Hg and diastolic BP < 90 mm Hg. During phase of active treatment, after 4 and 8 weeks, efficacy and safety of treatment was evaluated. Results 446 patients completed the study (255 M, 191 F; mean age 52 &plusmn; 13 years). The mean baseline BP value was 157/98 mm Hg. At the end of the study 9,9% of patients were treated with 2 mg, 72,9% of patients with 4 mg and 17,3% of patients with 6 mg of lacidipine. After 8 weeks lacidipine decreased significantly systolic BP by 23 mm Hg (p < 0,0001) and diastolic BP by 15 mm Hg (p < 0,0001) in the sitting position. Antihypertensive effect of lacidipine was not influenced by sex, age and BMI. Full normalization of BP was achieved in 71% of patients. Pulse pressure was significantly reduced by 8 mm Hg (p < 0,0001). Lacidipine was well tolerated, 21% of patients experienced adverse events. Conclusions Lacidipine is a highly effective drug in the monotherapy of mild to moderate essential hypertension. Antihypertensive effect of lacidipine is not influenced by age, sex and BMI. Lacidipine has a positive effect on pulse pressure. Lacidipine is well tolerated by patients. In conclusion, presented data point at the lacidipine as a useful drug in monotherapy of essential hypertension

Expression of maize Calcium-Dependent Protein Kinase (ZmCPK11) improves salt tolerance in transgenic Arabidopsis plants by regulating sodium and potassium homeostasis and stabilizing photosystem II

In plants, CALCIUM-DEPENDENT PROTEIN KINASES (CDPKs/CPKs) are involved in calcium signaling in response to endogenous and environmental stimuli. Here, we report that ZmCPK11, one of maize CDPKs, participates in salt stress response and tolerance. Salt stress induced expression and upregulated the activity of ZmCPK11 in maize roots and leaves. Activation of ZmCPK11 upon salt stress was also observed in roots and leaves of transgenic Arabidopsis plants expressing ZmCPK11. The transgenic plants showed a long-root phenotype under control conditions and a short-root phenotype under NaCl, abscisic acid (ABA) or jasmonic acid (JA) treatment. Analysis of ABA and JA content in roots indicated that ZmCPK11 can mediate root growth by regulating the levels of these phytohormones. Moreover, 4-week-old transgenic plants were more tolerant to salinity than the wild-type plants. Their leaves were less chlorotic and showed weaker symptoms of senescence accompanied by higher chlorophyll content and higher quantum efficiency of photosystem II. The expression of Na+/K+ transporters (HKT1, SOS1 and NHX1) and transcription factors (CBF1, CBF2, CBF3, ZAT6 and ZAT10) with known links to salinity tolerance was upregulated in roots of the transgenic plants upon salt stress. Furthermore, the transgenic plants accumulated less Na+ in roots and leaves under salinity, and showed a higher K+/Na+ ratio in leaves. These results show that the improved salt tolerance in ZmCPK11-transgenic plants could be due to an upregulation of genes involved in the maintenance of intracellular Na+ and K+ homeostasis and a protection of photosystem II against damage

ERCC1-deficient cells and mice are hypersensitive to lipid peroxidation

Lipid peroxidation (LPO) products are relatively stable and abundant metabolites, which accumulate in tissues of mammals with aging, being able to modify all cellular nucleophiles, creating protein and DNA adducts including crosslinks. Here, we used cells and mice deficient in the ERCC1-XPF endonuclease required for nucleotide excision repair and the repair of DNA interstrand crosslinks to ask if specifically LPO-induced DNA damage contributes to loss of cell and tissue homeostasis. Ercc1-/- mouse embryonic fibroblasts were more sensitive than wild-type (WT) cells to the LPO products: 4-hydroxy-2-nonenal (HNE), crotonaldehyde and malondialdehyde. ERCC1-XPF hypomorphic mice were hypersensitive to CCl4 and a diet rich in polyunsaturated fatty acids, two potent inducers of endogenous LPO. To gain insight into the mechanism of how LPO influences DNA repair-deficient cells, we measured the impact of the major endogenous LPO product, HNE, on WT and Ercc1-/- cells. HNE inhibited proliferation, stimulated ROS and LPO formation, induced DNA base damage, strand breaks, error-prone translesion DNA synthesis and cellular senescence much more potently in Ercc1-/- cells than in DNA repair-competent control cells. HNE also deregulated base excision repair and energy production pathways. Our observations that ERCC1-deficient cells and mice are hypersensitive to LPO implicates LPO-induced DNA damage in contributing to cellular demise and tissue degeneration, notably even when the source of LPO is dietary polyunsaturated fats

DMSO and TMAO&mdash;Differences in Interactions in Aqueous Solutions of the K-Peptide

Interactions between a solvent and their co-solute molecules in solutions of peptides are crucial for their stability and structure. The K-peptide is a synthetic fragment of a larger hen egg white lysozyme protein that is believed to be able to aggregate into amyloid structures. In this study, a complex experimental and theoretical approach is applied to study systems comprising the peptide, water, and two co-solutes: trimethylamide N-oxide (TMAO) or dimethyl sulfoxide (DMSO). Information about their interactions in solutions and on the stability of the K-peptide was obtained by FTIR spectroscopy and differential scanning microcalorimetry. The IR spectra of various osmolyte&ndash;water&ndash;model-peptide complexes were simulated with the DFT method (B3LYP/6-311++G(d,p)). The FTIR results indicate that both solutes are neutral for the K-peptide in solution. Both co-solutes affect the peptide to different degrees, as seen in the shape of its amide I band, and have different influences on its thermal stability. DFT calculations helped simplify the experimental data for easier interpretation

Sequence-specific p53 gene damage by chloroacetaldehyde and its repair kinetics in Escherichia coli

Oxidative stress and certain environmental carcinogens, e.g. vinyl chloride and its metabolite chloroacetaldehyde (CAA), introduce promutagenic exocyclic adducts into DNA, among them 1,N6-ethenoadenine (εA), 3,N4-ethenocytosine (εC) and N2,3-ethenoguanine (εG). We studied sequence-specific interaction of the vinyl-chloride metabolite CAA with human p53 gene exons 5-8, using DNA Polymerase Fingerprint Analysis (DPFA), and identified sites of the highest sensitivity. CAA-induced DNA damage was more extensive in p53 regions which revealed secondary structure perturbations, and were localized in regions of mutation hot-spots. These perturbations inhibited DNA synthesis on undamaged template. We also studied the repair kinetics of CAA-induced DNA lesions in E. coli at nucleotide resolution level. A plasmid bearing full length cDNA of human p53 gene was modified in vitro with 360 mM CAA and transformed into E. coli DH5α strain, in which the adaptive response system had been induced by MMS treatment before the cells were made competent. Following transformation, plasmids were re-isolated from transformed cultures 35, 40, 50 min and 1-24 h after transformation, and further subjected to LM-PCR, using ANPG, MUG and Fpg glycosylases to identify the sites of DNA damage. In adaptive response-induced E. coli cells the majority of DNA lesions recognized by ANPG glycosylase were removed from plasmid DNA within 35 min, while MUG glycosylase excised base modifications only within 50 min, both in a sequence-dependent manner. In non-adapted cells resolution of plasmid topological forms was perturbed, suggesting inhibition of one or more bacterial topoisomerases by unrepaired ε-adducts. We also observed delayed consequences of DNA modification with CAA, manifesting as secondary DNA breaks, which appeared 3 h after transformation of damaged DNA into E. coli, and were repaired after 24 h