Interaction of calmodulin with Sec61α limits Ca2+ leakage from the endoplasmic reticulum

In eukaryotes, protein transport into the endoplasmic reticulum (ER) is facilitated by a protein-conducting channel, the Sec61 complex. The presence of large, water-filled pores with uncontrolled ion permeability, as formed by Sec61 complexes in the ER membrane, would seriously interfere with the regulated release of calcium from the ER lumen into the cytosol, an essential mechanism for intracellular signalling. We identified a calmodulin (CaM)-binding motif in the cytosolic N-terminus of mammalian Sec61α that bound CaM but not Ca2+-free apocalmodulin with nanomolar affinity and sequence specificity. In single-channel measurements, CaM potently mediated Sec61-channel closure in Ca2+-dependent manner. At the cellular level, two different CaM antagonists stimulated calcium release from the ER through Sec61 channels. However, protein transport into microsomes was not modulated by Ca2+-CaM. Molecular modelling of the ribosome/Sec61/CaM complexes supports the view that simultaneous ribosome and CaM binding to the Sec61 complex may be possible. Overall, CaM is involved in limiting Ca2+ leakage from the ER.     

28-homobrassinolide improves growth and photosynthesis in <Emphasis Type="Italic">Cucumis sativus</Emphasis> L. through an enhanced antioxidant system in the presence of chilling stress

The ameliorative role of 28-homobrassinolide under chilling stress in various growth, photosynthesis, enzymes and biochemical parameters of cucumber (Cucumis sativus L.) were investigated. Cucumber seedlings were sprayed with 0 (control), 10⁻⁸, or 10⁻⁶ M of 28-homobrassinolide at the 30-day stage. 48 h after treatment plants were exposed for 18 h to chilling temperature (10/8°C, 5/3°C). The most evident effect of chilling stress was the marked reduction in plant growth, chlorophyll (Chl) content, and net photosynthetic rate, efficiency of photosystem II and activities of nitrate reductase and carbonic anhydrase. Moreover, the activities of antioxidant enzymes; catalase (E.C. 1.11.1.6), peroxidase (E.C.1.11.1.7), superoxide dismutase (E.C. 1.15.1.1) along with the proline content in leaves of the cucumber seedlings increased in proportion to chilling temperature. The stressed seedlings of cucumber pretreated with 28-homobrassinolide maintained a higher value of antioxidant enzymes and proline content over the control suggesting the protective mechanism against the ill-effect caused by chilling stress might be operative through an improved antioxidant system. Furthermore, the protective role of 28-homobrassinolide was reflected in improved growth, water relations, photosynthesis and maximum quantum yield of photosystem II both in the presence and absence of chilling stress.     

Physiological changes in major soldiers of Macrotermes gilvus (Isoptera: Termitidae) induced by the endoparasitoid Misotermes mindeni (Diptera: Phoridae)

The majority of true parasitoids manipulate their host’s physiology for their own benefit. In this study, we documented the physiological changes that occurred in major soldiers of the subterranean termite Macrotermes gilvus (Hagen) (Isoptera: Termitidae) parasitized by the koinobiont larval endoparasitoid Misotermes mindeni Disney and Neoh (Diptera: Phoridae). We compared the metabolic rate, body water content, body water loss rate, cuticular permeability, and desiccation tolerance between parasitized and unparasitized major soldiers. The metabolic rate of parasitized hosts was significantly higher than that of unparasitized termites. Mean total body water content of parasitized major soldiers (64.73 ± 3.26%) was significantly lower than that of unparasitized termites (71.99 ± 2.23%). Parasitized hosts also had significantly lower total body water loss rates (5.72 ± 0.06%/h) and higher cuticular permeability (49.37 ± 11.26 μg/cm/h/mmHg) than unparasitized major soldiers (6.75 ± 0.16%/h and 60.76 ± 24.98 μg/cm/h/mmHg, respectively). Parasitized major soldiers survived almost twice as long as unparasitized termites (LT₅₀ = 6.66 h and LT₅₀ = 3.40 h, respectively) and they had significantly higher tolerance to water loss compared to unparasitized termites (45.28 ± 6.79% and 32.84 ± 7.69%, respectively). Body lipid content in parasitized hosts (19.84 ± 6.27%) was significantly higher than that of unparasitized termites (6.17 ± 7.87%). Finally, parasitized hosts had a significantly lower percentage of cuticular water content than unparasitized major soldiers (10.97 ± 1.84% and 13.17 ± 2.21%, respectively). Based on these data, we conclude that the parasitism-induced physiological changes in the host are beneficial to the parasitoids as the alterations can clearly increase the parasite’s chances of survival when exposed to extreme environmental conditions and ensure that the parasitoids are able to complete their larval development successfully before the host dies.     

Hematological and serum biochemical analyses in experimental caprine besnoitiosis

This study was undertaken to investigate the hematological and biochemical changes in experimentally infected goats with Besnoitia caprae from the time of infection till 360 days post-infection (PI). Six male goats were inoculated subcutaneously with 13 × 10(7) bradyzoites of B. caprae, and blood samples were collected from the jugular vein. The total erythrocyte and total leukocyte counts, hematocrit value, and differential leukocyte counts were determined. Serum biochemical analysis, including the total protein, albumin, total globulin, cholesterol, triglyceride, chloride, testosterone, calcium (Ca(2+)), inorganic phosphorus, sodium (Na(+)), potassium (K(+)), iron (Fe(2+)), glucose, serum amyloid A (SAA), haptoglobin (Hp), fibrinogen, ceruloplasmin, aspartate aminotransferase, alanine aminotransferase, creatine kinase, lactate dehydrogenase, and alkaline phosphatase, was undertaken. Skin biopsy from the limbs were collected at weekly intervals and histologically examined for Besnoitia cysts. Cysts were present in the skin biopsies of the leg of the infected goats from day 28 PI. There were variations in hematological analyses, but no significant difference was seen. From day 30 to 360 PI, results showed that SAA, Hp, fibrinogen, and ceruloplasmin concentrations increased, whereas testosterone concentrations decreased. Infected goats exhibited decrease of albumin and increase of serum total protein and globulin concentrations. By contrast, there were no significant differences in the remained analyses concentrations.     

Protein and DNA destabilization by osmolytes: the other side of the coin

Osmolytes are naturally occurring small molecules accumulated intracellularly to protect organisms from various denaturing stresses. Similar to the two faces of a coin, several of these osmolytes are stabilizing and destabilizing proteins depending on the concentrations and/or solvent conditions. For example, the well known stabilizing osmolyte, trehalose destabilizes some proteins at high concentration and/or high pH. In spite of the fact that destabilizing aspects of osmolytes can modulate many cellular processes including regulation of protein homeostasis (proteostasis), protein-protein interaction, and protein-DNA interaction, researchers have mostly focused on the stabilizing aspects of osmolytes. Thus, it is important to look into both aspects of osmolytes to determine their precise role under physiological conditions. In this article, we have discussed both stabilizing and destabilizing/denaturant aspects of osmolytes to uncover both sides of the coin.     

Genotypic characterization of Iranian camel (Camelus dromedarius) isolates of Echinoccocus granulosus

In the present study, Echinoccocus granulosus isolates collected from camels (Camelus dromedarius) in eastern Iran were characterized based on the nucleotide sequences of mitochondrial CO1 and NDI genes. The molecular results for camel isolates demonstrated that at least 2 different genotypes are present, i.e., a buffalo genotype (G3) and the camel genotype (G6). Although the sequences of the Iranian camel genotype (G6) are completely homologous to the reference sequence of G6 (M84666) of E. granulosus , a nucleotide mutation (C to T at position 168) was detected in the CO1 sequences of the Iranian G3 isolates (HM626405) when compared with the reference G3 genotype (M84663). The findings of the present study represent the demonstration of the buffalo strain in camels. As previously reported, humans can be infected by this genotype; accordingly, the epidemiological importance of this genotype in the camel population should be considered in further studies.     

Over-expression of FoxM1 leads to epithelial-mesenchymal transition and cancer stem cell phenotype in pancreatic cancer cells

FoxM1 is known to play important role in the development and progression of many malignancies including pancreatic cancer. Studies have shown that the acquisition of epithelial-to-mesenchymal transition (EMT) phenotype and induction of cancer stem cell (CSC) or cancer stem-like cell phenotypes are highly inter-related, and contributes to drug resistance, tumor recurrence, and metastasis. The molecular mechanism(s) by which FoxM1 contributes to the acquisition of EMT phenotype and induction of CSC self-renewal capacity is poorly understood. Therefore, we established FoxM1 over-expressing pancreatic cancer (AsPC-1) cells, which showed increased cell growth, clonogenicity, and cell migration. Moreover, over-expression of FoxM1 led to the acquisition of EMT phenotype by activation of mesenchymal cell markers, ZEB1, ZEB2, Snail2, E-cadherin, and vimentin, which is consistent with increased sphere-forming (pancreatospheres) capacity and expression of CSC surface markers (CD44 and EpCAM). We also found that over-expression of FoxM1 led to decreased expression of miRNAs (let-7a, let-7b, let-7c, miR-200b, and miR-200c); however, re-expression of miR-200b inhibited the expression of ZEB1, ZEB2, vimentin as well as FoxM1, and induced the expression of E-cadherin, leading to the reversal of EMT phenotype. Finally, we found that genistein, a natural chemo-preventive agent, inhibited cell growth, clonogenicity, cell migration and invasion, EMT phenotype, and formation of pancreatospheres consistent with reduced expression of CD44 and EpCAM. These results suggest, for the first time, that FoxM1 over-expression is responsible for the acquisition of EMT and CSC phenotype, which is in part mediated through the regulation of miR-200b and these processes, could be easily attenuated by genistein.     

Genetic Variability of Thermal <Emphasis Type="Italic">Nymphaea</Emphasis> (Nymphaeaceae) Populations Based on ISSR Markers: Implications on Relationships,Hybridization, and Conservation

The globally widespread genus Nymphaea exhibits a wide range of morphological and taxonomical diversity. The intrusion of a cultivated variety by progressive propagation and its affect on aquatic habitat is demonstrated in this case study. We have studied the genetic diversity, population, and stand structure of the neophyte Nymphaea × ‘Panama Pacific’ as well as other species found in Lake Hévíz and dikes nearby using inter-simple sequence repeat (ISSR) markers. The ISSR assay revealed a low genetic variability for the small populations of Nymphaea caerulea, Nymphaea lotus var. thermalis, and a medium level for Nymphaea alba, Nymphaea rubra var. longiflora, and Nymphaea × ‘Panama Pacific’. The evolutionary genetic status of individuals found in the overlapping cultivation area of Nymphaea × ‘Panama Pacific’ and N. caerulea was affirmed to be of hybrid origin by reticulate network analysis and with morphological parameters. The Bayesian analysis of hybrid classes and the segregation of the ISSR markers also confirmed the hybrid origin of the individuals in question and revealed that they are falling into F2 or latter genotype frequency classes, indicating the viability and fertility of the hybrids. The set of analyzed species by phylogenetic network analysis of ISSR data has been divided into three major groups according to their evolutionary patterns (subg. Barachyceras, Lotos, and Nymphaea). Our results are in accordance with these three major subgenera within Nymphaea.     

Structure and stability analysis of cytotoxic complex of camel α-lactalbumin and unsaturated fatty acids produced at high temperature

α-Lactalbumin α-La), together with oleic acid can be converted to a complex, which kills tumor cells selectively. Cytotoxic α-La -oleic acid and α-La -linoleic acid complexes were generated by adding fatty acid to camel holo α-La at 60 ° C (referred to as La-OA-60 and La-LA-60 state, respectively). Structural properties of these complexes were studied and compared to the camel α-La. The experimental results show that linoleic acid induces α-La partial unfolding but oleic acid does not change the protein structure significantly. Also the stability of La-OA-60 and La-LA-60 toward thermal denaturation was measured. The order of temperature at the transition midpoint is as follows: La-LA-60 < La-OA-60 < α-La. La-OA-60 complex inhibited tubulin polymerization in vitro. Although the structures of La-OA-60 and La-LA-60 were different, these two complexes had similar cytotoxic effect to DU145 human prostate cancer cells. Samples of La-OA-60 that have been renatured after denaturation lost the specific biological activity toward tumor cells.