Two carbon fluxes to reserve starch in potato (Solanum tuberosum L.) tuber cells are closely interconnected but differently modulated by temperature

Parenchyma cells from tubers of Solanum tuberosum L. convert several externally supplied sugars to starch but the rates vary largely. Conversion of glucose 1-phosphate to starch is exceptionally efficient. In this communication, tuber slices were incubated with either of four solutions containing equimolar [U-1?C]glucose 1-phosphate, [U-1?C]sucrose, [U-1?C]glucose 1-phosphate plus unlabelled equimolar sucrose or [U-1?C]sucrose plus unlabelled equimolar glucose 1-phosphate. C1?-incorporation into starch was monitored. In slices from freshly harvested tubers each unlabelled compound strongly enhanced 1?C incorporation into starch indicating closely interacting paths of starch biosynthesis. However, enhancement disappeared when the tubers were stored. The two paths (and, consequently, the mutual enhancement effect) differ in temperature dependence. At lower temperatures, the glucose 1-phosphate-dependent path is functional, reaching maximal activity at approximately 20 °C but the flux of the sucrose-dependent route strongly increases above 20 °C. Results are confirmed by in vitro experiments using [U-1?C]glucose 1-phosphate or adenosine-[U-1?C]glucose and by quantitative zymograms of starch synthase or phosphorylase activity. In mutants almost completely lacking the plastidial phosphorylase isozyme(s), the glucose 1-phosphate-dependent path is largely impeded. Irrespective of the size of the granules, glucose 1-phosphate-dependent incorporation per granule surface area is essentially equal. Furthermore, within the granules no preference of distinct glucosyl acceptor sites was detectable. Thus, the path is integrated into the entire granule biosynthesis. In vitro C1?C-incorporation into starch granules mediated by the recombinant plastidial phosphorylase isozyme clearly differed from the in situ results. Taken together, the data clearly demonstrate that two closely but flexibly interacting general paths of starch biosynthesis are functional in potato tuber cells.     

Pyrrolovesamicols--synthesis, structure and VAChT binding of two 4-fluorobenzoyl regioisomers

This Letter describes the synthesis of two regioisomers of a new class of vesamicol analogs as possible ligands for imaging the vesicular acetylcholine transporter in future PET studies. The two pyrrolovesamicols (±)-6a and (±)-6b were synthesized by nucleophilic ring opening reaction of a tetrahydroindole epoxide precursor with 4-phenylpiperidine. The reaction mechanism of the synthesis was studied by HPLC and the molecular structures were determined by X-ray structure analysis. Unexpected low binding affinities to VAChT (K(i)=312±73 nM for (±)-6a and K(i)=7320±1840 nM for (±)-6b) were determined by competitive binding analysis using a cell line stably transfected with ratVAChT and (-)-[(3)H]vesamicol.     

Movement of Cicindela hirticollis Say Larvae in Response to Moisture and Flooding

The larvae of the tiger beetle, Cicindela hirticollis Say, inhabit sandy shoreline areas that flood periodically. This species has declined over much of its range and at least one subspecies is near extinction, possibly as a result of human alteration of waterways. In addition to physiological tolerance for anoxia, the larvae have physical and behavioral adaptations to avoid drowning. We hypothesized that C. hirticollis larvae would exhibit behavioral responses to soil moisture change and flooding because, unlike most other tiger beetles, they frequently relocate their burrows. Our laboratory studies demonstrated that larvae select surface soil moisture levels of 7–50% saturation in which to dig new burrows. Within 96 h of immersion, most larvae abandon burrows and larvae do not form new burrows in darkness. Larvae may relocate when flooded, suggesting a previously undocumented mechanism for dispersal; however, dams often eliminate suitable habitat areas downstream, suggesting that this behavior may be detrimental in riverine populations. Because larvae move during daylight hours, they also are likely to suffer mortality from trampling due to human recreational activity.     

Cyclophilin D links programmed cell death and organismal aging in Podospora anserina

Cyclophilin D (CYPD) is a mitochondrial peptidyl prolyl‐cis,trans‐isomerase involved in opening of the mitochondrial permeability transition pore (mPTP). CYPD abundance increases during aging in mammalian tissues and in the aging model organism Podospora anserina. Here, we show that treatment of the P. anserina wild‐type with low concentrations of the cyclophilin inhibitor cyclosporin A (CSA) extends lifespan. Transgenic strains overexpressing PaCypD are characterized by reduced stress tolerance, suffer from pronounced mitochondrial dysfunction and are characterized by accelerated aging and induction of cell death. Treatment with CSA leads to correction of mitochondrial function and lifespan to that of the wild‐type. In contrast, PaCypD deletion strains are not affected by CSA within the investigated concentration range and show increased resistance against inducers of oxidative stress and cell death. Our data provide a mechanistic link between programmed cell death (PCD) and organismal aging and bear implications for the potential use of CSA to intervene into biologic aging.     

Decrease of Extracellular Taurine in the Rat Dorsal Hippocampus After Central Nervous Administration of Vasopressin

The extracellular amino acid concentrations in the left and right dorsal hippocampus of male rats were studied before and during application of vasopressin into the right hippocampus. The method of intracerebral microdialysis was used for both arginine vasopressin administration and monitoring of the composition of the extracellular fluid. The concentrations of 16 amino acids were measured by HPLC in the perfusate samples. The level of taurine declined 20% in the right hippocampus during perfusion with vasopressin, whereas o-phosphoethanolamine decreased in both sides, the left 20% and the right 24%. These alterations may be related to cerebral osmoregulation. Also, the levels of tyrosine and phenylalanine increased 15% and 35%, respectively, during administration of vasopressin. No changes of other amino acids were observed.     

Modal gating of human CaV2.1 (P/Q-type) calcium channels: I. The slow and the fast gating modes and their modulation by beta subunits

The single channel gating properties of human CaV2.1 (P/Q-type) calcium channels and their modulation by the auxiliary beta1b, beta2e, beta3a, and beta4a subunits were investigated with cell-attached patch-clamp recordings on HEK293 cells stably expressing human CaV2.1 channels. These calcium channels showed a complex modal gating, which is described in this and the following paper (Fellin, T., S. Luvisetto, M. Spagnolo, and D. Pietrobon. 2004. J. Gen. Physiol. 124:463-474). Here, we report the characterization of two modes of gating of human CaV2.1 channels, the slow mode and the fast mode. A channel in the two gating modes differs in mean closed times and latency to first opening (both longer in the slow mode), in voltage dependence of the open probability (larger depolarizations are necessary to open the channel in the slow mode), in kinetics of inactivation (slower in the slow mode), and voltage dependence of steady-state inactivation (occurring at less negative voltages in the slow mode). CaV2.1 channels containing any of the four beta subtypes can gate in either the slow or the fast mode, with only minor differences in the rate constants of the transitions between closed and open states within each mode. In both modes, CaV2.1 channels display different rates of inactivation and different steady-state inactivation depending on the beta subtype. The type of beta subunit also modulates the relative occurrence of the slow and the fast gating mode of CaV2.1 channels; beta3a promotes the fast mode, whereas beta4a promotes the slow mode. The prevailing mode of gating of CaV2.1 channels lacking a beta subunit is a gating mode in which the channel shows shorter mean open times, longer mean closed times, longer first latency, a much larger fraction of nulls, and activates at more positive voltages than in either the fast or slow mode.     

Reflective and black mulch increase yield in pumpkins under virus disease pressure

Experiments were conducted over a 4-yr period that examined the effect reflective, black, and no-mulch had on aphid populations, incidence of virus infected pumpkin plants, and yield of pumpkin. Three different planting dates and their effect on viral infection and yield were also investigated. The use of reflective mulches significantly reduced the number of alates landing in these rows compared with black- or no-mulch rows. The reduction in aphid numbers resulted in a reduction in the percent of plants infected with virus. Approximately 10 times more plants were infected with virus in the black and no-mulch plots than in the reflective mulch plots in mid- to late July. However, by the end of August, all treatments had near 100% of plants infected with virus. This delay in viral infection in reflective mulch plots resulted in a 45 and 120% increase in pumpkin yield compared with black mulch and no-mulch plots, respectively. First plantings always had greater yields than later plantings. The interaction between mulch type and planting time was significant. Reflective mulches increased yields overall, but significantly increased yields compared with black or no-mulch in second and third plantings. The use of reflective mulch combined with timely planting is a cost effective way of delaying virus problems and increasing pumpkin yields in midwestern United States.     

Proliferating cell nuclear antigen (PCNA) as a proliferative marker during embryonic and adult zebrafish hematopoiesis

We investigated the expression of proliferative cell nuclear antigen (PCNA) in zebrafish to delineate the proliferative hematopoietic component during adult and embryonic hematopoiesis. Immunostaining for PCNA and enhanced green fluorescence protein (eGFP) was performed in wild-type and fli1-eGFP (endothelial marker) and gata1-eGFP (erythroid cell marker) transgenic fish. Expression of PCNA mRNA was examined in wild-type and chordin morphant embryos. In adult zebrafish kidney, the renal tubules are surrounded by endothelial cells and it is separated into hematopoietic and excretory compartments. PCNA was expressed in hematopoietic progenitor cells but not in mature neutrophils, eosinophils or erythroid cells. Some PCNA+ cells are scattered in the hematopoietic compartment of the kidney while others are closely associated with renal tubular cells. PCNA was also expressed in spermatogonial stem cells and intestine crypts, consistent with its role in cell proliferation and DNA synthesis. In embryos, PCNA is expressed in the brain, spinal cord and intermediate cell mass (ICM) at 24 h-post fertilization. In chordin morphants, PCNA is significantly upregulated in the expanded ICM. Therefore, PCNA can be used to mark cell proliferation in zebrafish hematopoietic tissues and to identify a population of progenitor cells whose significance would have to be further investigated.