Transport of galectin-3 between the nucleus and cytoplasm. II. Identification of the signal for nuclear export

Galectin-3, a factor involved in the splicing of pre-mRNA, shuttles between the nucleus and the cytoplasm. Previous studies have shown that incubation of fibroblasts with leptomycin B resulted in the accumulation of galectin-3 in the nucleus, suggesting that the export of galectin-3 from the nucleus may be mediated by the CRM1 receptor. A candidate nuclear export signal fitting the consensus sequence recognized by CRM1 can be found between residues 240 and 255 of the murine galectin-3 sequence. This sequence was engineered into the pRev(1.4) reporter system, in which candidate sequences can be tested for nuclear export activity in terms of counteracting the nuclear localization signal present in the Rev(1.4) protein. Rev(1.4)-galectin-3(240-255) exhibited nuclear export activity that was sensitive to inhibition by leptomycin B. Site-directed mutagenesis of Leu247 and Ile249 in the galectin-3 nuclear export signal decreased nuclear export activity, consistent with the notion that these two positions correspond to the critical residues identified in the nuclear export signal of the cAMP-dependent protein kinase inhibitor. The nuclear export signal activity was also analyzed in the context of a full-length galectin-3 fusion protein; galectin-3(1-263; L247A) showed more nuclear localization than wild-type, implicating Leu247 as critical to the function of the nuclear export signal. These results indicate that residues 240-255 of the galectin-3 polypeptide contain a leucine-rich nuclear export signal that overlaps with the region (residues 252-258) identified as important for nuclear localization.     

Molecular detection of predation by soil micro-arthropods on nematodes

The relative importance of the factors driving change in the population dynamics of nematodes in the soil is almost completely unknown. Top-down control by micro-arthropod predators may have a significant impact on nematode population dynamics. We report experiments showing that mites and Collembola were capable of reducing nematode numbers in the laboratory and were feeding on a targeted nematode species in the field. A PCR-based approach was developed for the detection of predation on three species of slug- and insect-pathogenic nematodes: Phasmarhabditis hermaphrodita, Heterorhabditis megidis and Steinernema feltiae. The collembolan Folsomia candida and the mesostigmatid mite Stratiolaelaps miles were employed as model predators to calibrate post-ingestion prey DNA detection times. Fragments of cytochrome oxidase I (COI) mtDNA were sequenced and species-specific primers were designed, amplifying 154-, 154- and 203-bp fragments for each of the nematode species. Detection times for nematode DNA within the guts of Collembola were longer than in mites, with half-lives (50% of samples testing positive) of 08.75 h and 05.03 h, respectively. F. candida significantly reduced numbers of the nematode H. megidis, with rates of predation of approximately 0.4 nematode infective juveniles per collembolan per hour over 10 h. Four taxa of field-caught micro-arthropod that had been exposed to the nematode P. hermaphrodita for a period of 12 h were analysed and significant numbers of three taxa tested positive. This is the first application of PCR techniques for the study of nematophagy and the first time these techniques have been used to measure predation on nematodes in the field.     

Compartmentation and flux characteristics of nitrate in spruce

The radiotracer¹³N was used to undertake compartmental analyses for NO ₃ ^– in intact non-mycorrhizal roots ofPicea glauca (Moench) Voss. seedlings. Three compartments were defined, with half-lives of exchange of 2.5 s, 20 s, and 7 min. These were identified as representing surface adsorption, apparent free space, and cytoplasm, respectively. Influx, efflux, and net flux as well as cytoplasmic and apparent-free-space nitrate concentrations were estimated for three different concentration regimes of external nitrate. After exposure to external NO ₃ ^– for 3 d, influx was calculated to be 0.09 mol·g^–1·h^–1 (at 10 M [NO ₃ ^– ]_o), 0.5mol·g^–1·h^–1 (at 100 M [NO _inf3 ^sup– ]_o), and 1.2 mol · g^–1· h^–1 (at 1.5 mM [NO ₃ ^– ]_o). Efflux increased with increasing [NO ₃ ^– ]_o, constituting 4% of influx at 10 M, 6% at 100 M, and 21% at 1.5 mM. Cytoplasmic [NO ₃ ^– ] was estimated to be 0.3 mM at 10 uM [NO ₃ ^– ]_o, 2mM at 100 M [NO ₃ ^– ]_o, and 4mM at 1.5 mM [NO ₃ ^– ]_o, while free-space [NO ₃ ^– ] was 16 M, 173 M, and 2.2 mM, respectively. A series of experiments was carried out to confirm the identity of the compartments resolved by efflux analysis. Pretreatment at high temperature or application of 2-chloro-ethanol, sodium dodecyl sulphate or hydrogen peroxide made it possible to distinguish the metabolic (cytoplasmic) phase from the remaining two (physical) phases. Likewise, varying [Pi] of the medium altered efflux and thereby [NO ₃ ^– ]_cyt, but did not affect [NO ₃ ^– ]_{free space}.Abbreviations and Symbols [NO ₃ ^– ]_cyt cytoplasmic NO ₃ ^– concentration - [NO ₃ ^– ]_{free space} apparent-free-space NO ₃ ^– concentration - [NO ₃ ^– ]_o concentration of NO ₃ ^– in the external solution - NO ₃ ^– flux - _co efflux from the cytoplasm - _oc influx to the cytoplasm - _net net flux - _xylem flux to the xylem - _red/vac combined flux to reduction and the vacuole The research was supported by a Natural Sciences and Engineering Research Council, Canada, grant to Dr. A.D.M. Glass and by a University of British Columbia Graduate Fellowship to Herbert J. Kronzucker. Our thanks go to Dr. M. Adam and Mr. P. Culbert at the particle accelerator facility TRIUMF on the University of British Columbia Campus for providing¹³NO ₃ ^– , Drs. R.D. Guy and S. Silim for providing plant material, and Dr. M.Y. Wang, Mr. J. Mehroke and Mr. P. Poon for assistance in experiments and for helpful discussions.     

The effect of density-dependent treatment and behavior change on the dynamics of HIV transmission

In this work, we propose a model for heterosexual transmission of HIV/AIDS in a population of varying size with an intervention program in which treatment and/or behavior change of the infecteds occur as an increasing function of the density of the infected class in the population. This assumption has socio-economic implications which is important for public health considerations since density-dependent treatment/behavior change may be more cost-saving than a program where treatment/behavior change occurs linearly with respect to the number of infecteds. We will make use of the conservation law of total sexual contacts which enables us to reduce the two-sex model to a simpler one-sex formulation. Analytical results will be given. Unlike a similar model with linear treatment/behavior change in Hsieh (1996) where conditions were obtained for the eradication of disease, we will show that density-dependent treatment/behavior change cannot eradicate the disease if the disease is able to persist without any treatment/behavior change. This work demonstrates the need to further understand how treatment/behavior change occurs in a society with varying population.     

Natural populations of the Closterium ehrenbergii (Desmidiales, Chlorophyta) species complex in Nepal

Several natural populations of the Closterium ehrenbergii Meneghini ex Ralfs species complex were collected in Nepal, in October–December 1982. Water temperature and pH were also recorded. Clonal isolates from these populations were identified to one of four mating groups (H, I, J and M) by test crossing with standard mating-type strains of known mating groups. Groups H and M have smooth walled zygospores, while Groups I and J have scrobiculated zygospore walls. Several undetermined isolates were found in some population samples. In contrast to the previously reported population samples from Nepal, especially from dried soil samples, some of these populations appeared to be rather heavily loaded with mutations that are deleterious to the sexual cycle (i.e. sexual compatibility, zygospore formation and germination). By genetic analysis, a zygote maturation-defective mutation (zym) was detected. One reason for such a heavy genetic load was suggested to be that most population samples had been maintained exclusively by asexual reproduction for a long period in large lakes and nearby ponds, or left-over vegetative populations in paddy fields after other members entered into dormancy through sexual reproduction. The significance of studying such mutations at sexual gene loci is discussed in the light of speciation problems in microalgae.     

A method for validating stochastic models of population viability: a case study of the mountain pygmy-possum (Burramys parvus)

1.  A method of validating stochastic models of population viability is proposed, based on assessing the mean and variance of the predicted population size.
2.  The method is illustrated with a model of the population dynamics of the mountain pygmy-possum ( Burramys parvus Broom 1895), based on annual census data collected from a single population in the Snowy Mountains of New South Wales, Australia between 1986 and 1997. The model incorporates density-dependence in survivorship and recruitment, and demographic and environmental stochasticity.
3.  The model appeared to make reasonable predictions for the three populations that were used for validation, provided the equilibrium population size was estimated accurately. This may require that differences in habitat quality between populations be taken into account.
4.  Following validation, the model was given new parameters using the additional data from the three populations, and the risk of population decline within the next 100 years was assessed. Although populations as small as 15 females are predicted to be relatively safe from extinction caused by stochastic processes, B. parvus appears vulnerable to loss of habitat and reductions in the population growth rate.
5.  The approach used in this paper is one of few attempts to validate a model of population viability using field data, and demonstrates that some aspects of stochastic population models can be tested.     

Genome analysis ofElymus angulatus andE. patagonicus (Poaceae) and their hybrids with N. and S. AmericanHordeum spp.

The results of genome analysis of five hybrids, viz.Elymus patagonicus ×Hordeum procerum, E. patagonicus ×H. tetraploidum, E. angulatus ×H. jubatum, E. angulatus ×H. lechleri, andE. angulatus ×H. parodii, are reported. The genomic constitution ofHordeum tetraploidum andH. jubatum is best given as H₁H₁H₂H₂, ofH. lechleri andH. parodii as H₁H₁H₂H₂H₄H₄, ofH. procerum as H₁H₁H₂H₂H₃H₃, and ofElymus patagonicus andE. angulatus as SSH₁H₁H₂H₂.     

Molecular analysis of Gigaspora (Glomales, Gigasporaceae)

This work presents a cooperative effort to integrate new molecular (isozyme and SSU analyses) characters into the morphological taxonomy of the genus Gigaspora (Glomales). Previous analyses of published Gigaspora SSU sequences indicated the presence of a few polymorphic nucleotides in the region delimited by primers NS71-SSU 1492'. In our study, the SSU of 24 isolates of arbuscular mycorrhizal (AM) fungi from the Gigasporaceae were amplified and the NS71-SSU 1492' region was directly sequenced. The corresponding sequences of four more isolates of AM fungi from Gigasporaceae, already published, were also included in our analyses. Three Gigaspora groups were identified on the basis of a 6 nucleotide-long 'molecular signature': Gigaspora rosea group ( G. rosea + G. albida ), Gigaspora margarita group ( G. margarita + G. decipiens ) and Gigaspora gigantea , which constituted a group by itself. The isozyme profiles (malate dehydrogenase, MDH) of 12 of these 28 isolates, and seven other isolates not sequenced, were compared. The results obtained further supported the grouping of isolates provided by the SSU analysis. Both SSU and MDH analysis indicated that two out of the 35 isolates had been misidentified, which was confirmed when their morphology was reassessed. The use of the Gigaspora intrageneric molecular signature as a quick, unambiguous and objective method to recognize Gigaspora isolates under any (field or laboratory) experimental conditions is suggested.