Evolution of additive and nonadditive genetic variance in development time along a cline in Drosophila serrata

Abstract. Latitudinal clines provide natural systems that may allow the effect of natural selection on the genetic variance to be determined. Ten clinal populations of Drosophila serrata collected from the eastern coast of Australia were used to examine clinal patterns in the trait mean and genetic variance of the life-history trait egg-to-adult development time. Development time significantly lengthened from tropical areas to temperate areas. The additive genetic variance for development time in each population was not associated with latitude but was associated with the population mean development time. Additive genetic variance tended to be larger in populations with more extreme development times and appeared to be consistent with allele frequency change. In contrast, the nonadditive genetic variance was not associated with the population mean but was associated with latitude. Levels of nonadditive genetic variance were greatest in the region of the cline where the gradient in the change in mean was greatest, consistent with Barton's (1999) conjecture that the generation of linkage disequilibrium may become an important component of the genetic variance in systems with a spatially varying optimum.     

Treponema denticola cystalysin catalyzes beta-desulfination of L-cysteine sulfinic acid and beta-decarboxylation of L-aspartate and oxalacetate

Pyridoxal 5'-phosphate-dependent cystalysin from Treponema denticola catalyzes the beta-displacement of the beta-substituent from both L-aspartate and L-cysteine sulfinic acid. The steady-state kinetic parameters for beta-desulfination of L-cysteine sulfinic acid, k(cat) and K(m), are 89+/-7 s(-1) and 49+/-9 mM, respectively, whereas those for beta-decarboxylation of L-aspartate are 0.8+/-0.1 s(-1) and 280+/-70 mM. Moreover, cystalysin in the pyridoxamine 5'-phosphate form has also been found to catalyze beta-decarboxylation of oxalacetate as shown by consumption of oxalacetate and a concomitant production of pyruvate. The k(cat) and K(m) of this reaction are 0.15+/-0.01 s(-1) and 13+/-2 mM, respectively. Possible mechanistic and physiological implications are discussed.     

Steady-state assemblages of phytoplankton in four temperate lakes (NE U.S.A.)

For four temperate lakes (Northeast U.S.A.) we identify periods of persistent phytoplankton assemblages and investigate the ecological conditions that correlate to these persistent assemblages. Periods of persistent assemblages, here considered as steady-state phases, were defined according to equilibrium criteria (two or three coexisting species, contributing to 80% of the standing biomass, for at least 2 weeks) defined by Sommer et al. (1993, Hydrobiologia 249: 1–7). For all four lakes, samples were taken weekly during the ice-free season and phytoplankton attributes (biomass, assemblages, diversity, species richness, change rates) and abiotic variables (temperature, I^* – as light mean in the mixing zone – z_mix, and nutrients) were analysed. Chodikee (CH), an eutrophic and rapidly flushed lake, did not show any persistent phase. The remaining three lakes showed single steady-state phases that occurred at varying times during the ice-free season. Steady-state phases occurred during early stratification in late spring in the stably stratified oligotrophic Mohonk Lake (MO), in the late summer stratification in the meso-eutrophic Stissing Lake (ST), and during spring mixing in Wononscopomuc Lake (WO). MO showed a 3-week period with dominance of F assemblage (Botryococcus braunii, Willea wilhelmii and Eutetramorus planctonicus), characteristic for clear epilimnia, tolerant to low nutrient and sensitive to high turbidity. For three weeks, ST had a stable assemblage with dominance of Lo(Woronichinia sp.), common assemblage in summer epilimnion of mesotrophic lakes and sensitive to prolonged or deep mixing; and P, assemblage able to live in eutrophic epilimnia with mild light and sensitive to stratification and silica depletion. In contrast, the mesotrophic Wononscopomuc Lake (WO) showed persistent assemblages during a 4-week period of spring circulation, when a dinoflagellate (Lo) was co-dominant with Nitzschia acicularis (C). The latter species is characteristic for mesotrophic lakes, tolerant to low light and sensitive to stratification and silica depletion. Both Lo and P assemblages, among seven others, had before been quoted, in literature, as dominant in maturing stages. We could not find consistent statistical differences between the periods classified as steady-state and non-steady-state. However, the data demonstrated that prolonged period of both mixing and stratification can maintain dominant assemblages. Although, historically sensed as opposite mechanisms, both mixing and stratification, if persistent, were observed maintaining dominant assemblages because both scenarios are characterized by environmental constancy.     

Bulla gouldiana period exhibits unique regulation at the mRNA and protein levels

The authors cloned the period (per) gene from the marine mollusk Bulla gouldiana, a well-characterized circadian model system. This allowed them to examine the characteristics of the per gene in a new phylum, and to make comparisons with the conserved PER domains previously characterized in insects and vertebrates. Only one copy of the per gene is present in the Bulla genome, and it is most similar to PER in two insects: the cockroach, Periplaneta americana, and silkmoth, Antheraea pernyi. Comparison with Drosophila PER (dPER) and murine PER 1 (mPER1) sequence reveals that there is greater sequence homology between Bulla PER (bPER) and dPER in the regions of dPER shown to be important to heterodimerization between dPER and Drosophila timeless. Although the structure suggests conservation between dPER and bPER, expression patterns differ. In all cells and tissues examined that are peripheral to the clock neurons in Bulla, bPer mRNA and protein are expressed constitutively in light:dark (LD) cycles. In the identified clock neurons, the basal retinal neurons (BRNs), a rhythm in bPer expression could be detected in LD cycles with a peak at zeitgeber time (ZT) 5 and trough expression at ZT 13. This temporal profile of expression more closely resembles that of mPER1 than that of dPER. bPer rhythms in the BRNs were not detected in continuous darkness. These analyses suggest that clock genes may be uniquely regulated in different circadian systems, but lead to similar control of rhythms at the cellular, tissue, and organismal levels.     

Photoinduced electron transfer and chemical alpha-deoxygenation of D-galactono-1,4-lactone. Synthesis of 2-deoxy-D-lyxo-hexofuranosides

Two simple procedures for the synthesis of 2-deoxy-D-lyxo-hexono-1,4-lactone are described. Reductive cleavage of a 2-O-tosyl derivative of D-galactono-1,4-lactone in the presence of sodium iodide afforded the 2-deoxy derivative. On the other hand, alpha-deoxygenation of D-galactono-1,4-lactone was easily achieved by photochemical electron transfer deoxygenation of HO-2 as the 3-(trifluoromethyl)benzoate. Methyl 2-deoxy-beta-D-lyxo-hexafuranoside ('methyl 2-deoxy-beta-D-galactofuranoside') was synthesized and tested as substrate for exo beta-D-galactofuranosidase from Penicillium fellutanum. The reaction was followed by HPAEC, showing that methyl 2-deoxy-beta-D-galactofuranoside was not hydrolyzed by incubation with the enzyme. Neither the 2-deoxy lactone, nor the 2-deoxy-beta-D-galactofuranoside acted as inhibitors of the reaction with the 4-nitrophenyl beta-D-galactofuranoside. The present and our previous results show that the hydroxyl groups at C-2, C-3 and C-6 of the galactofuranoside are essential for interaction with the exo beta-D-galactofuranosidase.     

Thiol protecting agents and antioxidants inhibit the mitochondrial permeability transition promoted by etoposide: implications in the prevention of etoposide-induced apoptosis

Etoposide (VP-16) is known to promote cell apoptosis either in cancer or in normal cells as a side effect. This fact is preceded by the induction of several mitochondrial events, including increase in Bax/Bcl-2 ratio followed by cytochrome c release and consequent activation of caspase-9 and -3, reduction of ATP levels, depolarization of membrane potential (DeltaPsi) and rupture of the outer membrane. These events are apoptotic factors essentially associated with the induction of the mitochondrial permeability transition (MPT). VP-16 has been shown to stimulate the Ca2+-dependent MPT induction similarly to prooxidants and to promote apoptosis by oxidative stress mechanisms, which is prevented by glutathione (GSH) and N-acetylcysteine (NAC). Therefore, the aim of this work was to study the effects of antioxidants and thiol protecting agents on MPT promoted by VP-16, attempting to identify the underlying mechanisms on VP-16-induced apoptosis. The increased sensitivity of isolated mitochondria to Ca2+-induced swelling, Ca2+ release, depolarization of DeltaPsi and uncoupling of respiration promoted by VP-16, which are prevented by cyclosporine A proving that VP-16 induces the MPT, are also efficiently prevented by ascorbate, the primary reductant of the phenoxyl radicals produced by VP-16. The thiol reagents GSH, dithiothreitol and N-ethylmaleimide, which have been reported to prevent the MPT induction, also protect this event promoted by VP-16. The inhibition of the VP-16-induced MPT by antioxidants agrees with the prevention of etoposide-induced apoptosis by GSH and NAC and suggests the generation of oxidant species as a potential mechanism underlying the MPT that may trigger the release of mitochondrial apoptogenic factors responsible for apoptotic cascade activation.     

In vitro chloroquine resistance modulation study on fresh isolates of Brazilian Plasmodium falciparum: intrinsic antimalarial activity of phenothiazine drugs

Phenothiazine drugs - fluphenazine, chlorpromazine, methotrimeprazine and trifluoperazine - were evaluated as modulating agents against Brazilian chloroquine-resistant fresh isolates of Plasmodium falciparum. Aiming to simulate therapeutic schedules, chloroquine was employed at the concentration used for sensitive falciparum malaria treatment and anti-psychotic therapeutic concentrations of the phenothiazine drugs were adopted in two-fold serial dilutions. The in vitro microtechnique for drug susceptibility was employed. Unlike earlier reported data, the phenothiazine modulating effect was not observed. However, all the drugs demonstrated intrinsic antiplasmodial activity in concentrations lower than those described in the literature. In addition, IC50 estimates have been shown to be inferior to the usual anti-psychotic therapeutic concentrations. Statistical analysis also suggested an increase in the parasitaemia rate or, even, a predominant antiparasitic effect of phenothiazine over chloroquine when used in combination.     

Niemann-Pick type C disease: mutations of NPC1 gene and evidence of abnormal expression of some mutant alleles in fibroblasts

We analyzed Niemann-Pick type C disease 1 (NPC1) gene in 12 patients with Niemann-Pick type C disease by sequencing both cDNA obtained from fibroblasts and genomic DNA. All the patients were compound heterozygotes. We found 15 mutations, eight of which previously unreported. The comparison of cDNA and genomic DNA revealed discrepancies in some subjects. In two unrelated patients carrying the same mutations (P474L and nt 2972del2) only one mutant allele (P474L), was expressed in fibroblasts. The mRNA corresponding to the other allele was not detected even in cells incubated with cycloheximide. The promoter variants (-1026T/G and -1186T/C or -238 C/G), found to be in linkage with 2972del2 allele do not explain the lack of expression of this allele, as they were also found in control subjects. In another patient, (N1156S/Q922X) the N1156S allele was expressed in fibroblasts while the expression of the other allele was hardly detectable. In a fourth patient cDNA analysis revealed a point mutation in exon 20 (P1007A) and a 56 nt deletion in exon 22 leading to a frameshift and a premature stop codon. The first mutation was confirmed in genomic DNA; the second turned out to be a T-->G transversion in exon 22, predicted to cause a missense mutation (V1141G). In fact, this transversion generates a donor splice site in exon 22, which causes an abnormal pre-mRNA splicing leading to a partial deletion of this exon. In some NPC patients, therefore, the comparison between cDNA and genomic DNA may reveal an unexpected expression of some mutant alleles of NPC1 gene.