Laser induced white emission generated by infrared excitation from Eu3+:Sr2CeO4 nanocrystals.

The laser induced white emission (LIWE) was observed from Eu3+:Sr2CeO4 nanocrystals. The samples were obtained in form of powders by the modified sol-gel route. The structure and morphology of the phosphors were investigated by powder X-ray diffraction and transmission electron microscope techniques. The intense LIWE occurred under reduced pressure and focused beam of near infrared laser excitation. The power and pressure dependencies exhibit evident threshold character typical for the avalanche effect. The photoconductivity of the Eu3+:Sr2CeO4 nanocrystals measured as a function of different powers of excitation source was analyzed.

Broadband anti-Stokes white emission of Sr2CeO4 nanocrystals induced by laser irradiation.

The laser induced white emission (LIWE) from Sr2CeO4 nanocrystals upon irradiation with a focused IR laser beam was investigated. It was observed to be a threshold phenomenon with its intensity increasing exponentially with the excitation power density. This process was investigated under double laser beam simultaneous excitation in the UV range leading to Stokes emission in the visible range and in the IR range leading to anti-Stokes LIWE. With increasing LIWE intensity, the Stokes emission intensity strongly decreased. The LIWE is accompanied by efficient photocurrent generation depending on laser excitation density followed by multiphoton absorption and ionization processes. Photoimpedance measurements showed a sharp increase of the dielectric constant by several orders of magnitude in the Sr2CeO4 nanocrystals during the LIWE process demonstrating a metallic-like behaviour. The mechanisms of LIWE include multiphoton absorption and ionization that lead to the creation of a coupled pair of Ce3+ and Ce4+ ions that allow for the intervalence charge transfer (IVCT) emission transitions in the white light range. A strong decrease of absorption band intensity of Sr2CeO4 with increasing LIWE intensity confirms the creation of (Ce3+, Ce4+) pairs.

Correlations between purebred and crossbred body weight traits in Limousin and Limousin-Angus populations.

The purpose of this study was to estimate correlations between purebred and F1 crossbred performance to verify the appropriateness of current models used in multibreed selection. Records on birth weight (WB) and weaning weight (WW) from purebred Limousins (LIM) and Limousin × Angus progeny (F1) were used to estimate genetic parameters using a multiple-trait (purebred and F1 weights were different traits) approach. For WB, there were 148,647 records for LIM and 17,981 for F1, and for WW, there were 81,585 records for LIM and 21,778 for F1. The fixed effect in models for LIM and F1 animals was contemporary group. Random effects for LIM animals were direct genetic, maternal genetic, and maternal permanent environment effects. Random effects for F1 were sire and dam. The pedigree for Angus dams used for crossing was unavailable and therefore these dams were assumed unrelated. The direct h2 estimates (SE) for purebred animals were 0.41 (0.05) and 0.24 (0.02) for WB and WW, respectively. For F1, the same estimates were 0.22 (0.09) and 0.32 (0.05). Genetic correlations estimates between purebreds and crossbreds were 0.84 (0.07) and 0.64 (0.18) for WB and WW, respectively. The genetic correlation for WW estimated in this study suggests that F1 and purebred information for this trait should not be treated, genetically, as the same trait due to different genetic effects molding it. However, the genetic correlation for WB was much higher, indicating that this trait in purebreds and F1 is essentially the same trait.

Heritability of individual egg hatching success versus hen hatchability in layers.

The purpose of the study was to test the genetics of individual egg hatchability. Hatching outcome (0,1) of each of the eggs (126,430) laid during hatching seasons of 5 generations of a Rhode Island White population was analyzed with models that attributed the direct additive effect either to an egg or to a hen. A Gibbs sampling procedure, accounting for dichotomous nature of the trait, was employed for variance component estimation. The egg/embryo direct additive component was negligibly small (h(2) = 0.007) from the point of view of the accuracy of the hatchability proof. The hen direct additive component, though more than 12 times higher (h(2) = 0.087) than that of the egg, was still more than 2 times smaller than the component because of her permanent environment (p(2) = 0.221). More accurate definition of hens' environmental needs may prove to be more effective for hatching outcome improvement than increasing the accuracy of the reproduction proof, because current selection has to be performed before the individual hatchability record is known.

Genotype by environment interaction for growth due to altitude in United States Angus cattle.

The objectives of this study were to determine if sires perform consistently across altitude and to quantify the genetic relationship between growth and survival at differing altitudes. Data from the American Angus Association included weaning weight (WW) adjusted to 205 (n = 77,771) and yearling weight adjusted to 365 (n = 39,450) d of age from 77,771 purebred Angus cattle born in Colorado between 1972 and 2007. Postweaning gain (PWG) was calculated by subtracting adjusted WW from adjusted yearling weight. Altitude was assigned to each record based upon the zip code of each herd in the database. Records for WW and PWG were each split into 2 traits measured at low and high altitude, with the records from medium altitude removed from the data due to inconsistencies between growth performance and apparent culling rate. A binary trait, survival (SV), was defined to account for censored records at yearling for each altitude. It was assumed that, at high altitude, individuals missing a yearling weight either died or required relocation to a lower altitude predominantly due to brisket disease, a condition common at high altitude. Model 1 considered each WW and PWG measured at 2 altitudes as separate traits. Model 2 treated PWG and SV measured as separate traits due to altitude. Models included the effects of weaning contemporary group, age of dam, animal additive genetic effects, and residual. Maternal genetic and maternal permanent environmental effects were included for WW. Heritability estimates for WW in Model 1 were 0.28 and 0.26 and for PWG were 0.26 and 0.19 with greater values in low altitude. Genetic correlations between growth traits measured at different altitude were moderate in magnitude: 0.74 for WW and 0.76 for PWG and indicate possibility of reranking of sires across altitude. Maternal genetic correlation between WW at varying altitude of 0.75 also indicates these may be different traits. In Model 2, heritabilities were 0.14 and 0.27 for PWG and 0.36 and 0.47 for SV. Genetic correlation between PWG measured at low and high altitude was 0.68. Favorable genetic correlations were estimated between SV and PWG within and between altitudes, suggesting that calves with genetics for increased growth from weaning to yearling also have increased genetic potential for SV. Genetic evaluations of PWG in different altitudes should consider preselection of the data, by using a censoring trait, like survivability to yearling.

Genotype by region and season interactions on weaning weight in United States Angus cattle.

The objective of this study was to determine if weaning weight performance is genetically consistent across different environments in the United States. The American Angus Association provided weight and pedigree data. Weaning weights observed in the Southeast (SoE) and Northwest (NW) were the focus of this study, as these regions are perceived as opposite extremes in climate. The 2 most represented calving seasons in each region were fall and winter in the SoE and winter and spring in the NW. The original data were edited to remove weaning weight records outside of 3 SD from the respective region-season mean, contemporary groups smaller than 20, and single-sire contemporary groups. The final dataset included 884,465 weaning weight records with 64,907 from fall-born calves in the SoE, 74,820 from winter-born calves in the SoE, 346,724 from winter-born calves in the NW and 398,014 from spring-born calves in the NW. Weaning weights of calves born in different region-season classes adjusted to 205 d of age were considered different but genetically correlated traits in a multivariate analysis. The sole fixed effect was weaning contemporary group and random effects included direct, maternal, maternal permanent environment, and a residual. Direct heritability estimates differed little across environments: 0.31 and 0.35 for weight in fall- and winter-born calves in the SoE, and 0.29 and 0.32 for winter- and spring-born calves in NW. Maternal heritability estimates ranged from 0.12 in the NW to 0.16 the SoE. Genetic correlations spanned from 0.69 to 0.93 among direct effects and from 0.65 to 0.95 among maternal effects. All heritability estimates had small (0.01 to 0.04) SE. The most distinct environments appeared to be winter in SoE and spring in NW (correlations of 0.69 and 0.65 for the direct and maternal effects). Different choices of sires for different environments might be justified to achieve the growth performance expected.

Estimation of genetic parameters for mature weight in Angus cattle.

The aim of this study was to estimate genetic parameters for BW of Angus cattle up to 5 yr of age and to discuss options for including mature weight (MW) in their genetic evaluation. Data were obtained from the American Angus Association. Only records from herds with at least 500 animals and with >10% of animals with BW at ≥ 2 yr of age were considered. Traits were weaning weight (WW, n = 81,525), yearling weight (YW, n = 62,721), and BW measured from 2 to 5 yr of age (MW2, n = 15,927; MW3, n = 12,404; MW4, n = 9,805; MW5, n = 7,546). Genetic parameters were estimated using an AIREML algorithm with a multiple-trait animal model. Fixed effects were contemporary group and departure of the actual age from standard age (205, 365, 730, 1,095, 1,460, and 1,825 d of age for WW, YW, MW2, MW3, MW4, and MW5, respectively). Random effects were animal direct additive genetic, maternal additive genetic, maternal permanent environment, and residual. Estimates of direct genetic variances (kg(2)) were 298 ± 71.8, 563 ± 15.1, 925 ± 52.1, 1,221 ± 65.8, 1,406 ± 80.4, and 1,402 ± 66.9; maternal genetic variances were 167 ± 4.8, 153 ± 6.1, 123 ± 9.1, 136 ± 12.25, 167 ± 18.0, and 110 ± 14.0; maternal permanent environment variances were 124 ± 2.9, 120 ± 4.3, 61 ± 7.5, 69 ± 11.9, 103 ± 15.9, and 134 ± 35.2; and residual variances were 258 ± 3.8, 608 ± 8.6, 829 ± 34.2, 1,016 ± 38.8, 1,017 ± 52.1, and 1,202 ± 63.22 for WW, YW, MW2, MW3, MW4, and MW5, respectively. The direct genetic correlation between WW and YW was 0.84 ± 0.14 and between WW and MW ranged from 0.66 ± 0.06 (WW and MW4) to 0.72 ± 0.11 (WW and MW2). Direct genetic correlations ranged from 0.77 ± 0.08 (YW and MW5) to 0.85 ± 0.07 (YW and MW2) between YW and MW, and they were ≥ 0.95 among MW2, MW3, MW4, and MW5. Maternal genetic correlations between WW and YW and MW ranged from 0.52 ± 0.05 (WW and MW4) to 0.95 ± 0.07 (WW and YW), and among MW they ranged from 0.54 ± 0.14 (MW4 and MW5) to 0.94 ± 0.07 (MW2 and MW3). Genetic correlations suggest that a genetic evaluation for MW may be MW2-based and that including BW from older ages could be accomplished by adjusting records to the scale of MW2.

Studies on changes of estimated breeding values of U.S. Holstein bulls for final score from the first to second crop of daughters.

The purpose of this study was to find ways of reducing changes of sire predicted transmitting ability for type's final scores (PTATs) from the first to second crop of daughters. The PTATs were estimated from two datasets: D01 (scores recorded up to 2001) and D05 (scores recorded up to 2005). The PTAT changes were calculated as the difference between the evaluations based on D01 and D05. The PTATs were adjusted to a common genetic base of all evaluated cows born in 1995. The single-trait (ST) animal model included the fixed effects of the herd-year-season-classifier, age by year group at classification, stage of lactation at classification, registry status of animals, and additive genetic and permanent environment random effects. Unknown parent groups (UPGs) were defined based on every other birth year starting from 1972. Modifications to the ST model included the usage of a single record per cow, separate UPGs for first and second crop daughters, separate UPGs for sires and dams, and deepened pedigrees for dams with missing phenotypic records. Also, the multiple-trait (MT) model treated records of registered and grade cows as correlated traits. The mean PTAT change, for all of the sires, was close to zero in all of the models analyzed. The estimated mean PTAT change for 145 sires with 40 to 100 first crop and ≥ 200 second crop daughters was -0.33, -0.20, -0.13, -0.28, and -0.12 with ST, only first records, only last records, updated pedigrees, and allowing separate parent groups (PGs) for sires and dams after updating the pedigrees, respectively. The percentages of sires showing PTAT decline were reduced from 74.5 (with ST) to 57.3 by using only the last records of cows, and to 56.4 by allowing separate UPGs for sires and dams after updating the pedigrees. Though updating of the pedigrees alone was not effective, separate UPGs for sires together with additional pedigree was helpful in reducing the bias.

Use of serial pig body weights for genetic evaluation of daily gain.

This study examined the utility of serial weights from FIRE (Feed Intake Recording Equipment, Osborne Industries, Inc., Osborne, KS, USA) stations for an analysis of daily gain. Data included 884 132 body weight records from 3888 purebred Duroc pigs. Pigs entered the feeder station at age 77-149 days and left at age 95-184 days. A substantial number of records were abnormal, showing body weight close to 0 or up to twice the average weight. Plots of body weights for some animals indicated two parallel growth curves. Initial editing used a robust regression, which was a two-step procedure. In the first step, a quadratic growth curve was estimated assuming small or 0 weights for points far away from the curve; the process is iterative. In the second step, weights more than 1.5 SD from the estimated growth curve were treated as outliers. The retained body weight records (607,597) were averaged to create average daily weight (170,443) and then used to calculate daily gains (152,636). Additional editing steps included retaining only animals with >or=50 body weight records and SD of the daily gain

Cell ATP level of Saccharomyces cerevisiae sensitively responds to culture growth and drug-inflicted variations in membrane integrity and PDR pump activity.

Cellular ATP level in Saccharomyces cerevisiae was measured during culture growth of strain US50-18C overproducing all major PDR pumps and its isogenic mutants variously deleted in these pumps. It was found to be inversely proportional to the intensity of cell metabolism during different growth phases and to the activity of PDR pumps, which are thus among major ATP consumers in the cells. The ATP level was increased when membrane integrity was affected by 0.5% butanol, and further increased by compound 23.1, a semisynthetic phenol lipid derivative that acts as inhibitor of Pdr5p and Snq2p pumps. The magnitude of increase in cell ATP caused by inhibition of Pdr5p pump by compound 23.1 and the Pdr5p pump inhibitor FK506 used for comparison reflects the activity and hence the energy demand of the pump. The rise in cell ATP caused by different PDR pump inhibitors can be thus used as an indicator of pump activity and the potency of the inhibitor.

Microsatellite markers may be ineffective in selection of laying hens for polygenic production traits.

Previous research on mapping QTL in a reference family of laying hens indicated that 5 microsatellite loci (MCW0133, MCW0170, MCW0114, MCW0139, and LEI0074) were significantly associated with genome regions affecting shell strength as well as egg and yolk weights. The aim of our investigation was to verify if those markers could be useful in selection of laying hens. The study involved 2 breeds of randomly segregating populations: Rhode Island Reds selected divergently and Green-legged Partridgenous chickens selected upwardly, over 4 generations, for the mentioned egg quality traits. The influence of marker genotype on bird performance was assessed through the prediction of breeding values using a model that distinguished the marker effect from that of the polygenic effect and by comparing breeding values between different genotypes at given marker loci. The effects of the linked QTL regions appeared too small to significantly differentiate the outcomes of classifications fitting or not fitting the marker genotype. Comparison of breeding values between microsatellite genotypes for laying and egg traits revealed that antagonistic pleiotropic effects exist between these 2 groups of traits, adding to the difficulty of accounting for marker genotypes in the selection of laying hens.

Genetic characteristics of the ostrich population using molecular methods.

A genetic analysis was performed on Polish ostriches from the 3 principal ostrich breeds: red-, blue-, and black-necks. The analysis was based on 2 molecular methods: DNA fingerprinting and microsatellites. The DNA fingerprinting patterns were obtained using the restriction enzyme HinfI and Jeffrey's 33.15 probe. The second method consisted of a PCR procedure, for which 5 VIAS-OS primers specific to the ostrich were used. The PCR products were separated on polyacrylamide gel using ALFexpress (Authomated Laser Fluorescent DNA Sequencer). The study aimed at assessing the genetic variability within and among the 3 ostrich breeds as well as evaluating the genetic distance between them, and represents the first report on the genetic characteristics of the ostrich breeds. The results obtained by both methods showed considerable compatibility, especially with regard to the relationship among the breeds analyzed. The diversity within breeds, obtained on the basis of the DNA fingerprinting analysis, proved to be low. Among the ostrich populations analyzed, the highest variability potential was observed for black-necked ostriches (the mean diversity of patterns amounted to 29.04%, whereas the mean heterozygosity was 0.30) and the lowest was observed for the red-necks. The largest genetic similarity was recorded between red- and blue-necked ostriches, but the greatest genetic distance was between the red- and black-necks. This means that the use of birds of those breeds in crosses should result in the highest heterotic effect. Both of these methods measured the genetic distance between the analyzed ostrich breeds that was expected from the geographic origin of these birds. The results obtained in the present study showed that both analytic methods used can be successfully applied when elaborating on the genetic characteristics of the ostrich.

Regeneration of flax ( Linum usitatissimum L.) plants from anther culture and somatic tissue with increased resistance to Fusarium oxysporum.

The aim of this study was to establish a protocol for the efficient production of flax plants of microspore origin. The results were compared to those obtained for plants regenerated from somatic explants from hypocotyls, cotyledons, leaves, stems and roots. All the plants obtained during the experiments were regenerated from callus that was grown for periods from a few weeks to a few months before the regeneration was achieved. Anther cultures were less effective in plant regeneration than somatic cell cultures. However, regenerants derived from anther cells showed valuable breeding features, including increased resistance to fungal wilt. The age of the donor plants and the season they grew in had a noticeable effect on their anther callusing and subsequent plant regeneration. Low temperature had a negative effect and dark pre-treatment a positive effect on callusing and plant regeneration. Different media were most effective for callus induction, shoot induction and rooting. For callus induction two carbon sources (2.5% sucrose and 2.5% glucose) were most effective; for shoots, only sucrose at lower concentration (2%) was effective. Rooting was most efficient in 1% sucrose and reduced (50%) mineral concentration in the medium. It was found that the length of in vitro cultivation significantly increases the ploidy and affects such features as regenerant morphological characteristics, petal colour, and resistance to Fusarium oxysporum-induced fungal wilt. The established plant regeneration system provides a basis for the creation of transgenic flax.

Effect of antioxidants on Saccharomyces cerevisiae mutants deficient in superoxide dismutases.

S. cerevisiae strain delta sodl lacking Cu,Zn-superoxide dismutase and delta sodl delta sod2 mutant lacking both Cu,Zn-SOD and Mn-superoxide dismutase displayed strongly reduced aerobic growth on glucose, glycerol and lactate; delta sod2 deletion had no effect on aerobic growth on glucose and largely precluded growth on glycerol and lactate. The oxygen-induced growth defects and their alleviation by antioxidants depended on growth conditions, in particular on oxygen supply to cells. Under strong aeration, vitamins A and E had a low effect, 100 mumol/L quercetin alleviated the growth defects of all three mutants while beta-carotene had no growth-restoring effect. The superoxide producer paraquat inhibited the aerobic growth of all three mutants in a concentration-dependent manner. Low concentrations of antioxidants had no effect on paraquat toxicity while higher concentrations supported the toxic effect of the agent.

Functional dissection of the ParB homologue (KorB) from IncP-1 plasmid RK2.

Active partitioning of low-copy number plasmids requires two proteins belonging to the ParA and ParB families and a cis-acting site which ParB acts upon. Active separation of clusters of plasmid molecules to the defined locations in the cell before cell division ensures stable inheritance of the plasmids. The central control operon of IncP-1 plasmids codes for regulatory proteins involved in the global transcriptional control of operons for vegetative replication, stable maintenance and conjugative transfer. Two of these proteins, IncC and KorB, also play a role in active partitioning, as the ParA and ParB homologues, respectively. Here we describe mapping the regions in KorB responsible for four of its different functions: dimerisation, DNA binding, repression of transcription and interaction with IncC. For DNA binding, amino acids E151 to T218 are essential, while repression depends not only on DNA binding but, additionally, on the adjacent region amino acids T218 to R255. The C-terminus of KorB is the main dimerisation domain but a secondary oligomerisation region is located centrally in the region from amino acid I174 to T218. Using three different methods (potentiation of transcriptional repression, potentiation of DNA binding and activation in the yeast two-hybrid system) we identify this region as also responsible for interactions with IncC. This IncC-KorB contact differs in location from the ParA-ParB/SopA-SopB interactions in P1/F but is similar to these systems in lying close to a masked oligomerisation determinant.

The antioxidant activity of BHT and new phenolic compounds PYA and PPA measured by chemiluminescence.

The antioxidative properties of two series of new phenolic, amphiphilic compounds were evaluated using the chemiluminescence (CL) method. 2,2'-Azobis (2-amidinopropane) dihydrochloride (AAPH) was used as a source of free radicals, to obtain high and prolonged CL. Three different kinds of buffers (organic and inorganic) were tested. The CL level varied only slightly depending on the buffer but increased significantly with the pH. Twelve newly synthesised compounds were compared with butylated hydroxytoluene (BHT), a commercially used antioxidant. The new antioxidants included two classes of quaternary ammonium salts with a phenol substituent functioning as an antioxidant. The salts were synthesised by quaternarization of pyrrolidine ethyl esters of dihydrocinnamic acid by n-alkoxymethyl bromides (PYA-n) or quaternarization of 2-dimethylaminoethyl esters by n-alkyl bromides (PPA-n). All the tested compounds quenched CL proportionally to their concentrations. In our experimental conditions 8.5 microM BHT quenched 50% of the CL. The PYA and PPA compounds had IC50 two to six times lower than BHT. CL inhibition was proportional to the pH for all antioxidants. The relationships between the structure and activity of the tested compounds are discussed.

Non-additive genetic effects in animal selection.

Genetic evaluation of purebred farm animals has been carried out for about half a century, employing additive approximation to describe the genetic background. An evaluated animal has been attributed a single breeding value for each trait of the breeding goal. The predicted additive genetic value of an animal equals the average breeding value of its parents. Although the selection based on the additive approach has proved successful, there still is a possibility of increasing the reliability of the breeding value estimation by accounting for non-additive genetic effects of dominance and epistasis, disregarded in the additive model. In the non-additive model, the expected quality of the progeny equals the average of the parents plus an effect resulting from the interaction between the parents. In this case, the evaluated animal may have as many breeding values as there are possible candidates to mate to, for each trait. The dominance and epistatic effects have already been accounted for in selecting animals or populations for some crossbreeding plans (combining ability, heterosis, and recombination loss). Also, using crossbreds for the sake of the breeding value estimation of purebred animals requires removing the non-additive effects from the crossbred performance and distributing the additive component between the purebreds. Combining ability is more and more discussed as a factor for matings within breed to produce terminal progeny.

In vivo evaluation of the context sequence of the translation initiation codon in plants.

Statistical analysis of the AUG initiation codon context in several plant organisms identified a nucleotide preference in some positions around the AUG. Sixteen AUG contexts were studied using transient expression in tobacco, maize and Norway spruce. Besides the importance of A or G at position -3, we revealed the role of positions -2, -1 for which AA or CC were found to be the best for tobacco and maize, respectively. GC (positions +4, +5) were also found to be important in both tobacco and maize. Finally, we identified a variation in context efficiency according to cell type, since A was better than G at position -3 in tobacco leaf protoplasts, while both nucleotides were equally efficient in tobacco suspension cells.

Spontaneous and radical-induced plasma membrane lipid peroxidation in differently oxidant-sensitive yeast species and its suppression by antioxidants.

Formation of thiobarbituric acid-reactive substances (TBRS; nmol/mg lipids) indicative of lipid peroxidation was measured in whole cells and in isolated plasma membrane lipids from three yeast species differing in oxidant sensitivity (Schizosaccharomyces pombe, Saccharomyces cerevisiae and Rhodotorula glutinis) after exposure to the Fenton reagent, FeII, H2O2, tert-butyl hydroperoxide (TBHP) and azo compounds (AAPH, ACHN). In whole cells, spontaneous TBRS formation rose in the sequence S. pombe < S. cerevisiae < R. glutinis (1:approximately 5:approximately 7). Oxidants increased the TBRS production 13-18 fold in the sequence FeII approximately TBHP > AAPH approximately ACHN approximately Fe-Fenton > H2O2. This increase need not be solely due to increased lipid peroxidation. In isolated plasma membrane lipids from all three species, the spontaneous TBRS production referred to 1 mg lipids was 9-13-fold higher than in whole cells. In S. pombe lipids, only TBHP increased the TBRS production. In lipids from S. cerevisiae and R. glutinis, all added oxidants increased the spontaneous TBRS production 2-3 times in the sequence TBHP > ACHN > AAPH > FeII > Fe-Fenton > H2O2. Oxidant-induced TBRS production in both whole cells and isolated membrane lipids was partially suppressed by the lipid peroxidation inhibitors 2,6-di-tert-butyl-4-methylphenol ("butylated hydroxytoluene"; BHT) and the newly synthesized PYA12 compound. Both agents were more effective in isolated lipids than in whole cells and against OH.-producing than against ROO.- or RO.-producing oxidants. Yeast membrane lipids, which are generally poor in polyunsaturated fatty acids, are thus subject to perceptible lipid peroxidation.

Signs of translational regulation within the transcript leader of a plant plasma membrane H(+)-ATPase gene.

Transcripts of most plant plasma membrane H(+)-ATPase genes possess a leader (5' untranslated region) that is unusually long and that contains a short upstream open reading frame (uORF), two features which suggest post-transcriptional regulation. To investigate the putative role of the transcript leader, we have placed the leader of pma3, one of the Nicotiana plumbaginifolia H(+)-ATPase genes, between the CaMV 35S promoter and the sequence coding for the beta-glucuronidase (GUS) reporter gene. Transient expression of this chimeric gene and of derived mutants was analysed in electroporated tobacco protoplasts. The whole leader had a positive effect on translation, since deletion of most of its sequence reduced GUS activity. Suppression of the uORF by point mutation of its initiating AUG increased GUS activity by about 55%. Analysis of various deletions and mutations suggested that the uORF is translated by at least two-thirds of scanning ribosomes, half of which subsequently reinitiate downstream translation under our experimental conditions. Reinitiation did not depend on the nucleotide sequence of the uORF, nor on that separating the uORF and the main open reading frame. We conclude that the pma3 transcript possesses features of translational regulation, whose mode of functioning has yet to be discovered.