Quantum random number generator based on photonic emission in semiconductors.

We report upon the realization of a novel fast nondeterministic random number generator whose randomness relies on the intrinsic randomness of the quantum physical processes of photonic emission in semiconductors and subsequent detection by the photoelectric effect. Timing information of detected photons is used to generate binary random digits-bits. The bit extraction method based on the restartable clock method theoretically eliminates both bias and autocorrelation while reaching efficiency of almost 0.5 bits per random event. A prototype has been built and statistically tested.

Extended life-span conferred by cotransporter gene mutations in Drosophila.

Aging is genetically determined and environmentally modulated. In a study of longevity in the adult fruit fly, Drosophila melanogaster, we found that five independent P-element insertional mutations in a single gene resulted in a near doubling of the average adult life-span without a decline in fertility or physical activity. Sequence analysis revealed that the product of this gene, named Indy (for I'm not dead yet), is most closely related to a mammalian sodium dicarboxylate cotransporter-a membrane protein that transports Krebs cycle intermediates. Indy was most abundantly expressed in the fat body, midgut, and oenocytes: the principal sites of intermediary metabolism in the fly. Excision of the P element resulted in a reversion to normal life-span. These mutations may create a metabolic state that mimics caloric restriction, which has been shown to extend life-span.

Cu, Zn superoxide dismutase deficiency accelerates the time course of an age-related marker in Drosophila melanogaster.

In the oxidative stress hypothesis of aging the random accumulation of oxidative damage over time is postulated to cause aging. The pace at which oxidative damage accrues determines the rate of aging, but it is less clear how the accumulation of random damage could cause the stereotypic pattern of aging. It has been proposed that oxidative damage induces changes in gene expression, translating a random input of damage into a patterned output. In support of this we show that in adult Drosophila melanogaster, with a deficiency in the anti-oxidant enzyme Cu, Zn superoxide dismutase (Sod), an increase in oxidative stress, and a shortened life span, there is acceleration in the normal age-related temporal pattern of wingless gene expression. The acceleration in the temporal pattern of wingless gene expression is proportional to the shortened life span suggesting that the shortened life span of Sod deficient animals is due, not to an abnormal pathological process, but to an increase in the rate of aging.

Regulation of gene expression is preserved in aging Drosophila melanogaster.

Aging, and the deterioration of biological performance that characterizes it, are routinely assumed to be due to a progressive global loss of homeostasis and a general increase in dysregulation [1-4] . We tested this hypothesis directly by measuring age-specific variability in gene expression. Analysis of the transcriptional activity of six genes in various inbred lines of Drosophila melanogaster unexpectedly failed to show an increase in variability among individuals as they age and die. Although regulation of gene expression is a central feature of life, a global decline in the control of gene expression does not appear to be either a cause or a consequence of the process of aging.

Patterns of expression of Hoxa-11 in micromass cultures of chick limb mesenchyme from various stages suggest a role for Hoxa-11 in the specification of the zeugopod.

During chick limb development the expression of Hoxa-11 specifically associated with the prospective zeugopod (radius + ulna) region. A series of micromass cultures were performed using mesenchyme from whole stage 18/19 or 20/21 limbs or mesenchyme from the tips of stage 22/23, 23/24 or 25 limbs. In such cultures, limb mesenchyme undergoes chondrogenesis paralleling its fate in vivo. Hoxa-11 and collagen type II mRNA were examined by dot blot and in situ analysis at different time points. The highest level of Hoxa-11 mRNA was found in micromass cultures of the tips of stage 22/23 and 23/24 limbs at 48 hours. Between stages 22 and 24, the determining events occur which subsequently lead to the formation of the zeugopod by the mesenchyme present in the tips at these stages. This correlation further supports a possible role of Hoxa-11 in the specification of the zeugopod.

Drosophila drop-dead mutations accelerate the time course of age-related markers.

Mutations of the drop-dead gene in Drosophila melanogaster lead to striking early death of the adult animal. At different times, after emergence from the pupa, individual flies begin to stagger and, shortly thereafter, die. Anatomical examination reveals gross neuropathological lesions in the brain. The life span of flies mutant for the drop-dead gene is four to five times shorter than for normal adults. That raises the question whether loss of the normal gene product might set into motion a series of events typical of the normal aging process. We used molecular markers, the expression patterns of which, in normal flies, change with age in a manner that correlates with life span. In the drop-dead mutant, there is an acceleration in the temporal pattern of expression of these age-related markers.

Spatial and temporal pattern of expression of the wingless and engrailed genes in the adult antenna is regulated by age-dependent mechanisms.

The spatial and temporal pattern of expression of enhancer trap lines reporting on the wingless (wg) and engrailed (en) genes was characterized in the adult antenna of Drosophila melanogaster. The time courses of expression seen for wg and en, although different from each other, reveal a complex well-controlled pattern of temporal expression, providing evidence that regulatory mechanisms are preserved throughout the life span of the adult fly. Altering the life span demonstrates that the temporal patterns of expression of both wg and en are linked to life span. These studies suggest that the expression of wg and en in the adult antenna is controlled by age-dependent mechanisms.

Timing of expression of a gene in the adult Drosophila is regulated by mechanisms independent of temperature and metabolic rate.

The examination of beta-galactosidase (beta-gal) expression in the third segment of the antenna of the 2216 enhancer trap line in Drosophila melanogaster reveals two distinct spatial and temporal regulatory patterns of expression during adult life. Type 1 expression is characterized by a decline in the level of beta-gal expression with increasing age. Starting from a maximal level of expression at the time of adult emergence, there is a decrease in the number of cells that express beta-gal so that by 40-50 days of adult life few cells express beta-gal. Varying the ambient temperature and using hyperactivity mutants (Hyperkinetic, Shaker) demonstrates that the rate of this decline is independent of temperature and metabolic rate. Type II expression is distinctly different in spatial distribution and temporal regulation from the first pattern. Type II expression is restricted in the antenna to a small (< 20-30) set of cells whose level of expression changes in a periodic manner with time. The regulation of this periodicity appears to be linked to ambient temperature.

Regulation of gene expression is linked to life span in adult Drosophila.

Examination of gene expression and aging in adult Drosophila reveals that the expression of some genes is regulated by age-dependent mechanisms. Genetic mutations, Hyperkinetic and Shaker, which are known to shorten life span through an acceleration of the aging process, were used to study the expression of an enhancer trap marked gene. The temporal pattern of expression for such a marked gene shows scaling with respect to life span; it is altered in direct proportion to the life expectancy of the adult animal. This demonstrates that expression of this gene is controlled through mechanisms coupled to physiologic as opposed to chronologic age. Results provide direct evidence for linkage between the regulation of gene expression and life span and establish a model system for the genetic analysis of aging.

Changes in gene expression during post-eclosional development in the olfactory system of Drosophila melanogaster.

We have found that the expression of some genes in Drosophila melanogaster changes during the life of the adult fly. These changes can be illustrated by the use of enhancer trap lines which mark the expression of particular genes in the adult fly. Although the fly is considered able to perform most necessary adult functions within the first 72 h after eclosion from the pupal case, we find changes in expression over the first 10 days of life in the antennae of several of the genes we have examined. Some genes change by increasing from an initially low level of expression of the marked gene, while other lines, which we have termed 'late-onset' genes, show no expression of the marked gene until 4-5 days following eclosion. In contrast, some genes decrease their expression during the first 10 days of life. The changes in gene expression seen over the first 10 days of the fly's adult life provides molecular evidence of the many maturational changes occurring during the early life of the adult fly.

Temporal patterns of gene expression in the antenna of the adult Drosophila melanogaster.

The time course of gene expression in the adult fruit fly has been partially characterized by using enhancer trap and reporter gene constructs that mark 49 different genes. The relative intensity of the reporter protein in individual cells of the antennae was measured as a function of adult age. Most genes showed a graduated expression, and the intensity of expression had a reproducible and characteristic time course. Different genes displayed different temporal patterns of expression and more often than not the pattern of expression was complex. We found a number of genes having patterns that scaled with life span. In these cases the intensity of gene expression was found to be invariant with respect to biological time, when expressed as a fraction of the life span of the line. The scaling was observed even when life span was varied as much as threefold. Such scaling serves to (1) further demonstrate that deterministic mechanisms such as gene regulation act to generate the temporal patterns of expression seen during adult life, (2) indicate that control of these regulatory mechanisms is linked to life span, and (3) suggest mechanisms by which this control is accomplished. We have concluded that gene expression in the adult fly is often regulated in a fashion that allows for graduated expression over time, and that the regulation itself is changing throughout adult life according to some prescribed program or algorithm.

The chicken homeobox gene Hoxd-11 encodes two alternatively spliced RNA species.

Two forms of cDNA clones corresponding to Hoxd-11 were obtained by screening a chicken limb bud cDNA library. Comparison of the two cDNA sequences with the mouse cDNA and genomic sequence, shows that the shorter cDNA is missing 3 nucleotides, which encode an alanine residue, at the junction between exons 1 and 2. Analysis of the chicken genomic sequence shows two tandem CAG repeats at the intron 1-exon 2 junction, both of which could serve as splice acceptor sequences. Thus we hypothesize that the two spliced forms result from alternative use of two tandem splice acceptor sequences. Using reverse transcription polymerase chain reaction we examined expression of the two forms in 3 to 14-day chick embryos. All samples showed both species of RNA with approximately 2 times more of the larger from compared with the smaller form.

The differential diagnostic capacity of serum amyloid A protein between infectious and non-infectious febrile episodes of neutropenic patients with acute leukemia.

We studied the behavior of four major acute phase proteins (SAA, CRP, ACT and AGP) in pyrexial occurrences of 16 neutropenic patients with acute leukemia. Altogether 37 febrile episodes were recorded; 27 were infectious in origin (microbiologically documented infection and clinically documented infection, MDI/CDI group) and 10 were pyrexias of unknown origin (PUO group). In the MDI/CDI group the mean value for the highest individual SAA concentration was 282 +/- 161 mg/l and in the PUO group 95 +/- 79 mg/l. The corresponding mean values were 4.0 mg/l (range 0.2-5.5 mg/l) in 10 control patients with 1 year remission and 0.8 mg/l (range < 0.1-1.2 mg/l) in 30 healthy adults. The peak value of SAA rose above 100 mg/l in 85% of our MDI/CDI pyrexias and in 40% of PUO. More reliable results were obtained when the difference between the value on the day when pyrexia occurred and the previous day was calculated. In that case, the difference was above 75 mg/l in 23 of 27 (85%) MDI/CDI pyrexias and in none of 10 (0%) PUO. In the MDI/CDI group the mean difference was 204 +/- 137 mg/l while it was only 26 +/- 19 mg/l in the PUO group. The statistical significance was very high (p < 0.0001). The CRP monitoring was very inferior to SAA while ACT and AGP monitorings were unsatisfactory.

The pattern of expression of the chicken homolog of HOX1I in the developing limb suggests a possible role in the ectodermal inhibition of chondrogenesis.

Homeobox-containing genes have been implicated in a variety of patterning events during vertebrate limb development. In an attempt to isolate cDNAs corresponding to 5' members of the chicken HOX 4 cluster of homeobox-containing genes, a cDNA library constructed from mRNAs expressed during early stages of chick limb development was screened with probes generated by the polymerase chain reaction (PCR) using oligonucleotide primers corresponding to sequences in the homeoboxes of the human HOX4C and HOX4F genes, the human homologs of Hox-4.4 and Hox-4.6. This screening resulted in the isolation of full length cDNAs for the chicken homolog of HOX4F (cognate of mouse Hox-4.6), which we have termed GHox-4.6, and the chicken homolog of human HOX1I, which we have named GHox-1i, a paralog of Hox-4.6 in the HOX 1 cluster. The homeodomains encoded by GHox-4.6 and GHox-1i differ by only three amino acids, and the two proteins show extensive similarity along their entire lengths. Despite their sequence similarity, in situ hybridization analysis has revealed that GHox-4.6 and GHox-1i exhibit strikingly different spatial patterns of expression during embryonic chick limb development. At early stages of limb development (stages 20-22), GHox-4.6 transcripts are present in high amounts throughout the posterior half of the limb mesoderm and are absent from the anterior half of the mesoderm, an expression pattern consistent with the possible involvement of GHox-4.6 in the specification of posterior positional identity. In contrast, GHox-1i exhibits no distinct anterior-posterior polarity of expression at stage 22, but rather is expressed in high amounts throughout the mesenchyme of the limb bud. At later stages of development (stage 25), GHox-1i continues to be expressed in high amounts throughout the undifferentiated mesenchyme subjacent to the apical ectodermal ridge, and, in addition, is expressed in the mesodermal cells in the proximal peripheral regions of the limb bud subjacent to the ectoderm which are differentiating into nonchondrogenic lineages. Conversely, little or no expression of GHox-1i is detectable in the proximal central core of the limb bud where chondrogenic differentiation is occurring. Thus, GHox-1i is expressed by the undifferentiated subridge mesenchymal cells and proximal peripheral mesenchymal cells of the limb bud that are being inhibited from undergoing chondrogenesis by the apical ectodermal ridge and nonridge ectoderm.(ABSTRACT TRUNCATED AT 400 WORDS)

Beta-thalassemia in Bulgaria.

Analyses of DNA from 64 patients with thalassemia major using the hybridization technique of amplified DNA with radiolabeled synthetic oligonucleotide probes identified 13 different beta-thalassemia mutations. The codon 39 (C----T) and IVS-I-110 (G----A) mutations occurred most frequently but seven additional mutations were observed which were present at frequencies of 3.9 to 10.2%. This broad spectrum of beta-thalassemia alleles complicates the analyses for institutions involved in prenatal diagnosis. Promoter mutations were rare and the frequencies of two other mild mutations [IVS-I-6 (T----C) and the poly A mutation] were relatively low indicating that beta-thalassemia is a severe disease among Bulgarians. The high frequencies of 4.7-5.5% for the four frameshifts at codons 5, 6, 8, and 8/9 may be specific for this population.