Biomedical subjects

John Tower

Publications and source records attributed to John Tower.

13 recordsLinked to original sources

PubMed2006

Sex-specific regulation of aging and apoptosis.

Genetic analysis of Drosophila, mice and humans indicates that gene alleles, mutations and transgenes that affect life span tend to do so differently depending on the sex of the organism. The likely reason for this is that the sexes are different genotypes (e.g., X/X vs. X/Y) and face quite different environments: e.g., to reproduce, males have to mate with females while females have to mate with males. Genes are subject to different genetic interactions and different gene-by-environment effects in male vs. female. The consequence is that through evolution certain genes are differently selected and optimized for each sex. Both the mitochondrial genome and the X chromosome are asymmetrically inherited in Drosophila and mammals; through evolution these genes spend relatively more time under selection in females and are therefore expected to be better optimized for function in the female than in the male. Consistent with this the Drosophila X chromosome has been found to be a hotspot for sexually antagonistic fitness variation. Old Drosophila and old mammals exhibit apoptosis-an observation consistent with the idea that the mitochondria are less functional during aging due to maternal-only inheritance. One feature of aging that is common to Drosophila and mammals is that females tend to live longer than males, and this may be due in part to sub-optimal mitochondrial function in males. The data support the conclusion that a significant part of the aging phenotype is due to antagonistic pleiotropy of gene function between the sexes. Liberal application of Occam's razor yields a molecular model for the co-regulation of sex, apoptosis and life span based on the on/off status of a single gene: Sxl in Drosophila melanogaster and Xist in humans. Aging may simply represent an ancient and conserved mechanism by which genes re-assort.

Aging↗

PubMed2006

The murmur of old broken heartstrings.

Although it has long been hypothesized that gene expression might become more variable or noisy during aging, direct evidence has been scarce so far. A new study reports detection, by PCR analysis of individual cells, of increased cell-to-cell variability in gene expression in aging mouse heart; moreover, a similar variability can be generated in cultured cells using oxidative stress (Bahar et al., 2006).

Aging↗

PubMed2005

Superoxide dismutase evolution and life span regulation.

Superoxide is among the most abundant reactive oxygen species (ROS) produced by the mitochondria, and is involved in cellular signaling pathways. Superoxide and other ROS can damage cellular macromolecules and levels of oxidative damage products are positively correlated with aging. Superoxide dismutase (SOD) enzymes catalyze the breakdown of superoxide into hydrogen peroxide and water and are therefore central regulators of ROS levels. Genetic and transgenic manipulation of SOD activities in model systems such as S. cereviseae, mouse and Drosophila are consistent with a central role for SOD enzymes in regulating oxidative stress resistance. Over-expression of SOD in S. cereviseae and Drosophila can reduce oxidative damage and extend life span, but the mechanism(s) are not yet clear. A phylogenetic analysis of publicly available SOD protein sequences suggests several additional conserved gene families. For example, in addition to the well-characterized soluble Cu/Zn enzyme (Sod) and mitochondrial manganese-containing form (Sod2), Drosophila melanogaster is found to contain a putative copper chaperone (CCS), an extracellular Cu/Zn enzyme (Sod3), and an extracellular protein distantly related to the Cu/Zn forms (Sodq). C. elegans and blue crab are unusual in having two Mn-containing SODs, and A. gambiae contains an unusual internally repeated SOD. The most parsimonius conclusion from the analysis of the extracellular SODs is that they evolved independently multiple times by addition of a signal peptide to cytoplasmic SOD.

Animals↗

PubMed2004

Similar gene expression patterns characterize aging and oxidative stress in Drosophila melanogaster.

Affymetrix GeneChips were used to measure RNA abundance for approximately 13,500 Drosophila genes in young, old, and 100% oxygen-stressed flies. Data were analyzed by using a recently developed background correction algorithm and a robust multichip model-based statistical analysis that dramatically increased the ability to identify changes in gene expression. Aging and oxidative stress responses shared the up-regulation of purine biosynthesis, heat shock protein, antioxidant, and innate immune response genes. Results were confirmed by using Northerns and transgenic reporters. Immune response gene promoters linked to GFP allowed longitudinal assay of gene expression during aging in individual flies. Immune reporter expression in young flies was partially predictive of remaining life span, suggesting their potential as biomonitors of aging.

Aging↗

PubMed2004

There's a problem in the furnace.

A long-sought-after mutation in the manganese superoxide dismutase gene MnSOD has been isolated in Drosophila melanogaster. The MnSOD null mutation is recessive lethal, with death occurring just after eclosion. In this Perspective, I discuss these results and related work in mice within the context of the oxidative damage theory of aging.

Aging↗

PubMed2004

Effects of simultaneous over-expression of Cu/ZnSOD and MnSOD on Drosophila melanogaster life span.

The FLP-out technique, based on yeast FLP recombinase, allows induced over-expression of transgenes in Drosophila adults. With FLP-out control and over-expressing flies have identical genetic backgrounds and therefore differences in life span must result from transgene induction. The amount of over-expression achieved varies between independent transgenic lines, and previously for both Cu/ZnSOD and MnSOD life span was found to be increased in proportion to the increase in enzyme activity. To determine if greater increases in enzyme and life span could be achieved with FLP-out, enzyme over-expression and life span were analyzed in eight lines containing two MnSOD transgenes, three lines containing three MnSOD transgenes, and three lines containing a MnSOD transgene plus a Cu/ZnSOD transgene. Life span was again found to be increased in proportion to the increase in MnSOD enzyme activity, with increases of up to 40% in mean and maximum life span. However the increases in enzyme activity and life span conferred per transgene were reduced when more than one transgene was present at the same time. When the reduced efficiency of enzyme over-expression per transgene was taken into account, simultaneous over-expression of MnSOD and Cu/ZnSOD was found to have partially additive effects on life span.

Animals↗

PubMed2004

Doxycycline-regulated over-expression of hsp22 has negative effects on stress resistance and life span in adult Drosophila melanogaster.

Drosophila hsp22 is a member of the small heat shock proteins family (shsps). The hsp22 is expressed in a tissue-general pattern in response to heat stress and during normal aging, and localizes to the mitochondrial matrix, however, its exact function and targets are unknown. Hsp22 was found to be rapidly induced in response to oxidative stress, indicating that hsp22 is also an oxidative stress response gene. To assay for effects of hsp22, a ubiquitous pattern of hsp22 gene expression was generated in young flies using the "tet-on" doxycycline-regulated promoter system. The hsp22 over-expression made flies more sensitive to heat and oxidative stress, while resistance to coumarin poisoning was not affected. Life span was also reduced, particularly at higher culture temperatures. Members of other hsp families have been shown to feedback-inhibit their own expression by interacting with the heat shock transcription factor (HSF) and preventing binding to the HSEs. Induction of hsp22:lacZ and hsp70:lacZ reporter transgenes in response to acute stress was normal in the presence of hsp22 protein over-expression and in old flies, indicating that the negative effects of hsp22 are downstream of the HSF/HSE pathway and the transcriptional heat shock response. The data demonstrate a specific over-expression phenotype for hsp22 and suggest that hsp22 interacts with heat and oxidative stress resistance pathways.

Aging↗

PubMed2004

Developmental gene amplification and origin regulation.

Developmentally regulated gene amplification serves to increase the number of templates for transcription, yielding greatly increased protein and/or RNA product for gene(s) at the amplified loci. It is observed with genes that are very actively transcribed and during narrow windows of developmental time where copious amounts of those particular gene products are required. Amplification results from repeated firing of origins at a few genomic loci, while the rest of the genome either does not replicate, or replicates to a lesser extent. As such, amplification is a striking exception to the once-and-only-once rule of DNA replication and may be informative as to that mechanism. Drosophila amplifies eggshell (chorion) genes in the follicle cells of the ovary to allow for rapid eggshell synthesis. Sciara amplifies multiple genes in larval salivary gland cells that encode proteins secreted in the saliva for the pupal case. Finally, Tetrahymena amplifies its rRNA genes several thousand-fold in the creation of the transcriptionally active macronucleus. Due to the ease of molecular and genetic analysis with these systems, the study of origin regulation has advanced rapidly. Comparisons reveal an evolutionarily conserved trans-regulatory apparatus and a similar organization of sequence-specific cis-regulatory replicator and origin elements. The studies indicate a regulatory role for chromatin structure and transcriptionally active genes near the origins.

Animals↗

PubMed2004

Sequence requirements for function of the Drosophila chorion gene locus ACE3 replicator and ori-beta origin elements.

The developmentally regulated amplification of the Drosophila third chromosome chorion gene locus requires multiple chromosomal elements. Amplification control element third chromosome (ACE3) appears to function as a replicator, in that it is required in cis for the activity of nearby DNA replication origin(s). Ori-beta is the major origin in the locus, and is a sequence-specific element that is sufficient for high-level amplification in combination with ACE3. Sequence requirements for amplification were examined using a transgenic construct that was buffered from chromosomal position effects by flanking insulator elements. The parent construct supported 18- to 20-fold amplification, and contained the 320 bp ACE3, the approximately 1.2 kb S18 chorion gene and the 840 bp ori-beta. Deletion mapping of ACE3 revealed that an evolutionarily conserved 142 bp core sequence functions in amplification in this context. Several deletions had quantitative effects, suggesting that multiple, partially redundant elements comprise ACE3. S. cerevisiae ARS1 origin sequences could not substitute for ori-beta, thereby confirming the sequence specificity of ori-beta. Deletion mapping of ori-beta identified two required components: a 140 bp 5' element and a 226 bp A/T-rich 3' element called the beta-region that has significant homology to ACE3. Antibody to the origin recognition complex subunit 2 (ORC2) recognizes large foci that localize to the endogenous chorion gene loci and to active transgenic constructs at the beginning of amplification. Mutations in Orc2 itself, or the amplification trans regulator satin eliminated the ORC2 foci. By contrast, with a null mutation of chiffon (dbf4-like) that eliminates amplification, diffuse ORC2 staining was still present, but failed to localize into foci. The data suggest a novel function for the Dbf4-like chiffon protein in ORC localization. Chromosomal position effects that eliminated amplification of transgenic constructs also eliminated foci formation. However, use of the buffered vector allowed amplification of transgenic constructs to occur in the absence of detectable foci formation. Taken together, the data suggest a model in which ACE3 and ori-beta nucleate the formation of a ORC2-containing chromatin structure that spreads along the chromosome in a mechanism dependent upon chiffon.

Animals↗

PubMed2003

A search for doxycycline-dependent mutations that increase Drosophila melanogaster life span identifies the VhaSFD, Sugar baby, filamin, fwd and Cctl genes.

BACKGROUND: A P-type transposable element called PdL has been engineered with a doxycycline-inducible promoter directed out through the 3' end of the element. Insertion of PdL near the 5' end of a gene often yields doxycycline-dependent overexpression of that gene and a mutant phenotype. This functional genomics strategy allows for efficient screening of large numbers of genes for overexpression phenotypes. RESULTS: PdL was mobilized to around 10,000 new locations in the Drosophila melanogaster genome and used to search for genes that would extend life span when overexpressed. Six lines were identified in which there was a 5-17% increase in life span in the presence of doxyxcycline. The mutations were molecularly characterized and in each case a gene was found to be overexpressed using northern blots. Two genes did not have previously known phenotypes and are implicated in membrane transport: VhaSFD encodes a regulatory subunit of the vacuolar ATPase proton pump (H+-ATPase), whereas Sugar baby (Sug) is related to a maltose permease from Bacillus. Three PdL mutations identified previously characterized genes: filamin encodes the homolog of an actin-polymerizing protein that interacts with presenilins. four wheel drive (fwd) encodes a phosphatidylinositol-4-kinase (PI 4-kinase) and CTP:phosphocholine cytidylyltransferase-l (Cctl) encodes the rate-limiting enzyme in phosphatidylcholine synthesis. Finally, an apparently novel gene (Red herring, Rdh) was found in the first intron of the encore gene. CONCLUSIONS: Screening for conditional mutations that increase Drosophila life span has identified genes implicated in membrane transport, phospholipid metabolism and signaling, and actin cytoskeleton organization.

1-Phosphatidylinositol 4-Kinase↗

PubMed2002

Setting a trap for aging-related genes in Drosophila.

In the October 2002 issue of Aging Cell, Seroude et al. used an enhancer trap technique to identify Drosophila genes whose expression changes with age. In this Perspective, the author discusses the implications of these results, including the possibility of using the enhancer trap lines to identify biomarkers of aging.

Animals↗

PubMed2002

Doxycycline-induced expression of sense and inverted-repeat constructs modulates phosphogluconate mutase (Pgm) gene expression in adult Drosophila melanogaster.

BACKGROUND: A tetracycline-regulated (conditional) system for RNA interference (RNAi) would have many practical applications. Such a strategy was developed using RNAi of the gene for phosphogluconate mutase (Pgm). Pgm is a candidate lifespan regulator: PgmS allele frequency is increased by selection for increased lifespan, whereas PgmM and PgmF allele frequencies are decreased. RESULTS: The Pgm alleles were cloned and sequenced and were found to differ by amino-acid substitutions consistent with the relative electrophoretic mobilities of the proteins. The 'tet-on' doxycycline-regulated promoter system was used to overexpress PgmS in a wild-type (PgmM) background. Enzyme activity increases of two- to five-fold were observed in five independent transgenic lines. Tet-on was also used to drive expression of an inverted-repeat fragment of Pgm coding region. The inverted-repeat transcript was expected to form a dsRNA hairpin, induce RNAi, and thereby reduce endogenous Pgm gene expression at the RNA level. Endogenous Pgm RNA levels in adult flies were found to be reduced or eliminated by doxycycline treatment in five independent inverted-repeat transgenic lines. Our results show that doxycycline-regulated expression of inverted-repeat constructs can cause a conditional reduction in specific gene expression. The effect of sense and inverted-repeat construct expression on lifespan was assayed in multiple transgenic lines. Under the conditions tested, altered Pgm gene expression had no detectable effect on adult Drosophila lifespan. CONCLUSIONS: A system for conditional RNAi in Drosophila adults shows promise for assay of gene functions during aging. Our results indicate that Pgm does not have a simple strong effect on longevity.

Aging↗

PubMed2002

Induced overexpression of mitochondrial Mn-superoxide dismutase extends the life span of adult Drosophila melanogaster.

A transgenic system ("FLP-out") based on yeast FLP recombinase allowed induced overexpression of MnSOD enzyme in adult Drosophila melanogaster. With FLP-out a brief heat pulse (HP) of young, adult flies triggered the rearrangement and subsequent expression of a MnSOD transgene throughout the adult life span. Control (no HP) and overexpressing (HP) flies had identical genetic backgrounds. The amount of MnSOD enzyme overexpression achieved varied among six independent transgenic lines, with increases up to 75%. Life span was increased in proportion to the increase in enzyme. Mean life span was increased by an average of 16%, with some lines showing 30-33% increases. Maximum life span was increased by an average of 15%, with one line showing as much as 37% increase. Simultaneous overexpression of catalase with MnSOD had no added benefit, consistent with previous observations that catalase is present in excess in the adult fly with regard to life span. Cu/ZnSOD overexpression also increases mean and maximum life span. For both MnSOD and Cu/ZnSOD lines, increased life span was not associated with decreased metabolic activity, as measured by O2 consumption.

Analysis of Variance↗