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PubMed2006

Resolving the genetic basis of invasiveness and predicting invasions.

Considerable effort has been invested in determining traits underlying invasiveness. Yet, identifying a set of traits that commonly confers invasiveness in a range of species has proven elusive, and almost nothing is known about genetic loci affecting invasive success. Incorporating genetic model organisms into ecologically relevant studies is one promising avenue to begin dissecting the genetic underpinnings of invasiveness. Molecular biologists are rapidly characterizing genes mediating developmental responses to diverse environmental cues, i.e., genes for plasticity, as well as to environmental factors likely to impose strong selection on invading species, e.g., resistance to herbivores and competitors, coordination of life-history events with seasonal changes, and physiological tolerance of heat, drought, or cold. Here, we give an overview of molecular genetic tools increasingly used to characterize the genetic basis of adaptation and that may be used to begin identifying genetic mechanisms of invasiveness. Given the divergent traits that affect invasiveness, "invasiveness genes" common to many clades are unlikely, but the combination of developmental genetic advances with further evolutionary studies and modeling may provide a framework for identifying genes that account for invasiveness in related species.

Arabidopsis↗
PubMed1986

Identification and sequence of a gene required for a developmentally regulated DNA excision in Anabaena.

Vegetative cells of the cyanobacterium Anabaena contain an 11 kb DNA element within the coding region of the nifD gene. This element is excised by site-specific recombination between directly repeated 11 bp sequences at each of its ends during differentiation of nitrogen-fixing cells called heterocysts. Site-specific recombination, leading to the same rejoined nifD gene, was observed during propagation in E. coli of a fragment containing the 11 kb element and flanking sequences. An assay for excision of the element in E. coli was developed, based on mini-Mu-lac transposition into the element. Since the 11 kb element lacks an origin of replication, its excision results in loss of lac and conversion of blue colony-forming cells to white on X-gal plates. Insertion and deletion mutagenesis identified a region of the element needed for excision. Mutations in this region could be complemented by a 6 kb fragment containing an open reading frame that runs counter to those of the nif genes, beginning 240 bp from the recombination site.

Base Sequence↗
PubMed1995

Gsh-2, a murine homeobox gene expressed in the developing brain.

A novel murine dispersed homeobox gene, designated Gsh-2, is described. Analysis of cDNA sequence, including the full open reading frame, reveals an encoded homeodomain that is surprisingly similar to those of the Antennapedia-type clustered Hox genes. In addition, the encoded protein includes polyhistidine and polyalanine tracts, as observed for several other genes of developmental significance. In situ hybridizations showed Gsh-2 expression in the developing central nervous system, including the ganglionic eminences of the forebrain, the diencephalon, which gives rise to the thalamus and hypothalamus, and in the hindbrain. Furthermore, a random oligonucleotide selection and PCR amplification procedure was used to define a target DNA binding sequence, CNAATTAG, as a first step towards the identification of downstream target genes.

Amino Acid Sequence↗
PubMed1996

Developmental regulation of perlecan gene expression in aortic smooth muscle cells.

Heparan sulfate proteoglycans (HSPGs) are believed to act as potent endogenous regulators of vascular smooth muscle cell (SMC) replication, migration, gene expression and differentiation. Here we describe the pattern of expression of perlecan, the predominant basement membrane HSPG, during aortic development in the rat. Expression of perlecan mRNA and protein in the aortic SMC was first significantly observed at day e19 (day 19 of embryonic development), a time which marks a dramatic switch in SMC replication rate and growth phenotype. Expression of perlecan message and protein was high throughout fetal and early neonatal life, and it remained readily detectable in the adult aorta. Using a double-labeling technique (in situ hybridization for perlecan message coupled with bromodeoxyuridine immunohistochemistry), we determined the relationship between DNA synthesis and perlecan mRNA expression in individual SMC at days e17-e21; we found that perlecan gene expression was largely limited to non-replicating cells. Consistent with the in vivo data, perlecan mRNA was undetectable in cultured e17 SMC by Northern or RT-PCR analysis, while in cultured adult SMC, perlecan mRNA was significantly higher in non-replicating (serum-starved) cultures compared to replicating cultures. Treatment of growth-arrested adult SMC cultures with heparin caused a further accumulation in perlecan mRNA levels. The data suggest that the expression of perlecan by vascular SMC is regulated by apparent developmental age as well as by cellular growth state. The developmentally times expression of perlecan in the aortic wall may contribute to the establishment and/or maintenance of vascular SMC differentiation and quiescence.

Animals↗
PubMed2001

Developmental regulation of leptin gene expression in rat brain and pituitary.

We have previously reported the expression of leptin mRNA and protein in adult rat brain and pituitary gland. We report here the presence of leptin and leptin receptor mRNA in neonatal female rat brain and pituitary using RT-PCR as well as leptin and leptin receptor immunoreactivity in neonatal rat brain. In addition, we describe age-related changes in leptin mRNA expression in female rat brain and pituitary from postnatal day 2 to 28, evaluated using semi-quantitative RT-PCR analysis. Age-related differences in leptin (ob) mRNA levels were tissue-dependent. The most striking developmental changes were noted in the pituitary and cerebral cortex. In the pituitary, ob mRNA levels were maximal during postnatal days 7-14 and fell sharply by postnatal day 22. In cortex, ob mRNA levels were low in neonatal pups (day 2-7) but increased significantly between postnatal days 14 and 28. Leptin mRNA was detectable at postnatal day 2 in hypothalamus and subcutaneous fat. No significant differences in the level of expression were observed between postnatal day 2 and 28. Serum leptin levels were highest at day 7-14 and decreased significantly by day 21-28, coincident with the fall in pituitary leptin expression. The high levels of leptin expression in the neonatal pituitary suggest that this gland may contribute to the circulating leptin levels during early postnatal development, when adipose deposits are minimal. These data indicate that regulation of leptin gene expression in the postnatal period is tissue-dependent, a finding, which suggests that local leptin expression may have important functional significance in the development of the brain-pituitary system.

Age Factors↗
PubMedDate not supplied

Expression of nine developmental stage-specific genes in human kidney and cultured renal cells.

BACKGROUND: Several genes transiently expressed during the maturation of the metanephrogenic mesenchyme have been reported in recent years while there is accumulating evidence of a reverted developmental pathway during tissue damage and loss of function. METHODS: Here we studied the expression of nine genes associated with kidney maturation from samples of normal human fetal, juvenile and adult kidneys and cultured glomerular cells using Northern blotting analysis. Subsequently, kidneys from patients with congenital nephrotic syndrome of the Finnish type (CNF), presenting with heavy proteinuria, and Wilms' tumor tissue were studied for the corresponding expression pattern for evidence of dedifferentiation/persistence of a fetal expression pattern. RESULTS: All the genes studied had their distinct expression pattern within the tissues and cells tested. No conclusive evidence of dedifferentiation could be obtained in the samples from CNF kidneys, whereas, as expected, the gene expression pattern of Wilms' tumor tissue was highly similar to that of fetal kidney tissue. CONCLUSION: Some genes thought to be only transiently expressed during kidney maturation were, however, constantly found in the samples from fetal to newborn and adult kidneys.

Adolescent↗
PubMed2002

Host recognition by pathogenic fungi through plant flavonoids.

A common characteristic among fungal pathogens of plants is that each specializes on a narrow range of specific plants as hosts. One adaptation to a specific host plant is the recognition of the host's chemicals which can be used to trigger genes or developmental pathways needed for pathogenesis. The production of characteristic flavonoids by plants, particularly those exuded from roots by legumes, appear to be used as signals for various microbes, including symbionts as well as pathogens. Nectria haematococca MPVI (anamorph: Fusarium solani) is a soil-borne pathogen of garden pea (Pisum sativum) which serves as a useful model in studying host flavonoid recognition. This fungus displays flavonoid induction of specific pathogenicity genes as well as stimulation of development needed for pathogenesis. Here, we summarize the study of flavonoid-inducible signal pathways which regulate these trait, through identification of transcription factors and regulatory components which control these responses. The characterization of the components a pathogen uses to specifically recognize its host provides insights into the host adaptation process at the molecular level.

Adaptation, Physiological↗
PubMed1993

Spatial restriction of cellular differentiation.

Myxococcus xanthus cells differentiate into myxospores within a fruiting body, an aggregate of approximately 10(5) cells. Previous work had discerned an inner and outer domain within the fruiting body differentiated by cell density and cell alignment. To test whether the two domains might play different roles in spore differentiation, developmentally regulated gene fusions were screened for expression restricted to one domain or the other. Transcriptional lacZ fusions to 80 developmentally regulated genes were examined and eight fusions were found that restricted expression to the inner domain, while one fusion, omega 7621, showed initial expression in the outer domain. Initial omega 7621 expression coincided with patches of spore precursors evident in bright-field microscopy. Later in development, both omega 7621 expression and the patches expanded inward, eventually filling both the inner and outer domains. Previous work had also shown that high cell density and cell alignment are required for transmission of the C-signal, which is needed to initiate spore differentiation. Evidence is presented for a novel morphogenetic mechanism in which C-signaling in the outer (high density) domain initiates spore differentiation. It is proposed that spore precursors are passively transported from the outer to the inner domain by the movements of undifferentiated rod cells. Reconstruction experiments showed that developing rod cells move with sufficient force to displace spores. Spore precursors thus accumulate in the inner domain where they express spore-specific genes at high levels and account for inner domain specific expression.

Biological Transport↗
PubMed2002

Molecular cloning and characterization of cDNAs encoding heterotrimeric G protein alpha and beta subunits from potato (Solanum tuberosum L.).

Two cDNAs, STGA2 and STGB2, that encode heterotrimeric G protein alpha and beta subunit proteins, respectively, were cloned from an early tuber cDNA library of potato (Solanum tuberosum cv. Superior). The cDNA of STGA2 encoded 384 amino acids, which showed 75-98% identities to plant Ga-subunits; STGB2 encoded 377 amino acids, which showed 83-92% identities to plant Gbeta-subunits. The transcript levels of the two genes were abundant in leaves, shoots, axially buds, unopened flowers, and active growing sprouts. However, the transcripts were barely detectable in roots. The expressions of STGA2 and STGB2 were up-regulated by light. Interestingly, the STGA2 and STGB2 gene expression showed synchronous patterns in the examined organs. During the early tuber development, the transcripts of STGA2 and STGB2 were abundant in unswollen stolons, swollen stolons, and new tubers, but were undetected in matured tubers. This indicates that potato Galpha- and beta-subunit genes are developmentally regulated. Based on these observations, we propose that heterotrimeric G proteins may be involved in the signaling pathway during potato tuber development.

Amino Acid Sequence↗
PubMed1992

DNA binding factors which interact with the Sp1 site of the chicken delta 1-crystallin promoter are developmentally regulated.

Transcription of the delta 1-crystallin gene is developmentally regulated in the embryonic chicken lens. Previous work defined a positive transcription regulatory element between positions -120 and -43 of the delta 1-crystallin promoter. This region contains a putative Sp1 binding site (-78 to -71), adjacent to a CAAT box (-70 to -67). Gel retardation assays using lens nuclear extracts revealed two protein-DNA complexes which involved the Sp1 site. The formation of the complexes increased from day 6 to day 11 of embryogenesis (period of lens organogenesis) peaked between days 11 and 15, then decreased in a non-parallel manner until hatching (day 21). A point mutation in the Sp1 binding site of the delta 1-crystallin promoter abolished formation of one of the complexes (complex 1, slower in mobility), while point mutations in the CAAT box had no effect on the formation of either complex. Studies using purified Sp1 protein and increasing amounts of embryonic chicken lens nuclear extracts showed cooperativity in the formation of both complexes, more remarkable with complex 1.

Aging↗
PubMed1997

Epidermal muscle attachment site-specific target gene expression and interference with myotube guidance in response to ectopic stripe expression in the developing Drosophila epidermis.

The egr-type zinc-finger transcription factor encoded by the Drosophila gene stripe (sr) is expressed in a subset of epidermal cells to which muscles attach during late stages of embryogenesis. We report loss-of-function and gain-of-function experiments indicating that sr activity provides ectodermal cells with properties required for the establishment of a normal muscle pattern during embryogenesis and for the differentiation of tendon-like epidermal muscle attachment sites (EMA). Our results show that sr encodes a transcriptional activator which acts as an autoregulated developmental switch gene. sr activity controls the expression of EMA-specific target genes in cells of ectodermal but not of mesodermal origin. sr-expressing ectodermal cells generate long-range signals that interfere with the spatial orientation of the elongating myotubes.

Animals↗
PubMed2000

Chromatin remodelling during the life cycle of trypanosomatids.

The mechanisms which control the expression of developmentally regulated genes in trypanosomatids remain unclear. The genes are grouped together into transcription units that are co-transcribed to yield polycistronic RNAs. Trans-splicing and polyadenylation give rise to mature, monocistronic mRNAs. It is difficult to imagine that expression of these genes is controlled at the level of transcription initiation because this would suggest that the genes are transcribed at the same rate. This is not the case, because at any given developmental stage in trypanosomes or Leishmania, genes transcribed from the same transcription unit are expressed at different levels within the cell. Consequently, these parasites must rely on post-transcriptional or post-translational mechanisms to generate the appropriate levels of gene product within the cell. There are no well-established examples of RNA polymerase II promoters in trypanosomes or Leishmania. However, the promoters for genes encoding the variant surface glycoprotein (VSG) and the procyclic acidic repetitive protein (PARP) have been identified and resemble ribosomal RNA polymerase I promoters. In higher eukaryotes where the mechanisms regulating transcription are clearer, there is increasing evidence that epigenetic factors, such as histones and modified bases, influence gene expression. Chemical modification of these factors can restructure chromatin and lead to gene activation or silencing. In trypanosomatids, an epigenetic mechanism for the control of developmentally expressed genes is a possibility. In this review, chromatin remodelling during the life and cell cycle of trypanosomes and Leishmania is explored, and the influence of epigenetic factors such as histones and modified bases on this process is discussed.

Animals↗
PubMed2006

Icariin-mediated expression of cardiac genes and modulation of nitric oxide signaling pathway during differentiation of mouse embryonic stem cells into cardiomyocytes in vitro.

AIM: To investigate effects of icariin on cardiac gene expression and the modulation of nitric oxide (NO) signal transduction during the differentiation of embryonic stem (ES) cells into cardiomyocytes in vitro. METHODS: The expression levels of cardiac developmental-dependent genes were measured using reverse transcription-polymerase chain reaction (RT-PCR). The chronotropic responses of cardiomyocytes to b-adrenoceptor stimulation were determined. The levels of cAMP and cGMP in ES cells were measured using radioimmunoassay. Endogenous NO levels were measured by using the Griess reaction. Aminoguanidine (AG) was used to confirm the influence of icariin on the endogenous NO signal pathway. RESULTS: Icariin significantly elevated mRNA levels of cardiac transcription factors GATA4 and Nkx2.5, and cardiac-specific alpha-MHC, MLC-2v and beta-AR genes in a concentration- and time-dependent manner (P<0.05). Cardiomyocytes derived from embryoid body (EB) treated with icariin were more sensitive to isoprenaline (P<0.01). Treatment of ES cells with icariin resulted in a continued elevation in the cAMP/cGMP ratio before a shift to the cardiomyocyte phenotype (P<0.05). AG decreased the NO level, and delayed and decreased the incidence of contracting EB to only approximately 35% on d 5+11, an effect that could be rescued by icariin. When cells were cocultured with icariin and AG, the percentage of beating EB reached a peak level of 73% on d 5+11 (P<0.05). CONCLUSION: The inducible effects of icariin were partly related to increase in the expression of cardiac developmental-dependent genes, and elevation of the cAMP/cGMP ratio in ES cells, as well as upregulation of endogenous NO generation during the early stages of cardiac development.

Animals↗
PubMed1997

Interplay of the E box, the cyclic AMP response element, and HTF4/HEB in transcriptional regulation of the neurospecific, neurotrophin-inducible vgf gene.

vgf is a neurotrophin response-specific, developmentally regulated gene that codes for a neurosecretory polypeptide. Its transcription in neuronal cells is selectively activated by the neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor, and neurotrophin 3, which induce survival and differentiation, and not by epidermal growth factor. We studied a short region of the rat vgf promoter which is essential for its regulated expression. A cyclic AMP response element (CRE) within this region is necessary for NGF induction of vgf transcription. Two sites upstream of CRE, an E box and a CCAAT sequence, bind nuclear protein complexes and are involved in transcriptional control. The E box has a dual role. It acts as an inhibitor in NIH 3T3 fibroblasts, together with a second E box located downstream, and as a stimulator in the NGF-responsive cell line PC12. By expression screening, we have isolated the cDNA for a basic helix-loop-helix transcription factor, a homolog of the HTF4/HEB E protein, that specifically binds the vgf promoter E box. The E protein was present in various cell lines, including PC12 cells, and was a component of a multiprotein nuclear complex that binds the promoter in vitro. The E box and CRE cooperate in binding to this complex, which may be an important determinant for neural cell-specific expression.

3T3 Cells↗
PubMed1997

Isolation of Aspergillus nidulans mutants that overcome brlA-induced growth arrest.

The Aspergillus nidulans brlA gene is a primary regulator of development-specific gene expression during conidiation. Forced activation of brlA in vegetative cells leads to inappropriate induction of conidiophore formation and causes growth to stop. In fact, when conidia containing a nutritionally inducible brlA gene fusion are placed on inducing medium, they fail to germinate. We used this phenotype to select 174 mutants that continue growing following such forced brlA activation. Forty-six of these mutants also produced abnormal developmental structures during air-induced conidiation as expected if the mutations resulted in an altered response to BrlA (designated sbr mutants for suppressors of brlA response). The predominant mutant class identified was defective in a known developmental regulatory gene, abaA. We also identified mutants with defects in the previously characterized early acting developmental regulatory genes flbB and flbD and in four previously undescribed loci designated sbrA-D. sbrA mutants represent the second largest group and are characterized by production of conidophore stalks that lack a normal vesicle and form branching sterigmata that rarely make spores. Because abaA expression could not be detected in sbrA mutants following brlA activation we propose that sbrA functions as a developmental modifier, participating in brlA-dependent activation of other developmental regulators.

Aspergillus nidulans↗
PubMed1998

Serum response factor is essential for mesoderm formation during mouse embryogenesis.

The transcription factor serum response factor (SRF), a phylogenetically conserved nuclear protein, mediates the rapid transcriptional response to extracellular stimuli, e.g. growth and differentiation signals. DNA- protein complexes containing SRF or its homologues function as nuclear targets of the Ras/MAPK signalling network, thereby directing gene activities associated with processes as diverse as pheromone signalling, cell-cycle progression (transitions G0-G1 and G2-M), neuronal synaptic transmission and muscle cell differentiation. So far, the activity of mammalian SRF has been studied exclusively in cultured cells. To study SRF function in a multicellular organism we generated an Srf null allele in mice. SRF-deficient embryos (Srf -/-) have a severe gastrulation defect and do not develop to term. They consist of misfolded ectodermal and endodermal cell layers, do not form a primitive streak or any detectable mesodermal cells and fail to express the developmental marker genes Bra (T), Bmp-2/4 and Shh. Activation of the SRF-regulated immediate early genes Egr-1 and c-fos, as well as the alpha-Actin gene, is severely impaired. Our study identifies SRF as a new and essential regulator of mammalian mesoderm formation. We therefore suggest that in mammals Ras/MAPK signalling contributes to mesoderm induction, as is the case in amphibia.

Animals↗
PubMed2003

Changes in gravitational force induce alterations in gene expression that can be monitored in the live, developing zebrafish heart.

Little is known about the effect of microgravity on gene expression, particularly in vivo during embryonic development. Using transgenic zebrafish that express the gfp gene under the influence of a beta-actin promoter, we examined the affect of simulated-microgravity on GFP expression in the heart. Zebrafish embryos, at the 18-20 somite-stage, were exposed to simulated-microgravity for 24 hours. The intensity of GFP fluorescence associated with the heart was then determined using fluorescence microscopy. Our measurements indicated that simulated-microgravity induced a 23.9% increase in GFP-associated fluorescence in the heart. In contrast, the caudal notochord showed a 17.5% increase and the embryo as a whole showed only an 8.5% increase in GFP-associated fluorescence. This suggests that there are specific effects on the heart causing the more dramatic increase. These studies indicate that microgravity can influence gene expression and demonstrate the usefulness of this in vivo model of 'reporter-gene' expression for studying the effects of microgravity.

Animals↗
PubMed1997

Vertebrate development in the environment of space: models, mechanisms, and use of the medaka.

With the advent of space travel, it is of immediate interest and importance to study the effects of exposure to various aspects of the altered environment of space, including microgravity, on Earth-based life forms. Initial studies of space travel have focused primarily on the short-term effects of radiation and microgravity on adult organisms. However, with the potential for increased lengths of time in space, it is critical to now address the effects of space on all phases of an organism's life cycle, from embryogenesis to post-natal development to reproduction. It is already possible for certain species to undergo multiple generations within the confines of the Mir Space Station. The possibility now exists for scientists to consider the consequences of even potentially subtle defects in development through multiple phases of an organism's life cycle, or even through multiple generations. In this discussion, we highlight a few of the salient observations on the effects of the space environment on vertebrate development and reproductive function. We discuss some of the many unanswered questions, in particular, in the context of the choice of appropriate models in which to address these questions, as well as an assessment of the availability of hardware already existing or under development which would be useful in addressing these questions.

Animals↗