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PubMed2003

Molecular characterization and spatial expression of the sunflower ABP1 gene.

We have used RT-PCR and low-stringency cDNA library screening to isolate the coding sequence of the sunflower auxin-binding protein (ABP1). All the clones analysed contained the same nucleotide sequence, suggesting that ABP1 is encoded by a single-copy gene in sunflower. The deduced amino acid sequence shows a high degree of similarity with ABP1 proteins from other plant species. Most remarkably, the sunflower protein lacks two cysteine residues present in all other plant ABPs known to date and shown to be involved in a disulfide bridge in the maize protein. Genomic Southern hybridization data support the existence of a single copy of the ABP1 gene in the sunflower genome. Northern hybridization corroborated earlier observations indicating that the steady-state level of ABP1 transcript is higher in actively dividing and growing organs than in the rest of the plant: it is more abundant in the shoot apex, floral buds and immature embryos than in mature leaves, stem, roots and ray flowers. To characterize the tissular ABP1 transcript distribution in sunflower, various organ sections were analysed upon in situ hybridization. Localized accumulation of the ABP1 transcript suggests that its spatial expression is highly regulated at the tissue level. In addition, the transcript preferentially accumulates in tissues having a high rate of cellular division, such as shoot and root apical meristems, leaf primordia and pro-vascular tissues. The ABP1 expression pattern was also studied at a temporal scale during lateral root formation. Real time PCR showed an elevation of the steady state level of the ABP1 transcript in root axes after 36 h of seed germination. In situ hybridization revealed that this global increase is the result of local accumulation of the ABP1 transcript in lateral root primordia, which are known to develop under auxin action. The possibility that a high ABP1 expression level correlates with a high cellular sensitivity to auxin is discussed.

Amino Acid Sequence↗

PubMed2002

Nod factor induction of reactive oxygen species production is correlated with expression of the early nodulin gene rip1 in Medicago truncatula.

Plant genes that are specifically activated by the rhizobial lipochitooligosaccharide signal molecule (Nod factor) in legume hosts are collectively referred to as nodulins. Although nodulin gene expression is both spatially and temporally correlated with symbiosis, the function of these genes and the molecular events underlying their expression remain unknown. Sequence analysis of rip1, an early nodulin gene encoding a putative peroxidase protein, revealed the existence of sequence motifs with homology to reactive oxygen species (ROS) responsive cis elements. Here we report that recognition of compatible Nod factor rapidly stimulates a spatially localized production of reactive oxygen species in legume roots. Sinorhizobium meliloti mutants that produce an altered Nod factor structure and a nonnodulating plant mutant, dmi1-1, that is implicated in Nod factor signal transduction are equally impaired in the ability to elicit ROS production and rip1 expression. Interestingly, both rip1 transcription and ROS production exhibit the same tissue-specific pattern of localization. Moreover, exogenous hydrogen peroxide is sufficient to activate rip1 transcription. Taken together, these results suggest that ROS production is a consequence of specific Nod factor perception and implicate H2O2 produced during this response as a mediator of Nod factor-induced rip1 expression.

Gene Expression Regulation, Developmental↗

PubMed2004

Microarray analysis of gene expression on herbal glycoside recipes improving deficient ability of spatial learning memory in ischemic mice.

In order to reveal the mechanism of herbal glycoside recipes retrieving deficient ability of spatial learning memory in mice suffering from cerebral ischemia/reperfusion, a microarray system was used to analyze gene expression in those groups with increasing ability of spatial learning memory who were different from ischemic mice. In this work, we reported a comprehensive characterization of gene expression profiles of mouse hippocampus by the use of cDNA microarray system containing 1176 known genes in middle cerebral artery occlusion (MCAO) ischemic mice after treating with different dosage recipes of glycoside herbs (30, 90, and 270 mg/kg). The ability of spatial learning memory in ischemic mice was found to be decreased. The pathological process in ischemic mouse brain showed that a complex related to 100 genes' expression yielded 1.8-fold. Dose-dependent effects showed an improvement in the deficient ability and reduction in infarct volume when treated with glycoside recipes. Many genes (38-46) in expression were found greater than 1.8-fold in those effective recipes groups, including genes in cell cycle regulation, signal transduction, nerve system transcription factors, DNA binding protein, etc. Nine genes related to retrieving deficient ability of spatial learning memory treated with glycoside recipes were also found in this study. These results suggest that microarray analysis of gene expression might be useful for elucidating the mechanisms of pharmacological function of recipes.

14-3-3 Proteins↗

PubMed2004

Gibberellin induces alpha-amylase gene in seed coat of Ipomoea nil immature seeds.

Two full-length cDNAs encoding gibberellin 3-oxidases, InGA3ox1 and InGA3ox2, were cloned from developing seeds of morning glory (Ipomoea nil (Pharbitis nil) Choisy cv. Violet) with degenerate-PCR and RACEs. The RNA-blot analysis for these clones revealed that the InGA3ox2 gene was organ-specifically expressed in the developing seeds at 6-18 days after anthesis. In situ hybridization showed the signals of InGA3ox2 mRNA in the seed coat, suggesting that active gibberellins (GAs) were synthesized in the tissue, although no active GA was detected there by immunohistochemistry. In situ hybridization analysis for InAmy1 (former PnAmy1) mRNA showed that InAmy1 was also synthesized in the seed coat. Both InGA3ox2 and InAmy1 genes were expressed spatially overlapped without a clear time lag, suggesting that both active GAs and InAmy1 were synthesized almost simultaneously in seed coat and secreted to the integument. These observations support the idea that GAs play an important role in seed development by inducing alpha-amylase.

Amino Acid Sequence↗

PubMed1999

Proliferative hemangiomas: analysis of cytokine gene expression and angiogenesis.

Hemangiomas are benign vascular tumors of childhood that can lead to disfigurement and/or life-threatening consequences. The pathogenesis of hemangioma formation is likely to involve increased angiogenesis. Basic fibroblast growth factor and vascular endothelial growth factor are cytokines that stimulate angiogenesis in multiple in vivo and in vitro models. Proliferative hemangiomas have been found to have elevated levels of basic fibroblast growth factor and vascular endothelial growth factor protein, but the gene expression of these cytokines in human specimens has not been previously studied. We examined the gene expression and spatial distribution of basic fibroblast growth factor and vascular endothelial growth factor messenger RNA in proliferative versus involuted human hemangioma specimens using nonisotopic in situ hybridization techniques. Thirteen hemangioma specimens were harvested during initial surgical excision. In situ hybridization was performed on frozen sections of both proliferative and involuted hemangioma specimens using genetically engineered antisense probes specific for basic fibroblast growth factor and vascular endothelial growth factor messenger RNA. Controls were an interleukin-6 sense sequence and a transforming growth factor-beta 1 antisense sequence. A large number of cells within the specimens of proliferative hemangiomas revealed localized gene expression of basic fibroblast growth factor and vascular endothelial growth factor messenger RNA (626 +/- 129 and 1660 +/- 371 cells/mm2, respectively). The majority of the cells were endothelial in origin. In contrast, involuted hemangioma specimens revealed significantly lower numbers of cells staining positive for basic fibroblast growth factor and vascular endothelial growth factor messenger RNA (44 +/- 11 and 431 +/- 76 cells/mm2, respectively; p < 0.05). Transforming growth factor-beta 1 messenger RNA was slightly more expressed by involuted hemangiomas (117 +/- 30 cells/mm2). There were very low levels of transforming growth factor-beta 1 gene expression from proliferative hemangiomas (37 +/- 24 cells/mm2; p < 0.02). These data demonstrate that (1) in situ hybridization allows identification and relative quantitation of cells expressing messenger RNA for specific growth factors in human hemangioma specimens; (2) basic fibroblast growth factor and vascular endothelial growth factor messenger RNA are up-regulated in proliferative hemangiomas; and (3) transforming growth factor-beta 1 messenger RNA remains low in both proliferative and involuted hemangiomas. Because basic fibroblast growth factor and vascular endothelial growth factor messenger RNA have been implicated in the pathobiology of human hemangioma formation, biochemical modulation of these angiogenic cytokines may eventually help inhibit proliferation and promote regression of hemangiomas.

Child, Preschool↗

PubMed2002

The human involucrin gene contains spatially distinct regulatory elements that regulate expression during early versus late epidermal differentiation.

Human involucrin (hINV) is a keratinocyte protein that is expressed in the suprabasal compartment of the epidermis and other stratifying surface epithelia. Involucrin gene expression is initiated early in the differentiation process and is maintained until terminal cell death. The distal regulatory region (DRR) is a segment of the hINV promoter (nucleotides -2473/-1953) that accurately recapitulates the normal pattern of suprabasal (spinous and granular layer) expression in transgenic mouse epithelia. To identify sequences that mediate expression at specific stages of differentiation, we divided the DRR into two segments, a 376 nucleotide upstream region (DRR(-2473/-2100)) and a 147 nucleotide downstream region (DRR(-2100/-1953)), and evaluated the ability of these sequences to drive expression in transgenic mice. The DRR(-2473/-2100) segment drives expression at a level comparable to that observed for the DRR, but expression is restricted to the upper granular layers (i.e., no spinous layer expression). In contrast, the DRR(-2100/-1953) segment does not drive expression. However, reassembling the DRR restores the complete range of expression. These results suggest that two distinct, spatially-separate elements are required to specify the complete differentiation-dependent program of involucrin gene expression. To identify specific transcription factor binding sites involved in this regulation, we mutated an activator protein-1 binding site, AP1-5, located within DRR(-2473/-2100) segment. This site binds AP1 transcription factors present in mouse epidermal extracts, and its mutation eliminates appropriate hINV expression. This result suggests that AP1 factors participate as components of a multi-component transcription factor complex that is required for regulation.

Animals↗

PubMed2002

Conditional gene expression in the respiratory epithelium of the mouse.

Transgenic mouse models mediating conditional temporal and spatial regulation of gene expression to the respiratory epithelium were developed utilizing the reverse tetracycline transactivator (rtTA) expressed under the control of SP-C and CCSP promoters. Luciferase activity was detected in the lungs of fetal and adult double transgenic mice but was not detected in other tissues or in single transgenic mice. In adult mice, maximal luciferase activity was detected 16 h after the administration of doxycycline in the drinking water, or 2 h after the injection of doxycycline. Activation of the transgene was observed after the administration of doxycycline in food pellets. After prolonged exposure to doxycycline, luciferase activity decreased slowly following removal of doxycycline, suggesting the importance of tissue pools which maintained expression of the transgene. In SP-C-rtTA mice, exposure of the pregnant dam to doxycycline induced luciferase activity in fetal lung tissue as early as E10.5. Luciferase activity was maintained in the lung tissue of pups during the period of lactation when the mother received doxycycline in the drinking water. In the CCSP-rtTA mice, luciferase was not detected in the absence of doxycycline. In the SP-C-rtTA mice, luciferase activity was detected in the absence of doxycycline but was enhanced approximately 10-fold by administration of drugs. The SP-C-rtTA and CCSP-rtTA activator mice control the expression of transgenes in the developing and mature respiratory epithelium, and will be useful for the study of gene function in the lung.

Animals↗

PubMed2006

Prenatal alcohol exposure alters GABA(A)alpha5 expression: a mechanism of alcohol-induced learning dysfunction.

OBJECTIVE: In a model for fetal alcohol syndrome (FAS), we have previously found an alteration in NMDA receptors suggesting mediation, at least in part, of alcohol-related learning deficit. NMDA and GABA receptors interact in a multisynaptic circuit for the regulation of the inhibitory tone through the CNS. The GABA receptor subunit GABA(A)alpha5 is involved in learning and is developmentally regulated, as it is excitatory in the perinatal brain and inhibitory in the adult. We were interested to evaluate alcohol's effect on GABA(A)alpha5 expression to further understand alcohol-induced learning dysfunction. STUDY DESIGN: Timed, pregnant C57B16/J mice were treated on gestational day 8 with alcohol (25% alcohol, 0.03 mL/kg i.p.) or control (saline). Embryos and brains were harvested 10 days after treatment, and brains from adult offspring were collected after evaluation in the Morris Water Maze, a well-established test for spatial learning. Gene expression included samples from at least 3 litters per timepoint, and calibrator-normalized relative real-time polymerase chain reaction (PCR) was performed to quantify GABA(A)alpha5 with GAPDH standardization. Statistical analysis included analysis of variance (ANOVA). RESULTS: Prenatal alcohol exposure significantly decreased GABA(A)alpha5 expression in the embryo (P < .02) and fetal brains (P < .01) 10 days after therapy. However, in adult brains GABA(A)alpha5 expression was increased versus controls (P < .01). As previously demonstrated, prenatal alcohol exposure resulted in deficits in adults learning the Morris Water Maze with controls learning faster (P < .05). CONCLUSION: Prenatal alcohol exposure alters developmental GABA(A)alpha5 expression. This may further explain the long-lasting damage of alcohol on learning skills. Both the alcohol-induced reduction in the GABA(A)alpha5 subunit during development and up-regulation in adult brain may be related to learning deficits resulting in decreased learning potential caused by the developmental defect and an increased inhibition of learning resulting from increased expression as an adult. In combination with our previous findings, these suggest that alcohol-induced learning impairment is likely the result of alterations of both NMDA and GABA expression and function.

Animals↗

PubMed1999

Brain insulin receptors and spatial memory. Correlated changes in gene expression, tyrosine phosphorylation, and signaling molecules in the hippocampus of water maze trained rats.

Evidence accumulated from clinical and basic research has indirectly implicated the insulin receptor (IR) in brain cognitive functions, including learning and memory (Wickelgren, I. (1998) Science 280, 517-519). The present study investigates correlative changes in IR expression, phosphorylation, and associated signaling molecules in the rat hippocampus following water maze training. Although the distribution of IR protein matched that of IR mRNA in most forebrain regions, a dissociation of the IR mRNA and protein expression patterns was found in the cerebellar cortex. After training, IR mRNA in the CA1 and dentate gyrus of the hippocampus was up-regulated, and there was increased accumulation of IR protein in the hippocampal crude synaptic membrane fraction. In the CA1 pyramidal neurons, changes in the distribution pattern of IR in particular cellular compartments, such as the nucleus and dendritic regions, was observed only in trained animals. Although IR showed a low level of in vivo tyrosine phosphorylation, an insulin-stimulated increase of in vitro Tyr phosphorylation of IR was detected in trained animals, suggesting that learning may induce IR functional changes, such as enhanced receptor sensitivity. Furthermore, a training-induced co-immunoprecipitation of IR with Shc-66 was detected, along with changes in in vivo Tyr phosphorylation of Shc and mitogen-activated protein kinase, as well as accumulation of Shc-66, Shc-52, and Grb-2 in hippocampal synaptic membrane fractions following training. These findings suggest that IR may participate in memory processing through activation of its receptor Tyr kinase activity, and they suggest possible engagement of Shc/Grb-2/Ras/mitogen-activated protein kinase cascades.

3T3 Cells↗

PubMed2001

PDE1 encodes a P-type ATPase involved in appressorium-mediated plant infection by the rice blast fungus Magnaporthe grisea.

Plant infection by the rice blast fungus Magnaporthe grisea is brought about by the action of specialized infection cells called appressoria. These infection cells generate enormous turgor pressure, which is translated into an invasive force that allows a narrow penetration hypha to breach the plant cuticle. The Magnaporthe pde1 mutant was identified previously by restriction enzyme-mediated DNA integration mutagenesis and is impaired in its ability to elaborate penetration hyphae. Here we report that the pde1 mutation is the result of an insertion into the promoter of a P-type ATPase-encoding gene. Targeted gene disruption confirmed the role of PDE1 in penetration hypha development and pathogenicity but highlighted potential differences in PDE1 regulation in different Magnaporthe strains. The predicted PDE1 gene product was most similar to members of the aminophospholipid translocase group of P-type ATPases and was shown to be a functional homolog of the yeast ATPase gene ATC8. Spatial expression studies showed that PDE1 is expressed in germinating conidia and developing appressoria. These findings implicate the action of aminophospholipid translocases in the development of penetration hyphae and the proliferation of the fungus beyond colonization of the first epidermal cell.

Adenosine Triphosphatases↗

PubMed2002

It takes guts: the Drosophila hindgut as a model system for organogenesis.

The Drosophila hindgut is fruitful territory for investigation of events common to many types of organogenesis. The development of the Drosophila hindgut provides, in microcosm, a genetic model system for studying processes such as establishment (patterning) of an epithelial primordium, its internalization by gastrulation, development of left--right asymmetric looping, patterning in both the anteroposterior and dorsoventral axes, innervation, investment of an epithelium with mesoderm, reciprocal epitheliomesenchymal interactions, cell shape change, and cell rearrangement. We review the genetic control of these processes during development of the Drosophila hindgut, and compare these to related processes in other bilaterians, particularly vertebrates. We propose that caudal/Cdx, brachyenteron/Brachyury, fork head/HNF-3, and wingless/Wnt constitute a conserved "cassette" of genes expressed in the blastopore and later in the gut, involved in posterior patterning, cell rearrangement, and gut maintenance. Elongation of the internalized Drosophila hindgut primordium is similar to elongation of the archenteron and also of the entire embryonic axis (both during and after gastrulation), as well as of various tubules (e.g., nephric ducts, Malpighian tubules), as it is driven by cell rearrangement. The genes drumstick, bowl, and lines (which encode putative transcriptional regulators) are required for this cell rearrangement, as well as for spatially localized gene expression required to establish the three morphologically distinct subregions of the hindgut. Expression of signaling molecules regulated by drumstick, bowl, and lines, in particular of the JAK/STAT activator Unpaired at the hindgut anterior, may play a role in controlling hindgut cell rearrangement. Other cell signaling molecules expressed in the hindgut epithelium are required to establish its normal size (Dpp and Hh), and to establish and maintain the hindgut visceral mesoderm (Wg and Hh). Both maternal gene activity and zygotic gene activity are required for asymmetric left--right looping of the hindgut. Some of the same genes (caudal and brachyenteron) required for embryonic hindgut development also act during pupation to construct a new hindgut from imaginal cells. Application of the plethora of genetic techniques available in Drosophila, including forward genetic screens, should identify additional genes controlling hindgut development and thus shed light on a variety of common morphogenetic processes.

Animals↗

PubMed1992

Individual stage selector element mutations lead to reciprocal changes in beta- vs. epsilon-globin gene transcription: genetic confirmation of promoter competition during globin gene switching.

Biochemical and genetic analysis of the embryonic to adult beta-like globin gene switch in chickens has led to the hypothesis that competition between the promoters of the cis-linked epsilon- and beta-globin genes for interaction with a shared enhancer mediates the developmental changes in expression of beta-globin protein isotypes. To test specific predictions of this promoter competition model, a sensitive RNA/polymerase chain reaction assay has been used to investigate the effects of individual beta-globin promoter mutations on expression of the two linked genes in transiently transfected erythroid cells. Mutations that attenuated adult beta-globin transcription resulted concomitantly in a proportional increase in expression of the embryonic epsilon-globin gene. Consistent with the model, mutations disrupting the binding sites for either of two adult stage-specific transcription factors (NF-E4 and beta CTF) indicate that these sites are essential both for induction of beta-globin gene expression and for indirect suppression (through promoter competition) of epsilon-globin transcription in definitive (adult) erythroid cells. These results provide direct evidence that stage-specific transcription factors affect the equilibrium existing between multiple interacting globin cis-regulatory elements. We conclude that promoter competition is an important mechanism through which developmental regulation of chicken beta-globin gene switching is achieved and that such competitive interactions may prove to be generally applicable to the regulation of a variety of other temporally or spatially restricted gene expression patterns.

Animals↗

PubMed1995

Regulation of the acetylcholine receptor epsilon subunit gene by recombinant ARIA: an in vitro model for transynaptic gene regulation.

Structural specialization of the postsynaptic skeletal muscle membrane is in part mediated by the motor neuron-induced transcriptional regulation of synaptic muscle nuclei. ARIA, a factor that stimulates production of acetylcholine receptors (AChRs), is a candidate signaling molecule for such regulation. Here we examine the transynaptic inducing potential of this polypeptide factor. ARIA immunoreactivity is detectable at synaptic sites in vivo. In vitro, recombinant heregulin beta 1 (rHRG beta 1), the human homolog of ARIA, induces expression of the AChR epsilon gene, the subunit most sensitive to synaptic input. The inducing property of rHRG beta 1 is demonstrated most dramatically in primary muscle cultures from transgenic mice bearing an epsilon promoter-nuclear lacZ reporter transgene. Transient transfection experiments using the Sol 8 muscle cell line indicate that sequences that confer responsiveness to ARIA are located within a 150 bp epsilon subunit promoter region and are E box-independent. These results suggest that ARIA performs a vital role by directing spatially restricted gene expression at the neuromuscular junction.

Animals↗

PubMed2004

Hox cluster disintegration with persistent anteroposterior order of expression in Oikopleura dioica.

Tunicate embryos and larvae have small cell numbers and simple anatomical features in comparison with other chordates, including vertebrates. Although they branch near the base of chordate phylogenetic trees, their degree of divergence from the common chordate ancestor remains difficult to evaluate. Here we show that the tunicate Oikopleura dioica has a complement of nine Hox genes in which all central genes are lacking but a full vertebrate-like set of posterior genes is present. In contrast to all bilaterians studied so far, Hox genes are not clustered in the Oikopleura genome. Their expression occurs mostly in the tail, with some tissue preference, and a strong partition of expression domains in the nerve cord, in the notochord and in the muscle. In each tissue of the tail, the anteroposterior order of Hox gene expression evokes spatial collinearity, with several alterations. We propose a relationship between the Hox cluster breakdown, the separation of Hox expression domains, and a transition to a determinative mode of development.

Animals↗

PubMed2001

Regulation of gene transcription in the epididymis.

The epididymis exhibits region-specific as well as cell-specific patterns of gene expression within the epithelium. The spatial and temporal patterns of gene expression originate during development and are critical to the formation and maintenance of a fully functional epididymis. Despite the number of mechanisms reported to contribute to the regulation of eukaryotic gene expression, little is known about the specific mechanisms involved in the control of epididymal gene expression. This review will outline some of the cis-DNA elements and associated transcription factors that have been identified in the epididymis, in addition to discussing the potential role of co-regulator molecules and changes in chromatin structure as critical control points of gene expression. Although gene expression can be controlled at several points, discussion will focus on gene regulation at the transcriptional level. The role of post-transcriptional control, with particular attention to mRNA stability, will also be discussed.

Animals↗

PubMed2026

A High-Resolution Stereo-Seq Spatial Transcriptomic Resource for Adult Holstein Cattle Liver.

The bovine liver is a highly compartmentalized organ that plays essential roles in continuous gluconeogenesis and nitrogen recycling; however, its spatial molecular architecture has remained largely uncharacterized due to the limitations of traditional bulk and single-cell approaches. To address this gap, Spatial Enhanced Resolution Omics-sequencing (Stereo-seq) was utilized to generate a subcellular-resolution (500 nm) transcriptomic map of an adult Holstein cattle liver, and a refined reference-guided workflow was implemented to overcome standard annotation limitations in livestock. Raw sequencing data were processed using the Stereo-seq Analysis Workflow and analyzed with Stereopy, Seurat, SingleR, and reference-guided workflows. Spatial aggregation was evaluated at Bin20, Bin50, Bin100, Bin150, and Bin200. Increasing bin size increased molecular identifier counts and detected-gene complexity while progressively reducing spatial granularity. Bin50, corresponding to 50 × 50 DNA nanoballs and an approximate nominal footprint of 25 × 25 µm, was therefore selected as a practical intermediate aggregation level for the primary analyses. Quality-control assessment, Leiden clustering, UMAP visualization, reference-based cell-type annotation, cluster-marker analysis, and spatial mapping of canonical hepatic genes demonstrated preservation of biologically interpretable liver transcriptional organization. Raw sequencing data processed spatial matrices, annotated objects, and analysis code are publicly available to support reanalysis and computational benchmarking. In summary, we present a Stereo-seq spatial transcriptomic resource generated from liver tissue of an adult Holstein cow. This initial resource provides a valuable foundation for future studies of bovine liver biology, comparative genomics, and the spatial basis of livestock health and production traits.

Animals↗

PubMed2004

Rat tyrosine hydroxylase promoter directs tetracycline-inducible foreign gene expression in dopaminergic cell types.

A prerequisite for creating animal models in which gene expression is spatially and temporally controlled is the development of promoters to target genetic switches to specific populations of cells. Here we used the dopaminergic biosynthetic enzyme, tyrosine hydroxylase (TH) to test various combinations of tetracycline (Tet) system elements to determine the optimal configuration for inducible, tissue-specific expression. The present study shows that the degree of expression and level of leakiness associated with the Tet transactivators rtTA, rtTA2S-M2, tTS/rtTA or tTS/rtTA2S-M2 was dependent upon both the promoter and cell type utilized. Specifically, CMV-driven tTS/rtTA2S-M2 exhibited the highest level of inducibility in HEK cells (approximately 1000-fold) versus the dopaminergic cell line, MN9D (approximately 70-fold). In contrast, TH-driven rtTA2S-M2 yielded the highest level of expression with the least background in dopaminergic cell types versus HEK cells. Moreover, the TH promoter could be combined with the bi-directional Tet response system, BiTetO, allowing for the co-expression and regulation of two genes in the same cell. To further test the feasibility of this system we replaced the reporter gene with human Bcl-2. Consistent with previous studies, induction of Bcl-2 expression in dopaminergic cell types attenuated cell death due to the neurotoxin, MPP+. Taken together, these data suggest that targeted, inducible gene expression can be achieved in dopaminergic cell types.

Animals↗

PubMed1986

Spatial regulation of Antennapedia and bithorax gene expression by the Polycomb locus in Drosophila.

Transcripts encoded by six of the homeotic lethal complementation groups within the Antennapedia and bithorax gene complexes (ANT-C and BX-C) accumulate primarily in nonoverlapping regions of the embryonic central nervous system (CNS). It has been previously proposed that the maintenance of these selective patterns of expression involve hierarchical cross-regulatory interactions among ANT-C and BX-C loci. Selective expression of the ANT-C and BX-C also involves regulation by a number of unlinked genetic loci, including the Polycomb (Pc) locus. Here we show that the ANT-C and BX-C transcript distribution patterns are dramatically altered in the CNS of Pc- embryos. We discuss these altered patterns of expression in the context of different models for the spatial regulation of ANT-C and BX-C gene expression.

Animals↗