Biomedical subjects

Michael Levine

Publications and source records attributed to Michael Levine.

52 records · Page 3Linked to original sources

PubMed2002

The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins.

The first chordates appear in the fossil record at the time of the Cambrian explosion, nearly 550 million years ago. The modern ascidian tadpole represents a plausible approximation to these ancestral chordates. To illuminate the origins of chordate and vertebrates, we generated a draft of the protein-coding portion of the genome of the most studied ascidian, Ciona intestinalis. The Ciona genome contains approximately 16,000 protein-coding genes, similar to the number in other invertebrates, but only half that found in vertebrates. Vertebrate gene families are typically found in simplified form in Ciona, suggesting that ascidians contain the basic ancestral complement of genes involved in cell signaling and development. The ascidian genome has also acquired a number of lineage-specific innovations, including a group of genes engaged in cellulose metabolism that are related to those in bacteria and fungi.

Alleles↗

PubMed2002

Characterization of the intergenic RNA profile at abdominal-A and Abdominal-B in the Drosophila bithorax complex.

The correct spatial expression of two Drosophila bithorax complex (BX-C) genes, abdominal-A (abdA) and Abdominal-B (AbdB), is dependent on the 100-kb intergenic infraabdominal (iab) region. The iab region is known to contain a number of different domains (iab2 through iab8) that harbor cis-regulatory elements responsible for directing expression of abdA and AbdB in the second through eighth abdominal segments. Here, we use in situ hybridization to perform high-resolution mapping of the transcriptional activity in the iab control regions. We show that transcription of the control regions themselves is abundant and precedes activation of the abdA and AbdB genes. As with the homeotic genes of the BX-C, the transcription patterns of the RNAs from the iab control regions demonstrate colinearity with the sequence of the iab regions along the chromosome and the domains in the embryo under the control of the specific iab regions. These observations suggest that the intergenic RNAs may play a role in initiating cis regulation at the BX-C early in development.

Animals↗

PubMed2002

Whole-genome analysis of dorsal-ventral patterning in the Drosophila embryo.

The maternal Dorsal regulatory gradient initiates the differentiation of several tissues in the early Drosophila embryo. Whole-genome microarray assays identified as many as 40 new Dorsal target genes, which encode a broad spectrum of cell signaling proteins and transcription factors. Evidence is presented that a tissue-specific form of the NF-Y transcription complex is essential for the activation of gene expression in the mesoderm. Tissue-specific enhancers were identified for new Dorsal target genes, and bioinformatics methods identified conserved cis-regulatory elements for coordinately regulated genes that respond to similar thresholds of the Dorsal gradient. The new Dorsal target genes and enhancers represent one of the most extensive gene networks known for any developmental process.

Animals↗

PubMed2002

Promoter-proximal tethering elements regulate enhancer-promoter specificity in the Drosophila Antennapedia complex.

Insulator DNAs and promoter competition regulate enhancer-promoter interactions within complex genetic loci. Here we provide evidence for a third mechanism: promoter-proximal tethering elements. The Scr-ftz region of the Antennapedia gene complex includes two known enhancers, AE1 and T1. AE1 selectively interacts with the ftz promoter to maintain pair-rule stripes of ftz expression during gastrulation and germ-band elongation. The T1 enhancer, located 3' of the ftz gene and approximately 25 kb 5' of the Scr promoter, selectively activates Scr expression in the prothorax and posterior head segments. A variety of P element minigenes were examined in transgenic embryos to determine the basis for specific AE1-ftz and T1-Scr interactions. A 450-bp DNA fragment located approximately 100 bp 5' of the Scr transcription start site is essential for T1-Scr interactions and can mediate long-range activation of a ftz/lacZ reporter gene when placed 5' of the ftz promoter. We suggest that the Scr450 fragment contains tethering elements that selectively recruit T1 to the Scr promoter. Tethering elements might regulate enhancer-promoter interactions at other complex genetic loci.

Animals↗

PubMed2002

Control of intercalation is cell-autonomous in the notochord of Ciona intestinalis.

Dishevelled signaling plays a critical role in the control of cell intercalation during convergent extension in vertebrates. This study presents evidence that Dishevelled serves a similar function in the Ciona notochord. Embryos transgenic for mutant Dishevelled fail to elongate their tails, and notochord cells fail to intercalate, though notochord cell fates are unaffected. Analysis of mosaic transgenics revealed that the effects of mutant Dishevelled on notochord intercalation are cell-autonomous in Ciona, though such defects have nonautonomous effects in Xenopus. Furthermore, our data indicate that notochord cell intercalation in Ciona does not require the progressive signals which coordinate cell intercalation in the Xenopus notochord, highlighting an important difference in how mediolateral cell intercalation is controlled in the two animals. Finally, this study establishes the Ciona embryo as an effective in vivo system for the study of the molecular control of morphogenetic cell movements in chordates.

Animals↗

PubMed2002

Dorsal gradient networks in the Drosophila embryo.

Here, we describe one of the major maternal regulatory gradients, Dorsal, and threshold outputs of gene expression that result from the graded distribution of this transcription factor. The analysis of a large number of authentic and synthetic target genes suggests that the Dorsal gradient directly specifies at least four, and possibly as many as seven, different thresholds of gene activity and tissue differentiation. These thresholds initiate the differentiation of the three primary embryonic tissues: the mesoderm, neurogenic ectoderm, and dorsal ectoderm. Moreover, primary readouts of the Dorsal gradient create asymmetries that subdivide each tissue into multiple cell types during gastrulation. Dorsal patterning thresholds represent the culmination of one of the most complete gene regulation network known in development, which begins with the asymmetric positioning of the oocyte nucleus within the egg chamber and leads to the localized activation of the Toll-Dorsal signaling pathway in ventral regions of the early embryo.

Animals↗

PubMed2002

Genome-wide identification of tissue-specific enhancers in the Ciona tadpole.

Less than 100 cis-regulatory DNAs have been characterized in the context of transgenic metazoan embryos. Here we investigate the feasibility of conducting a genome-wide search for tissue-specific enhancers in the ascidian Ciona intestinalis. A total of 138 random genomic DNA fragments with an average size of 1.7 kb were separately placed 5' of a lacZ reporter gene. Eleven of the lacZ fusion genes displayed localized patterns of expression in tadpole-stage Ciona embryos. At least five of these transgenes appear to contain bona fide tissue-specific enhancers that direct expression in the cerebral vesicle, neural tube, primordial adhesive organ, notochord, and tail epidermis. One of the enhancers maps near Distalless (Ci-Dll-A) and recapitulates most aspects of the endogenous expression pattern, including localized expression in the anterior-most regions of the neurogenic ectoderm. We discuss the prospects of creating a regulatory atlas of the Ciona genome, whereby every enhancer is identified for every gene.

Animals↗

PubMed2002

Tail morphogenesis in the ascidian, Ciona intestinalis, requires cooperation between notochord and muscle.

We present evidence that notochord and muscle differentiation are crucial for morphogenesis of the ascidian tail. We developed a novel approach for embryological manipulation of the developing larval tissues using a simple method to introduce DNA into Ciona intestinalis and the several available tissue-specific promoters. With such promoters, we misexpressed the Xenopus homeobox gene bix in notochord or muscle of Ciona embryos as a means of interfering with development of these tissues. Ciona embryos expressing bix in the notochord from the 64-cell stage develop into larvae with very short tails, in which the notochord precursors fail to intercalate and differentiate. Larvae with mosaic expression of bix have intermediate phenotypes, in which a partial notochord is formed by the precursor cells that did not receive the transgene while the precursors that express the transgene cluster together and fail to undergo any of the cell-shape changes associated with notochord differentiation. Muscle cells adjacent to differentiated notochord cells are properly patterned, while those next to the notochord precursor cells transformed by bix exhibit various patterning defects. In these embryos, the neural tube extends in the tail to form a nerve cord, while the endodermal strand fails to enter the tail region. Similarly, expression of bix in muscle progenitors impairs differentiation of muscle cells, and as a result, notochord cells fail to undergo normal extension movements. Hence, these larvae have a shorter tail, due to a block in the elongation of the notochord. Taken together, these observations suggest that tail formation in ascidian larvae requires not only signaling from notochord to muscle cells, but also a "retrograde" signal from muscle cells to notochord.

Animals↗

PubMed2002

Exploiting transcription factor binding site clustering to identify cis-regulatory modules involved in pattern formation in the Drosophila genome.

A major challenge in interpreting genome sequences is understanding how the genome encodes the information that specifies when and where a gene will be expressed. The first step in this process is the identification of regions of the genome that contain regulatory information. In higher eukaryotes, this cis-regulatory information is organized into modular units [cis-regulatory modules (CRMs)] of a few hundred base pairs. A common feature of these cis-regulatory modules is the presence of multiple binding sites for multiple transcription factors. Here, we evaluate the extent to which the tendency for transcription factor binding sites to be clustered can be used as the basis for the computational identification of cis-regulatory modules. By using published DNA binding specificity data for five transcription factors active in the early Drosophila embryo, we identified genomic regions containing unusually high concentrations of predicted binding sites for these factors. A significant fraction of these binding site clusters overlap known CRMs that are regulated by these factors. In addition, many of the remaining clusters are adjacent to genes expressed in a pattern characteristic of genes regulated by these factors. We tested one of the newly identified clusters, mapping upstream of the gap gene giant (gt), and show that it acts as an enhancer that recapitulates the posterior expression pattern of gt.

Animals↗

PubMed2002

Decoding cis-regulatory DNAs in the Drosophila genome.

Cis-regulatory DNAs control the timing and sites of gene expression during metazoan development. Changes in gene expression are responsible for the morphological diversification of metazoan body plans. However, traditional methods for the identification and characterization of cis-regulatory DNAs are tedious. During the past year, computational methods have been used to identify novel cis-DNAs within the entire Drosophila genome. These methods change the way that cis-DNAs will be analyzed in future studies and offer the promise of unraveling complex gene networks.

Animals↗

PubMed2002

Whole-genome expression profiles identify gene batteries in Drosophila.

Microarray studies make it possible to obtain gene expression data on a whole-genome scale. Arbeitman et al. present microarray data generated at multiple time points throughout Drosophila melanogaster development, and identify new genes engaged in a broad spectrum of processes, including the patterning of the early embryo and senescence in adults.

Animals↗

PubMed2002

Analyzing gene regulation in ascidian embryos: new tools for new perspectives.

Ascidians are marine protochordates at the evolutionary boundary between invertebrates and vertebrates. Ascidian larvae provide a simple system for unraveling gene regulation networks underlying the formation of the basic chordate body plan. After being used for over a century as a model for embryological studies, ascidians have become, in the past decade, an increasingly popular organism for studying gene regulation. Part of the renewed appeal of this system is the use of electroporation to introduce transgenic DNAs into developing embryos. This method is considerably more efficient than conventional microinjection assays and permits the simultaneous transformation of hundreds of embryos. Electroporation has allowed the identification and characterization of cis-regulatory DNAs that mediate gene expression in a variety of tissues, including the notochord, tail muscles, CNS, and endoderm. Electroporation has also provided a simple method for misexpressing patterning genes and producing dominant mutant phenotypes. Recent studies have used electroporation to create "knock-out" phenotypes by overexpressing dominant negative forms of particular proteins. Here we review the past and present uses of electroporation in ascidian development, and speculate on potential future uses.

Animals↗

PubMed2002

Linear signaling in the Toll-Dorsal pathway of Drosophila: activated Pelle kinase specifies all threshold outputs of gene expression while the bHLH protein Twist specifies a subset.

Differential activation of the Toll receptor leads to the formation of a broad Dorsal nuclear gradient that specifies at least three patterning thresholds of gene activity along the dorsoventral axis of precellular embryos. We investigate the activities of the Pelle kinase and Twist basic helix-loop-helix (bHLH) transcription factor in transducing Toll signaling. Pelle functions downstream of Toll to release Dorsal from the Cactus inhibitor. Twist is an immediate-early gene that is activated upon entry of Dorsal into nuclei. Transgenes misexpressing Pelle and Twist were introduced into different mutant backgrounds and the patterning activities were visualized using various target genes that respond to different thresholds of Toll-Dorsal signaling. These studies suggest that an anteroposterior gradient of Pelle kinase activity is sufficient to generate all known Toll-Dorsal patterning thresholds and that Twist can function as a gradient morphogen to establish at least two distinct dorsoventral patterning thresholds. We discuss how the Dorsal gradient system can be modified during metazoan evolution and conclude that Dorsal-Twist interactions are distinct from the interplay between Bicoid and Hunchback, which pattern the anteroposterior axis.

Animals↗

PubMed2002

The Snail repressor positions Notch signaling in the Drosophila embryo.

The maternal Dorsal nuclear gradient initiates the differentiation of the mesoderm, neurogenic ectoderm and dorsal ectoderm in the precellular Drosophila embryo. Each tissue is subsequently subdivided into multiple cell types during gastrulation. We have investigated the formation of the mesectoderm within the ventral-most region of the neurogenic ectoderm. Previous studies suggest that the Dorsal gradient works in concert with Notch signaling to specify the mesectoderm through the activation of the regulatory gene sim within single lines of cells that straddle the presumptive mesoderm. This model was confirmed by misexpressing a constitutively activated form of the Notch receptor, Notch(IC), in transgenic embryos using the eve stripe2 enhancer. The Notch(IC) stripe induces ectopic expression of sim in the neurogenic ectoderm where there are low levels of the Dorsal gradient. sim is not activated in the ventral mesoderm, due to inhibition by the localized zinc-finger Snail repressor, which is selectively expressed in the ventral mesoderm. Additional studies suggest that the Snail repressor can also stimulate Notch signaling. A stripe2-snail transgene appears to induce Notch signaling in 'naïve' embryos that contain low uniform levels of Dorsal. We suggest that these dual activities of Snail, repression of Notch target genes and stimulation of Notch signaling, help define precise lines of sim expression within the neurogenic ectoderm.

Animals↗

PubMedDate not supplied

Concordance of field and emergency department assessment in the prehospital management of patients with dyspnea.

OBJECTIVE: Dyspnea is a common complaint of patients treated by emergency medical services (EMS). Few studies have examined the ability of paramedics to distinguish between etiologies of dyspnea. The authors evaluated the degree of agreement related to cardiac versus noncardiac sources of dyspnea between field and emergency department (ED) assessment of patients transported at the advanced life support level. METHODS: This was a retrospective, cohort study of consecutive patients aged > or =35 years transported by paramedics with dyspnea. The authors compared the concordance between the EMS and ED diagnoses. They also investigated whether patients whose assessments were discordant had worse outcomes. RESULTS: Paramedics correctly assessed the cause of dyspnea in 172 of 222 (77%) patients (kappa=0.60; 95% confidence interval [CI]=0.51, 0.69). Among single-source (i.e., cardiac or noncardiac) dyspnea patients, prehospital providers correctly assessed 70 of 84 (83%) noncardiac causes and 98 of 114 (86%) cardiac causes (kappa=0.69; 95% CI=0.59, 0.79). When the ED diagnosis included both cardiac and noncardiac etiologies, paramedics treated seven of 24 (29%) patients as noncardiac, 13 of 24 (54%) as cardiac, and four of 24 (17%) as combined-source dyspnea. The authors did not observe any statistically significant differences in in-hospital mortality, intubation frequency, or hospital length of stay in patients whose prehospital dyspnea diagnosis was discordant. CONCLUSION: The authors conclude that in this EMS system, field assessment of dyspnea by paramedics is in agreement with that arrived at in the ED in a high proportion of patients with dyspnea from a single source. However, field assessment of dyspnea from multiple etiologies is less concordant.

Adult↗

PubMedDate not supplied

A swollen joint: why all the fuss?

Acute arthritis may be a potential medical emergency. An infected joint causes rapid cartilaginous destruction and risk of future osteoarthritis. Prompt attention to the historical clues and potential causative organisms ensures appropriate therapy. Numerous microbes have been identified as the causative agent in septic arthritis, and various populations have distinct susceptibilities to these specific organisms. One broad classification of septic arthritis is differentiating gonococcal and nongonococcal organisms. This classification is important, as not only do the organisms differ, but the age of the patient and the portal of entry also differ. Aspiration of synovial fluid is paramount for proper diagnosis and management of septic arthritis. In addition, the timely administration of properly chosen antibiotic agents is essential for reducing morbidity and mortality.

Anti-Bacterial Agents↗