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1.  MicroRNA-205 Controls Neonatal Expansion of Skin Stem Cells by Modulating the PI3K Pathway 
Nature cell biology  2013;15(10):1153-1163.
Skin stem cells (SCs) are specified and rapidly expanded to fuel body growth during early development. However, molecular mechanisms that govern the amplification of skin SCs remain unclear. Here we report an essential role for miR-205, one of the most highly expressed miRNAs in skin SCs, in promoting neonatal expansion of these cells. Unlike most mammalian miRNAs, genetic deletion of miR-205 causes neonatal lethality with severely compromised epidermal and hair follicle growth. In the miR-205 KO skin SCs, phospho-Akt is significantly downregulated, and the SCs prematurely exit the cell cycle. In the hair follicle, this accelerates the transition of the neonatal SCs towards quiescence. We identify multiple miR-205 targeted negative regulators of PI3K signaling that mediate the repression of phospho-Akt and restrict the proliferation of SCs. Our findings reveal an essential role for miR-205 in maintaining the expansion of skin SCs by antagonizing negative regulators of PI3K signaling.
PMCID: PMC3789848  PMID: 23974039
2.  Homeostatic control of Argonaute stability by microRNA availability 
Homeostatic mechanisms regulate the abundance of many small RNA components. We used Drosophila and mammalian systems to demonstrate a conserved homeostatic system in which the status of miRNA biogenesis controls Argonaute protein stability. Clonal analyses of multiple mutants of core Drosophila miRNA factors revealed that stability of the miRNA effector AGO1 is dependent on miRNA biogenesis. Reciprocally, ectopic transcription of miRNAs within in vivo clones induced accumulation of AGO1, as did genetic interference with the ubiquitin-proteasome system. In mammals, we found that the stability of mAgo2 declined in Dicer knockout cells, and was rescued by proteasome blockade or introduction of either Dicer plasmid or Dicer-independent miRNA constructs. Importantly, Dicer-dependent miRNA constructs generated pre-miRNAs that bind Ago2, but did not rescue Ago2 stability. We conclude that Argonaute levels are finely tuned by cellular availability of mature miRNAs and the ubiquitin-proteasome system.
PMCID: PMC3702675  PMID: 23708604
3.  Intertwined pathways for Argonaute-mediated microRNA biogenesis in Drosophila 
Nucleic Acids Research  2013;42(3):1987-2002.
Although Dicer is essential for general microRNA (miRNA) biogenesis, vertebrate mir-451 is Dicer independent. Instead, its short pre-miRNA hairpin is ‘sliced’ by Ago2, then 3′-resected into mature miRNAs. Here, we show that Drosophila cells and animals generate functional small RNAs from mir-451-type precursors. However, their bulk maturation arrests as Ago-cleaved pre-miRNAs, which mostly associate with the RNAi effector AGO2. Routing of pre-mir-451 hairpins to the miRNA effector AGO1 was inhibited by Dicer-1 and its partner Loqs. Loss of these miRNA factors promoted association of pre-mir-451 with AGO1, which sliced them and permitted maturation into ∼23–26 nt products. The difference was due to the 3′ modification of single-stranded species in AGO2 by Hen1 methyltransferase, whose depletion permitted 3′ trimming of Ago-cleaved pre-miRNAs in AGO2. Surprisingly, Nibbler, a 3′–5′ exoribonuclease that trims ‘long’ mature miRNAs in AGO1, antagonized miR-451 processing. We used an in vitro reconstitution assay to identify a soluble, EDTA-sensitive activity that resects sliced pre-miRNAs in AGO1 complexes. Finally, we use deep sequencing to show that depletion of dicer-1 increases the diversity of small RNAs in AGO1, including some candidate mir-451-like loci. Altogether, we document unexpected aspects of miRNA biogenesis and Ago sorting, and provide insights into maturation of Argonaute-cleaved miRNA substrates.
PMCID: PMC3919586  PMID: 24220090
4.  Drosophila Argonaute 1 and its miRNA biogenesis partners are required for oocyte formation and germline cell division 
Developmental biology  2012;365(2):384-394.
Argonaute 1 (Ago1) is a member of the Argonaute/PIWI protein family involved in small RNA-mediated gene regulation. In Drosophila, Ago1 plays a specific role in microRNA (miRNA) biogenesis and function. Previous studies have demonstrated that Ago1 regulates the fate of germline stem cells. However, the function of Ago1 in other aspects of oogenesis is still elusive. Here we report the function of Ago1 in developing egg chambers. We find that Ago1 protein is enriched in the oocytes and also highly expressed in the cytoplasm of follicle cells. Clonal analysis of multiple ago1 mutant alleles shows that many mutant egg chambers contain only 8 nurse cells without an oocyte which is phenocopied in dicer-1, pasha and drosha mutants. Our results suggest that Ago1 and its miRNA biogenesis partners play a role in oocyte determination and germline cell division in Drosophila.
PMCID: PMC3763516  PMID: 22445511
5.  The evolution and functional diversification of animal microRNA genes 
Cell research  2008;18(10):985-996.
microRNAs (miRNAs) are an abundant class of ~22 nucleotide (nt) regulatory RNAs that are pervasive in higher eukaryotic genomes. In order to fully understand their prominence in genomes, it is necessary to elucidate the molecular mechanisms that can diversify miRNA activities. In this review, we describe some of the many strategies that allow novel miRNA functions to emerge, with particular emphasis on how miRNA genes evolve in animals. These mechanisms include changes in their sequence, processing, or expression pattern; acquisition of miRNA* functionality or antisense processing; and de novo gene birth. The facility and versatility of miRNAs to evolve and change likely underlies how they have become dominant constituents of higher genomes.
PMCID: PMC2712117  PMID: 18711447
microRNA; evolution; subfunctionalization; neofunctionalization
6.  Dicer-independent, Ago2-mediated microRNA biogenesis in vertebrates 
Cell Cycle  2010;9(22):4455-4460.
A canonical biogenesis pathway involving sequential cleavage by the Drosha and Dicer RNAse III enzymes governs the maturation of most animal microRNAs. However, there exist a variety of alternative miRNA biogenesis pathways, most of which bypass Drosha processing. Recently, three groups described for the first time a vertebrate microRNA pathway that bypasses Dicer cleavage. This mechanism was characterized with respect to the highly conserved vertebrate gene mir-451, for which Drosha processing yields a short (42 nucleotide) hairpin that is directly loaded into Ago2, the sole vertebrate “Slicer” Argonaute. Ago2-mediated cleavage of this hairpin yields a 30 nucleotide intermediate, whose 3′ end is resected to generate the dominantly cloned ∼23 nucleotide mature miR-451. Knowledge of this pathway provides an unprecedented tool with which to express microRNAs and small interfering RNAs in Dicer mutant cells. More generally, the mir-451 backbone constitutes a new platform for gene silencing that complements existing shRNA technology.
PMCID: PMC3048044  PMID: 21088485
mir-451; Ago2; Slicer; Dicer-independent; erythropoiesis
7.  Evolution of mir-92a Underlies Natural Morphological Variation in Drosophila melanogaster 
Current Biology  2013;23(6):523-528.
Identifying the genetic mechanisms underlying phenotypic change is essential to understanding how gene regulatory networks and ultimately the genotype-to-phenotype map evolve. It is recognized that microRNAs (miRNAs) have the potential to facilitate evolutionary change [1–3]; however, there are no known examples of natural morphological variation caused by evolutionary changes in miRNA expression. Therefore, the contribution of miRNAs to evolutionary change remains unknown [1, 4]. Drosophila melanogaster subgroup species display a portion of trichome-free cuticle on the femur of the second leg called the “naked valley.” It was previously shown that Ultrabithorax (Ubx) is involved in naked valley variation between D. melanogaster and D. simulans [5, 6]. However, naked valley size also varies among populations of D. melanogaster, ranging from 1,000 up to 30,000 μm2. We investigated the genetic basis of intraspecific differences in the naked valley in D. melanogaster and found that neither Ubx nor shavenbaby (svb) [7, 8] contributes to this morphological difference. Instead, we show that changes in mir-92a expression underlie the evolution of naked valley size in D. melanogaster through repression of shavenoid (sha) [9]. Therefore, our results reveal a novel mechanism for morphological evolution and suggest that modulation of the expression of miRNAs potentially plays a prominent role in generating organismal diversity.
► mir-92a represses shavenoid in the posterior femur to modulate naked valley size ► cis-regulatory changes in mir-92a cause the evolution of morphology in Drosophila ► Trichome pattern changes are caused by different regulatory factors ► Changes in miRNA expression might play a prominent role in phenotypic change
PMCID: PMC3605577  PMID: 23453955
8.  Mirtrons: microRNA biogenesis via splicing 
Biochimie  2011;93(11):1897-1904.
A well-defined mechanism governs the maturation of most microRNAs (miRNAs) in animals, via stepwise cleavage of precursor hairpin transcripts by the Drosha and Dicer RNase III enzymes. Recently, several alternative miRNA biogenesis pathways were elucidated, the most prominent of which substitutes Drosha cleavage with splicing. Such short hairpin introns are known as mirtrons, and their study has uncovered related pathways that combine splicing with other ribonucleolytic machinery to yield Dicer substrates for miRNA biogenesis. In this review, we consider the mechanisms of splicing-mediated miRNA biogenesis, computational strategies for mirtron discovery, and the evolutionary implications of the existence of multiple miRNA biogenesis pathways. Altogether, the features of mirtron pathways illustrate unexpected flexibility in combining RNA processing pathways, and highlight how multiple functions can be encoded by individual transcripts.
PMCID: PMC3185189  PMID: 21712066
mirtron; microRNA; small RNA biogenesis; splicing
9.  Synchronous right hepatectomy and cesarean section in a pregnant lady with hepatocellular carcinoma 
Cancer in pregnancy is rare and hepatocellular carcinoma (HCC) during pregnancy is even rarer. Due to limited experience, management of these patients remains challenging.
A 33-year old pregnant lady presented with HCC at 28 weeks of gestation. She underwent synchronous cesarean section and right hepatectomy at 32 weeks of gestation. The post-operative course was uneventful. She was discharged home on day 10 after surgery. Histolopathology confirmed HCC. The surgical resection margins were clear. At a follow-up of 3 months after surgery, the mother was disease free and the infant was well.
HCC during pregnancy is extremely rare. The experience in its management and outcomes are lacking. In managing any patient diagnosed with a malignant neoplasm in pregnancy, both the mother and the fetus have to be considered.
With adequate preoperative assessment and a good management strategy, good results can be obtained for both the mother and the baby for a pregnant patient with HCC.
PMCID: PMC3537950  PMID: 23159911
Hepatocellular carcinoma; Hepatectomy; Pregnancy; Cesarean section
10.  The long and short of inverted repeat genes in animals 
Cell cycle (Georgetown, Tex.)  2008;7(18):2840-2845.
MicroRNAs (miRNAs) are endogenous transcripts that contain intramolecular double stranded RNA (dsRNA) and are processed by Dicer. Their mature products are ~21-24 nucleotides in length, and they collectively regulate a broad network of endogenous transcripts. A subset of animal miRNAs are produced from mirtrons, short hairpin introns whose splicing bypasses the normal nuclear processing of canonical miRNAs. Recent studies revealed novel, extended intramolecular dsRNA produced by defined transcription units in flies and mammals, termed hairpin RNAs (hpRNAs). Detailed biogenesis studies in Drosophila showed that hpRNAs are not merely “long” miRNAs, but are actually processed by a distinct biogenesis pathway that is related to the canonical RNA interference pathway. We compare and contrast the miRNA and hpRNA pathways in this review, and describe some of the key questions that the recognition of this novel pathway raises.
PMCID: PMC2697033  PMID: 18769156
microRNA; mirtron; hairpin RNA; siRNA; Drosophila
11.  microRNA control of cell-cell signaling during development and disease 
Cell cycle (Georgetown, Tex.)  2008;7(15):2327-2332.
MicroRNAs (miRNAs) are critical post-transcriptional regulators that may collectively control a majority of animal genes. With thousands of miRNAs identified, a pressing challenge is now to understand their specific biological activities. Many predicted miRNA:target interactions only subtly alter gene activity. It has consequently not been trivial to deduce how miRNAs are relevant to phenotype, and by extension, relevant to disease. We note that the major signal transduction cascades that control animal development are highly dose-sensitive and frequently altered in human disorders. On this basis, we hypothesize that developmental cell signaling pathways represent prime candidates for mediating some of the major phenotypic consequences of miRNA deregulation, especially under gain-of-function conditions. This perspective reviews the evidence for miRNA targeting of the major signaling pathways, and discusses its implications for how aberrant miRNA activity might underlie human disease and cancer.
PMCID: PMC2697031  PMID: 18677099
microRNA; cell-cell signaling; disease; cancer; apoptosis
12.  Transgenic Inhibitors of RNA Interference in Drosophila 
Fly  2007;1(6):311-316.
RNA silencing functions as an adaptive antiviral defense in both plants and animals. In turn, viruses commonly encode suppressors of RNA silencing, which enable them to mount productive infection. These inhibitor proteins may be exploited as reagents with which to probe mechanisms and functions of RNA silencing pathways. In this report, we describe transgenic Drosophila strains that allow inducible expression of the viral RNA silencing inhibitors Flock House virus-B2, Nodamura virus-B2, vaccinia virus-E3L, influenza A virus-NS1 and tombusvirus P19. Some of these, especially the B2 proteins, are effective transgenic inhibitors of double strand RNA-induced gene silencing in flies. On the other hand, none of them is effective against the Drosophila microRNA pathway. Their functional selectivity makes these viral silencing proteins useful reagents with which to study biological functions of the Drosophila RNA interference pathway.
PMCID: PMC2714256  PMID: 18820441
RNAi; virus; microRNA; post-transcriptional; regulation
13.  Alternative miRNA biogenesis pathways and the interpretation of core miRNA pathway mutants 
Molecular cell  2011;43(6):892-903.
Since the establishment of a canonical animal microRNA biogenesis pathway driven by the RNase III enzymes Drosha and Dicer, an unexpected variety of alternative mechanisms that generate functional microRNAs have emerged. We review here the many Drosha-independent and Dicer-independent microRNA biogenesis strategies characterized over the past few years. Beyond reflecting the flexibility of small RNA machineries, the existence of non-canonical pathways has consequences for interpreting mutants in the core microRNA machinery. Such mutants are commonly used to assess the consequences of “total” microRNA loss, and indeed, they exhibit many overall phenotypic similarities. Nevertheless, ongoing studies reveal a growing number of settings in which alternative microRNA pathways contribute to distinct phenotypes amongst core microRNA biogenesis mutants.
PMCID: PMC3176435  PMID: 21925378
14.  Discovery of functional elements in 12 Drosophila genomes using evolutionary signatures 
Nature  2007;450(7167):219-232.
Sequencing of multiple related species followed by comparative genomics analysis constitutes a powerful approach for the systematic understanding of any genome. Here, we use the genomes of 12 Drosophila species for the de novo discovery of functional elements in the fly. Each type of functional element shows characteristic patterns of change, or ‘evolutionary signatures’, dictated by its precise selective constraints. Such signatures enable recognition of new protein-coding genes and exons, spurious and incorrect gene annotations, and numerous unusual gene structures, including abundant stop-codon readthrough. Similarly, we predict non-protein-coding RNA genes and structures, and new microRNA (miRNA) genes. We provide evidence of miRNA processing and functionality from both hairpin arms and both DNA strands. We identify several classes of pre- and post-transcriptional regulatory motifs, and predict individual motif instances with high confidence. We also study how discovery power scales with the divergence and number of species compared, and we provide general guidelines for comparative studies.
PMCID: PMC2474711  PMID: 17994088
15.  Control of microRNA biogenesis and transcription by cell signaling pathways 
A limited set of cell-cell signaling pathways presides over the vast majority of animal developmental events. The typical raison d'etre for signal transduction is to control the transcription of protein-coding genes. However, with the recent appreciation of microRNAs, growing attention has been paid towards understanding how signaling pathways intertwine with microRNA-mediated regulation. This review highlights recent studies that uncover unexpected modes of microRNA regulation by cell signaling pathways. Not only can miRNA transcription be positively or negatively regulated by cell signaling, the TGF-β/BMP pathways and Ras/MAPK pathways have now been shown to directly influence microRNA biogenesis to mediate substantial cellular phenotypes.
PMCID: PMC3149747  PMID: 21592778
16.  Ars2 maintains neural stem cell identity via direct transcriptional activation of Sox2 
Nature  2011;481(7380):195-198.
Fundamental questions concern the transcriptional networks that control the identity and self-renewal of neural stem cells (NSCs), a specialized subset of astroglial cells endowed with stem properties and neurogenic capacity. We observed that the zinc finger protein Ars2 is expressed by adult NSCs from the subventricular zone (SVZ). Selective knockdown of Ars2 in GFAP+ cells within the adult SVZ depleted NSC number and their neurogenic capacity. These phenotypes were recapitulated in the postnatal SVZ of hGFAP-Cre::Ars2fl/fl conditional knockouts, but were more severe. Ex vivo assays showed that Ars2 was necessary and sufficient to promote NSC self-renewal, by positively regulating the expression of Sox2. Although plant1–3 and animal4,5 orthologs of Ars2 are known for their conserved roles in microRNA biogenesis, we unexpectedly observed that Ars2 retained capacity to promote self-renewal in Drosha and Dicer knockout NSCs. Instead, chromatin immunoprecipitation revealed that Ars2 bound a specific region within the 6kb NSC enhancer of Sox2. This association was RNA-independent, and the bound region was required for Ars2-mediated activation of Sox2. We used gel-shift analysis to confirm direct interaction, and to refine the region bound by Ars2 to a specific conserved DNA sequence. The importance of Sox2 as a critical downstream effector was shown by its ability to restore the self-renewal and multipotency defects of Ars2 knockout NSCs. Altogether, we reveal Ars2 as a novel transcription factor that controls the multipotent progenitor state of NSCs via direct activation of the pluripotency factor Sox2.
PMCID: PMC3261657  PMID: 22198669
17.  Global Patterns of Tissue-Specific Alternative Polyadenylation in Drosophila 
Cell reports  2012;1(3):277-289.
We analyzed the usage and consequences of alternative cleavage and polyadenylation (APA) in Drosophila melanogaster by using >1 billion reads of stranded mRNA-seq across a variety of dissected tissues. Beyond demonstrating that a majority of fly transcripts are subject to APA, we observed broad trends for 3′ untranslated region (UTR) shortening in the testis and lengthening in the central nervous system (CNS); the latter included hundreds of unannotated extensions ranging up to 18 kb. Extensive northern analyses validated the accumulation of full-length neural extended transcripts, and in situ hybridization indicated their spatial restriction to the CNS. Genes encoding RNA binding proteins (RBPs) and transcription factors were preferentially subject to 3′ UTR extensions. Motif analysis indicated enrichment of miRNA and RBP sites in the neural extensions, and their termini were enriched in canonical cis elements that promote cleavage and polyadenylation. Altogether, we reveal broad tissue-specific patterns of APA in Drosophila and transcripts with unprecedented 3′ UTR length in the nervous system.
PMCID: PMC3368434  PMID: 22685694
18.  Neurophysiological Defects and Neuronal Gene Deregulation in Drosophila mir-124 Mutants 
PLoS Genetics  2012;8(2):e1002515.
miR-124 is conserved in sequence and neuronal expression across the animal kingdom and is predicted to have hundreds of mRNA targets. Diverse defects in neural development and function were reported from miR-124 antisense studies in vertebrates, but a nematode knockout of mir-124 surprisingly lacked detectable phenotypes. To provide genetic insight from Drosophila, we deleted its single mir-124 locus and found that it is dispensable for gross aspects of neural specification and differentiation. On the other hand, we detected a variety of mutant phenotypes that were rescuable by a mir-124 genomic transgene, including short lifespan, increased dendrite variation, impaired larval locomotion, and aberrant synaptic release at the NMJ. These phenotypes reflect extensive requirements of miR-124 even under optimal culture conditions. Comparison of the transcriptomes of cells from wild-type and mir-124 mutant animals, purified on the basis of mir-124 promoter activity, revealed broad upregulation of direct miR-124 targets. However, in contrast to the proposed mutual exclusion model for miR-124 function, its functional targets were relatively highly expressed in miR-124–expressing cells and were not enriched in genes annotated with epidermal expression. A notable aspect of the direct miR-124 network was coordinate targeting of five positive components in the retrograde BMP signaling pathway, whose activation in neurons increases synaptic release at the NMJ, similar to mir-124 mutants. Derepression of the direct miR-124 target network also had many secondary effects, including over-activity of other post-transcriptional repressors and a net incomplete transition from a neuroblast to a neuronal gene expression signature. Altogether, these studies demonstrate complex consequences of miR-124 loss on neural gene expression and neurophysiology.
Author Summary
microRNAs are abundant ∼22 nucleotide RNAs inferred to mediate pervasive post-transcriptional control of most genes. Still, relatively little is understood about their endogenous requirements and impact, especially in animal systems. We analyzed a knockout of Drosophila mir-124, which is conserved in sequence and neuronal expression across the animal kingdom, and predicted to have hundreds of mRNA targets. While dispensable for gross neural specification and differentiation, deletion of mir-124 caused short lifespan, increased variation in dendrite numbers, impaired larval locomotion, and aberrant synaptic release at the NMJ. These phenotypes reflect extensive requirements of miR-124 even under optimal culture conditions. Loss of miR-124 broadly upregulated its direct targets but did not support the proposed mutual exclusion model, as its functional target genes were relatively highly expressed in neurons. One notable aspect of the direct miR-124 network was coordinate targeting of five positive components in the retrograde BMP signaling pathway, whose activation in neurons phenocopies loss of miR-124. Derepression of the direct miR-124 target network had many secondary effects, including over-activity of other post-transcriptional repressors and impaired transition from neuroblast to neuronal transcriptome signatures. Altogether, we demonstrate complex requirements for this conserved miRNA on gene expression and neurophysiology.
PMCID: PMC3276548  PMID: 22347817
19.  Inhibition of Hyaluronan Synthase-3 Decreases Subcutaneous Colon Cancer Growth by Increasing Apoptosis 
Hyaluronan (HA) and hyaluronan synthases (HAS) have been implicated in cancer growth and progression. We previously have shown that HAS3 and HA mediate tumor growth in SW620 colon cancer cells, but the mechanism remains poorly understood. In addition, the effect of HAS3 inhibition on tumor growth with other cells lines has not been explored. We therefore hypothesized that inhibition of HAS3 in highly tumorigenic HCT116 colon cancer cells would decrease tumor growth and that the underlying mechanism would involve altering proliferation and/or apoptosis. HAS3 expression was inhibited by transfection with siRNA; a scrambled sequence served as a control. Stable transfectants were injected into the flanks of nude mice and tumor growth followed for 30 days. Proliferation and apoptosis were then assessed in the harvested tumors. Results were compared using the Students’ t-test and ANOVA where appropriate. siRNA transfection decreased HAS3 expression, protein production, and pericellular HA retention, and decreased in vivo tumor growth. Proliferation was unaffected in the HCT116 tumors, but increased slightly in the SW620 tumors. In contrast, HAS3 inhibition significantly increased apoptosis in all tumors. HAS3 inhibition decreases subcutaneous tumor growth by colon cancer cells and significantly increases apoptosis with less effect on proliferation. These data show that HAS3 and HA mediate colon cancer growth by inhibiting apoptosis.
PMCID: PMC3265824  PMID: 21453239
Colon carcinoma; hyaluronan; hyaluronan synthase; apoptosis proliferation; HCT116; SW620
20.  The Drosophila miR-310 cluster negatively regulates synaptic strength at the neuromuscular junction 
Neuron  2010;68(5):879-893.
Emerging data implicate microRNAs (miRNAs) in the regulation of synaptic structure and function, but we know little about their role in the regulation of neurotransmission in presynaptic neurons. Here we demonstrate that the miR-310-313 cluster is required for normal synaptic transmission at the Drosophila larval neuromuscular junction. Loss of miR-310-313 cluster leads to a significant enhancement of neurotransmitter release, which can be rescued with temporally restricted expression of mir-310-313 in larval presynaptic neurons. Kinesin family member, Khc-73 is a functional target for miR-310-313 as its expression is increased in mir-310-313 mutants and reducing it restores normal synaptic function. Cluster mutants show an increase in the active zone protein Bruchpilot accompanied by an increase in electron dense T-bars. Finally, we show that repression of Khc-73 by miR-310-313 cluster influences the establishment of normal synaptic homeostasis. Our findings establish a role for miRNAs in the regulation of neurotransmitter release.
PMCID: PMC3034365  PMID: 21145002
21.  Identification of Functional Elements and Regulatory Circuits by Drosophila modENCODE 
Roy, Sushmita | Ernst, Jason | Kharchenko, Peter V. | Kheradpour, Pouya | Negre, Nicolas | Eaton, Matthew L. | Landolin, Jane M. | Bristow, Christopher A. | Ma, Lijia | Lin, Michael F. | Washietl, Stefan | Arshinoff, Bradley I. | Ay, Ferhat | Meyer, Patrick E. | Robine, Nicolas | Washington, Nicole L. | Di Stefano, Luisa | Berezikov, Eugene | Brown, Christopher D. | Candeias, Rogerio | Carlson, Joseph W. | Carr, Adrian | Jungreis, Irwin | Marbach, Daniel | Sealfon, Rachel | Tolstorukov, Michael Y. | Will, Sebastian | Alekseyenko, Artyom A. | Artieri, Carlo | Booth, Benjamin W. | Brooks, Angela N. | Dai, Qi | Davis, Carrie A. | Duff, Michael O. | Feng, Xin | Gorchakov, Andrey A. | Gu, Tingting | Henikoff, Jorja G. | Kapranov, Philipp | Li, Renhua | MacAlpine, Heather K. | Malone, John | Minoda, Aki | Nordman, Jared | Okamura, Katsutomo | Perry, Marc | Powell, Sara K. | Riddle, Nicole C. | Sakai, Akiko | Samsonova, Anastasia | Sandler, Jeremy E. | Schwartz, Yuri B. | Sher, Noa | Spokony, Rebecca | Sturgill, David | van Baren, Marijke | Wan, Kenneth H. | Yang, Li | Yu, Charles | Feingold, Elise | Good, Peter | Guyer, Mark | Lowdon, Rebecca | Ahmad, Kami | Andrews, Justen | Berger, Bonnie | Brenner, Steven E. | Brent, Michael R. | Cherbas, Lucy | Elgin, Sarah C. R. | Gingeras, Thomas R. | Grossman, Robert | Hoskins, Roger A. | Kaufman, Thomas C. | Kent, William | Kuroda, Mitzi I. | Orr-Weaver, Terry | Perrimon, Norbert | Pirrotta, Vincenzo | Posakony, James W. | Ren, Bing | Russell, Steven | Cherbas, Peter | Graveley, Brenton R. | Lewis, Suzanna | Micklem, Gos | Oliver, Brian | Park, Peter J. | Celniker, Susan E. | Henikoff, Steven | Karpen, Gary H. | Lai, Eric C. | MacAlpine, David M. | Stein, Lincoln D. | White, Kevin P. | Kellis, Manolis
Science (New York, N.Y.)  2010;330(6012):1787-1797.
To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- and tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation.
PMCID: PMC3192495  PMID: 21177974
22.  Middle-preserving pancreatectomy for synchronous ampullary carcinoma and solid-pseudopapillary tumor of distal pancreas 
Total pancreatectomy is the treatment of choice for multicentric diseases involving the pancreas. Middle-preserving pancreatectomy is a recently reported alternative procedure when the pancreatic body is spared from disease.
Presentation of case
We report a 63-year old lady who underwent a combined Whipple's operation and distal splenopancreatectomy for her synchronous ampullary carcinoma and solid-pseudopapillary tumor of the distal pancreas.
For multiple tumors of the pancreas, the choice of surgery should be based on the nature of pathology and follow the principle of oncological resection.
Middle-preserving pancreatectomy is a safe and feasible option for patient with multicentric or synchronous pancreatic pathologies.
PMCID: PMC3215245  PMID: 22096749
Pancreaticoduodenectomy; Distal pancreatectomy; Middle-preserving pancreatectomy; Carcinoma of ampulla; Solid-pseudopapillary tumor of the pancreas
23.  A view from Drosophila: multiple biological functions for individual microRNAs 
microRNAs (miRNAs) comprise an extensive class of post-transcriptional regulatory molecules in higher eukaryotes. Intensive research in Drosophila has revealed that miRNAs control myriad developmental and physiological processes. Interestingly, several of the best-studied miRNAs impact multiple biological processes, often by regulating distinct key target genes in each setting. Here we discuss the roles of some of these pleiotropic miRNAs, and their implications for studying and interpreting the roles of miRNAs in gene regulatory networks.
PMCID: PMC2919623  PMID: 20211749
24.  R2D2 Organizes Small Regulatory RNA Pathways in Drosophila▿ †  
Molecular and Cellular Biology  2010;31(4):884-896.
Drosophila microRNAs (miRNAs) and small interfering RNAs (siRNAs) are generally produced by different Dicer enzymes (Dcr-1 and Dcr-2) and sorted to functionally distinct Argonaute effectors (AGO1 and AGO2). However, there is cross talk between these pathways, as highlighted by the recognition that Drosophila miRNA* strands (the partner strands of mature miRNAs) are generated by Dcr-1 but are preferentially sorted to AGO2. Here, we show that a component of the siRNA loading complex, R2D2, is essential both to load endogenously encoded siRNAs (endo-siRNAs) into AGO2 and to prevent endo-siRNAs from binding to AGO1. Northern blot analysis and deep sequencing showed that in the r2d2 mutant, all classes of endo-siRNAs were unable to load AGO2 and instead accumulated in the AGO1 complex. Such redirection was specific to endo-siRNAs and was not observed with miRNA* strands. We observed functional consequences of altered sorting in RNA interference (RNAi) mutants, since endo-siRNAs generated from cis-natural antisense transcripts (cis-NAT-siRNA) exhibited evidence for biased maturation as single strands in AGO1 according to thermodynamic asymmetry and a hairpin-derived endo-siRNA formed cleavage-competent complexes with AGO1 upon mutation of r2d2. Finally, we demonstrated a direct role for the R2D2/Dcr-2 heterodimer in sensing central mismatch positions that direct miRNA* strands to AGO2. Together, these data reveal new roles of R2D2 in organizing small RNA networks in Drosophila.
PMCID: PMC3028645  PMID: 21135122
25.  Integrative Analysis of the Caenorhabditis elegans Genome by the modENCODE Project 
Gerstein, Mark B. | Lu, Zhi John | Van Nostrand, Eric L. | Cheng, Chao | Arshinoff, Bradley I. | Liu, Tao | Yip, Kevin Y. | Robilotto, Rebecca | Rechtsteiner, Andreas | Ikegami, Kohta | Alves, Pedro | Chateigner, Aurelien | Perry, Marc | Morris, Mitzi | Auerbach, Raymond K. | Feng, Xin | Leng, Jing | Vielle, Anne | Niu, Wei | Rhrissorrakrai, Kahn | Agarwal, Ashish | Alexander, Roger P. | Barber, Galt | Brdlik, Cathleen M. | Brennan, Jennifer | Brouillet, Jeremy Jean | Carr, Adrian | Cheung, Ming-Sin | Clawson, Hiram | Contrino, Sergio | Dannenberg, Luke O. | Dernburg, Abby F. | Desai, Arshad | Dick, Lindsay | Dosé, Andréa C. | Du, Jiang | Egelhofer, Thea | Ercan, Sevinc | Euskirchen, Ghia | Ewing, Brent | Feingold, Elise A. | Gassmann, Reto | Good, Peter J. | Green, Phil | Gullier, Francois | Gutwein, Michelle | Guyer, Mark S. | Habegger, Lukas | Han, Ting | Henikoff, Jorja G. | Henz, Stefan R. | Hinrichs, Angie | Holster, Heather | Hyman, Tony | Iniguez, A. Leo | Janette, Judith | Jensen, Morten | Kato, Masaomi | Kent, W. James | Kephart, Ellen | Khivansara, Vishal | Khurana, Ekta | Kim, John K. | Kolasinska-Zwierz, Paulina | Lai, Eric C. | Latorre, Isabel | Leahey, Amber | Lewis, Suzanna | Lloyd, Paul | Lochovsky, Lucas | Lowdon, Rebecca F. | Lubling, Yaniv | Lyne, Rachel | MacCoss, Michael | Mackowiak, Sebastian D. | Mangone, Marco | McKay, Sheldon | Mecenas, Desirea | Merrihew, Gennifer | Miller, David M. | Muroyama, Andrew | Murray, John I. | Ooi, Siew-Loon | Pham, Hoang | Phippen, Taryn | Preston, Elicia A. | Rajewsky, Nikolaus | Rätsch, Gunnar | Rosenbaum, Heidi | Rozowsky, Joel | Rutherford, Kim | Ruzanov, Peter | Sarov, Mihail | Sasidharan, Rajkumar | Sboner, Andrea | Scheid, Paul | Segal, Eran | Shin, Hyunjin | Shou, Chong | Slack, Frank J. | Slightam, Cindie | Smith, Richard | Spencer, William C. | Stinson, E. O. | Taing, Scott | Takasaki, Teruaki | Vafeados, Dionne | Voronina, Ksenia | Wang, Guilin | Washington, Nicole L. | Whittle, Christina M. | Wu, Beijing | Yan, Koon-Kiu | Zeller, Georg | Zha, Zheng | Zhong, Mei | Zhou, Xingliang | Ahringer, Julie | Strome, Susan | Gunsalus, Kristin C. | Micklem, Gos | Liu, X. Shirley | Reinke, Valerie | Kim, Stuart K. | Hillier, LaDeana W. | Henikoff, Steven | Piano, Fabio | Snyder, Michael | Stein, Lincoln | Lieb, Jason D. | Waterston, Robert H.
Science (New York, N.Y.)  2010;330(6012):1775-1787.
We systematically generated large-scale data sets to improve genome annotation for the nematode Caenorhabditis elegans, a key model organism. These data sets include transcriptome profiling across a developmental time course, genome-wide identification of transcription factor–binding sites, and maps of chromatin organization. From this, we created more complete and accurate gene models, including alternative splice forms and candidate noncoding RNAs. We constructed hierarchical networks of transcription factor–binding and microRNA interactions and discovered chromosomal locations bound by an unusually large number of transcription factors. Different patterns of chromatin composition and histone modification were revealed between chromosome arms and centers, with similarly prominent differences between autosomes and the X chromosome. Integrating data types, we built statistical models relating chromatin, transcription factor binding, and gene expression. Overall, our analyses ascribed putative functions to most of the conserved genome.
PMCID: PMC3142569  PMID: 21177976

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