Life Sciences Commons^™

Mcnamara 201412 Nih Scap Innocentive Challenge Solution - T-Bow Rainbow T-Cells And Tumor Cells Spatial Multiplexing Gene Expression Reporter System – Plus Supplement Plus Posters - 20151027 - Please Download "75" Instead, George Mcnamara

George McNamara

McNamara 201412 NIH SCAP InnoCentive Challenge Solution - T-Bow Rainbow T-cells and Tumor Cells Spatial Multiplexing Gene Expression Reporter System – plus supplement plus posters - 20151027.

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Please download the current 20151027 (October 27, 2015) Tattletales and T-Bow update from

http://works.bepress.com/gmcnamara/75/

The bepress web site is not letting me replace the old pdf here at "65" with the additional 10 pages update.

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The download is my/Cooper lab solution (submission) to the 2014 NIH Single Cell Analysis Program (SCAP) InnoCentive Challenge, "Follow That Cell". I submitted the Solution on 20141215Mon (with 20 minutes to spare). The Challenge web page …

Tattletales And T-Bow Update 20151027tue, George Mcnamara

George McNamara

20151027Tue this "75"

http://works.bepress.com/gmcnamara/75

is my update of "65" posting

See text at

http://works.bepress.com/gmcnamara/65/

for text summary. The PDf here in "75" supersedes "65".

The PDF here has 10 pages added to the end from the "65" version (pages 40-49 of PDF when including the bepress cover page)..

here is the text in my cover page (bepress may add its own cover):

20151027Tue: added 10 page e-poster at bottom explaining Binary Tattletales and T-Bow. That is, binary with respect to protein components. For one color (number of repeats, epitope tags, FPs are examples, here rounded to convenient numbers):

1. 100 …

Mechanisms Of Neuroprotection Against Ischemic Insult By Stress-Inducible Phosphoprotein-1, Jason Xu

Electronic Thesis and Dissertation Repository

Stress-inducible phosphoprotein-1 (STI1) levels are increased in the brain following ischemia. STI1 is a co-chaperone for Hsp70/Hsp90 modulating protein folding. STI1 can also be secreted by a number of cells and function to activate extracellular signalling by the prion protein (PrP^C) and type-I bone morphogenetic protein receptor ALK2. However, the mechanisms by which STI1 can protect neurons against ischemia are currently unknown. A caspase-3 reporter mouse line was used to evaluate the consequences of increased extracellular STI1 levels. Neurons were treated with recombinant STI1 and specific agonists/antagonists for PrP^C, α7nAChR, and ALK2 prior to oxygen-glucose deprivation …

Rna Recognition By The Caenorhabditis Elegans Oocyte Maturation Determinant Oma-1, Ebru Kaymak, Sean Ryder

Sean P. Ryder

Maternally supplied mRNAs encode proteins that pattern early embryos in many species. In the nematode Caenorhabditis elegans, a suite of RNA-binding proteins regulates expression of maternal mRNAs during oogenesis, the oocyte to embryo transition, and early embryogenesis. To understand how these RNA-binding proteins contribute to development, it is necessary to determine how they select specific mRNA targets for regulation. OMA-1 and OMA-2 are redundant proteins required for oocyte maturation--an essential part of meiosis that prepares oocytes for fertilization. Both proteins have CCCH type tandem zinc finger RNA-binding domains. Here, we define the RNA binding specificity of OMA-1 and demonstrate that …

Migration In The Social Stage Of Dictyostelium Discoideum Amoebae Impacts Competition, Chandra N. Jack, Neil J. Buttery, Boahemaa Adu-Oppong, Michael Powers, Christopher R.L. Thompson, David Queller, Joan E. Strassmann

Biology Faculty Publications & Presentations

Interaction conditions can change the balance of cooperation and conflict in multicellular groups. After aggregating together, cells of the social amoeba Dictyostelium discoideum may migrate as a group (known as a slug) to a new location. We consider this migration stage as an arena for social competition and conflict because the cells in the slug may not be from a genetically homogeneous population. In this study, we examined the interplay of two seemingly diametric actions, the solitary action of kin recognition and the collective action of slug migration in D. discoideum, to more fully understand the effects of social competition …

The Generation, Exploitation And Future Of Induced Pluripotent Stem Cells, Jacob Steenwyk

Scholarly Undergraduate Research Journal at Clark (SURJ)

The foundational advancements of John Gurdon and Shinya Yamanaka have improved understanding of dedifferen- tiation of cells to a pluripotent state. The seminal discovery established a novel system to study disease pathogenesis, drug screening, and toxicity, as well as sprouted the new field of regenerative medicine. In this article, the method- ology to obtain dedifferentiated cells, known as induced pluripotent stem (iPS) cells, subsequent validation, and application of which are reviewed. The experiments investigated here aim to demonstrate the capacity of iPS cells to replace the ethically-gray human embryonic cells by developing human livers and viable, healthy animals. It is …

Cardiomyocyte Polarity And Embryonic Heart Development: Role Of Rac1, Carmen Leung

Electronic Thesis and Dissertation Repository

Congenital heart defects (CHDs) are the most common human birth defect and the leading cause of death from a birth defect in the first year of life. Thus, a further understanding of the mechanisms underlying CHDs, which could lead to improved diagnosis and treatment, is crucial. The small GTPase, Rac1, acts as a pleiotropic effector of numerous cellular processes; however, little is known about its role in embryonic heart development. The aim of this thesis was to investigate the role of Rac1 signaling in cardiac development. Using the Cre/loxP system, mouse models with an anterior second heart field (SHF) or …

Dynamic Ubiquitination Drives Herpesvirus Neuroinvasion, Nicholas J. Huffmaster, Patricia J. Sollars, Alexsia L. Richards, Gary E. Pickard, Gregory A. Smith

School of Veterinary and Biomedical Sciences: Faculty Publications

Neuroinvasive herpesviruses display a remarkable propensity to enter the nervous system of healthy individuals in the absence of obvious trauma at the site of inoculation. We document a repurposing of cellular ubiquitin during infection to switch the virus between two invasive states. The states act sequentially to defeat consecutive host barriers of the peripheral nervous system and together promote the potent neuroinvasive phenotype. The first state directs virus access to nerve endings in peripheral tissue, whereas the second delivers virus particles within nerve fibers to the neural ganglia. Mutant viruses locked in either state remain competent to overcome the corresponding …

Characterization And Diurnal Measurement Of Oral Inflammation In Association With Glycemic Control, Periodontal Status, & Glucose Stimulation, Melanie N. Kuehl

USF Tampa Graduate Theses and Dissertations

Diabetes has afflicted 8.3%, approximately 25.8 million, of the United States population and is the seventh leading cause of death [1]. Type I diabetes (T1D) accounts for 5 to 10% of all diagnosed cases of diabetes in the United States [2]. If present trends continue, the rate of T1D incidence among children under the age of 14 will increase by 3% globally [3]. T1D is an autoimmune disorder in which the β-cells of the pancreatic islets are destroyed, leading to high blood sugar. Hyperglycemia and loss of immunological tolerance to self-antigens are common associations of T1D [4]. Periodontal disease impacts …

Cladribine Analogues Via O6-(Benzotriazolyl) Derivatives Of Guanine Nucleosides, Sakilam Satishkumar, Prasanna K. Vuram, Siva Subrahmanyam Relangi, Venkateshwarlu Gurram, Hong Zhou, Robert J. Kreitman, Michelle M. Martínez Montemayor, Lijia Yang, Muralidharan Kaliyaperumal, Somesh Sharma, Narender Pottabathini, Mahesh K. Lakshman

Publications and Research

Cladribine, 2-chloro-2′-deoxyadenosine, is a highly efficacious, clinically used nucleoside for the treatment of hairy cell leukemia. It is also being evaluated against other lymphoid malignancies and has been a molecule of interest for well over half a century. In continuation of our interest in the amide bond-activation in purine nucleosides via the use of (benzotriazol-1yl-oxy)tris(dimethylamino)phosphonium hexafluorophosphate, we have evaluated the use of O6-(benzotriazol-1-yl)-2′-deoxyguanosine as a potential precursor to cladribine and its analogues. These compounds, after appropriate deprotection, were assessed for their biological activities, and the data are presented herein. Against hairy cell leukemia (HCL), T-cell lymphoma (TCL) and chronic lymphocytic …

Real-Time Detection Of Breast Cancer Cells Using Peptidefunctionalized Microcantilever Arrays, Hashem Etayash, Keren Jiang, Sarfuddin Azmi, Thomas Thundat, Kamaljit Kaur

Pharmacy Faculty Articles and Research

Ligand-directed targeting and capturing of cancer cells is a new approach for detecting circulating tumor cells (CTCs). Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch®) is currently used clinically to enumerate CTCs. Here we report the use of a peptide moiety in conjunction with a microcantilever array system to selectively detect CTCs resulting from cancer, specifically breast cancer. A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells. …

Reactive Oxygen Species-Mediated Neurodegeneration Is Independent Of The Ryanodine Receptor In Caenorhabditis Elegans, Lyndsay E.A. Young, Daniel C. Williams

Journal of the South Carolina Academy of Science

Despite the significant impacts on human health caused by neurodegeneration, our understanding of the degeneration process is incomplete. The nematode Caenorhabditis elegans is emerging as a genetic model organism well suited for identification of conserved cellular mechanisms and molecular pathways of neurodegeneration. Studies in the worm have identified factors that contribute to neurodegeneration, including excitotoxicity and stress due to reactive oxygen species (ROS). Disruption of the gene unc-68, which encodes the ryanodine receptor, abolishes excitotoxic cell death, indicating a role for calcium (Ca²⁺) signaling in neurodegeneration. We tested the requirement for unc-68 in ROS-mediated neurodegeneration using the …

Caenorhabditis Elegans Alg-1 Antimorphic Mutations Uncover Functions For Argonaute In Microrna Guide Strand Selection And Passenger Strand Disposal, Anna Y. Zinovyeva, Isana Veksler-Lublinsky, Ajay A. Vashisht, James A. Wohlschlegel, Victor R. Ambros

Victor R. Ambros

MicroRNAs are regulators of gene expression whose functions are critical for normal development and physiology. We have previously characterized mutations in a Caenorhabditis elegans microRNA-specific Argonaute ALG-1 (Argonaute-like gene) that are antimorphic [alg-1(anti)]. alg-1(anti) mutants have dramatically stronger microRNA-related phenotypes than animals with a complete loss of ALG-1. ALG-1(anti) miRISC (microRNA induced silencing complex) fails to undergo a functional transition from microRNA processing to target repression. To better understand this transition, we characterized the small RNA and protein populations associated with ALG-1(anti) complexes in vivo. We extensively characterized proteins associated with wild-type and mutant ALG-1 and found that the mutant …

Robust Distal Tip Cell Pathfinding In The Face Of Temperature Stress Is Ensured By Two Conserved Micrornas In Caenorhabditis Elegans, Samantha L. Burke, Molly Hammell, Victor R. Ambros

Victor R. Ambros

Biological robustness, the ability of an organism to maintain a steady-state output as genetic or environmental inputs change, is critical for proper development. MicroRNAs have been implicated in biological robustness mechanisms through their post-transcriptional regulation of genes and gene networks. Previous research has illustrated examples of microRNAs promoting robustness as part of feedback loops and genetic switches and by buffering noisy gene expression resulting from environmental and/or internal changes. Here we show that the evolutionarily conserved microRNAs mir-34 and mir-83 (homolog of mammalian mir-29) contribute to the robust migration pattern of the distal tip cells in Caenorhabditis elegans by specifically …

Control Of Stem Cell Self-Renewal And Differentiation By The Heterochronic Genes And The Cellular Asymmetry Machinery In Caenorhabditis Elegans, Omid F. Harandi, Victor Ambros

Victor R. Ambros

Transitions between asymmetric (self-renewing) and symmetric (proliferative) cell divisions are robustly regulated in the context of normal development and tissue homeostasis. To genetically assess the regulation of these transitions, we used the postembryonic epithelial stem (seam) cell lineages of Caenorhabditis elegans. In these lineages, the timing of these transitions is regulated by the evolutionarily conserved heterochronic pathway, whereas cell division asymmetry is conferred by a pathway consisting of Wnt (Wingless) pathway components, including posterior pharynx defect (POP-1)/TCF, APC related/adenomatosis polyposis coli (APR-1)/APC, and LIT-1/NLK (loss of intestine/Nemo-like kinase). Here we explore the genetic regulatory mechanisms underlying stage-specific transitions between self-renewing …

Developmental Decline In Neuronal Regeneration By The Progressive Change Of Two Intrinsic Timers, Yan Zou, Hui Chiu, Anna Zinovyeva, Victor Ambros, Chiou-Fen Chuang, Chieh Chang

Victor R. Ambros

Like mammalian neurons, Caenorhabditis elegans neurons lose axon regeneration ability as they age, but it is not known why. Here, we report that let-7 contributes to a developmental decline in anterior ventral microtubule (AVM) axon regeneration. In older AVM axons, let-7 inhibits regeneration by down-regulating LIN-41, an important AVM axon regeneration-promoting factor. Whereas let-7 inhibits lin-41 expression in older neurons through the lin-41 3' untranslated region, lin-41 inhibits let-7 expression in younger neurons through Argonaute ALG-1. This reciprocal inhibition ensures that axon regeneration is inhibited only in older neurons. These findings show that a let-7-lin-41 regulatory circuit, which was previously …

The Evolution Of Our Thinking About Micrornas, Victor Ambros

Victor R. Ambros

Our appreciation of the significance of microRNAs to biology at large continues to be an evolving process.

Victor Ambros: The Broad Scope Of Micrornas. Interview By Caitlin Sedwick, Victor R. Ambros

Victor R. Ambros

Interview with Victor Ambros, who studies how microRNAs impact development.

Mutations In Conserved Residues Of The C. Elegans Microrna Argonaute Alg-1 Identify Separable Functions In Alg-1 Mirisc Loading And Target Repression, Anna Y. Zinovyeva, Samir Bouasker, Martin J. Simard, Christopher M. Hammell, Victor R. Ambros

Victor R. Ambros

microRNAs function in diverse developmental and physiological processes by regulating target gene expression at the post-transcriptional level. ALG-1 is one of two Caenorhabditis elegans Argonautes (ALG-1 and ALG-2) that together are essential for microRNA biogenesis and function. Here, we report the identification of novel antimorphic (anti) alleles of ALG-1 as suppressors of lin-28(lf) precocious developmental phenotypes. The alg-1(anti) mutations broadly impair the function of many microRNAs and cause dosage-dependent phenotypes that are more severe than the complete loss of ALG-1. ALG-1(anti) mutant proteins are competent for promoting Dicer cleavage of microRNA precursors and for associating with and stabilizing microRNAs. However, …

The Developmental Timing Regulator Hbl-1 Modulates The Dauer Formation Decision In Caenorhabditis Elegans, Xantha Karp, Victor Ambros

Victor R. Ambros

Animals developing in the wild encounter a range of environmental conditions, and so developmental mechanisms have evolved that can accommodate different environmental contingencies. Harsh environmental conditions cause Caenorhabditis elegans larvae to arrest as stress-resistant "dauer" larvae after the second larval stage (L2), thereby indefinitely postponing L3 cell fates. HBL-1 is a key transcriptional regulator of L2 vs. L3 cell fate. Through the analysis of genetic interactions between mutations of hbl-1 and of genes encoding regulators of dauer larva formation, we find that hbl-1 can also modulate the dauer formation decision in a complex manner. We propose that dynamic interactions between …

Mir-14 Regulates Autophagy During Developmental Cell Death By Targeting Ip3-Kinase 2, Charles Nelson, Victor Ambros, Eric Baehrecke

Victor R. Ambros

Macroautophagy (autophagy) is a lysosome-dependent degradation process that has been implicated in age-associated diseases. Autophagy is involved in both cell survival and cell death, but little is known about the mechanisms that distinguish its use during these distinct cell fates. Here, we identify the microRNA miR-14 as being both necessary and sufficient for autophagy during developmentally regulated cell death in Drosophila. Loss of miR-14 prevented induction of autophagy during salivary gland cell death, but had no effect on starvation-induced autophagy in the fat body. Moreover, misexpression of miR-14 was sufficient to prematurely induce autophagy in salivary glands, but not in …

Micrornas And Developmental Timing, Victor Ambros

Victor R. Ambros

MicroRNAs regulate temporal transitions in gene expression associated with cell fate progression and differentiation throughout animal development. Genetic analysis of developmental timing in the nematode Caenorhabditis elegans identified two evolutionarily conserved microRNAs, lin-4/mir-125 and let-7, that regulate cell fate progression and differentiation in C. elegans cell lineages. MicroRNAs perform analogous developmental timing functions in other animals, including mammals. By regulating cell fate choices and transitions between pluripotency and differentiation, microRNAs help to orchestrate developmental events throughout the developing animal, and to play tissue homeostasis roles important for disease, including cancer.

Dauer Larva Quiescence Alters The Circuitry Of Microrna Pathways Regulating Cell Fate Progression In C. Elegans, Xantha Karp, Victor Ambros

Victor R. Ambros

In C. elegans larvae, the execution of stage-specific developmental events is controlled by heterochronic genes, which include those encoding a set of transcription factors and the microRNAs that regulate the timing of their expression. Under adverse environmental conditions, developing larvae enter a stress-resistant, quiescent stage called 'dauer'. Dauer larvae are characterized by the arrest of all progenitor cell lineages at a stage equivalent to the end of the second larval stage (L2). If dauer larvae encounter conditions favorable for resumption of reproductive growth, they recover and complete development normally, indicating that post-dauer larvae possess mechanisms to accommodate an indefinite period …

The Embryonic Mir-35 Family Of Micrornas Promotes Multiple Aspects Of Fecundity In Caenorhabditis Elegans, Katherine Mcjunkin, Victor R. Ambros

Victor R. Ambros

MicroRNAs guide many aspects of development in all metazoan species. Frequently, microRNAs are expressed during a specific developmental stage to perform a temporally defined function. The C. elegans mir-35-42 microRNAs are expressed abundantly in oocytes and early embryos and are essential for embryonic development. Here, we show that these embryonic microRNAs surprisingly also function to control the number of progeny produced by adult hermaphrodites. Using a temperature-sensitive mir-35-42 family mutant (a deletion of the mir-35-41 cluster), we demonstrate three distinct defects in hermaphrodite fecundity. At permissive temperatures, a mild sperm defect partially reduces hermaphrodite fecundity. At restrictive temperatures, somatic gonad …

Caenorhabditis Elegans Micrornas Of The Let-7 Family Act In Innate Immune Response Circuits And Confer Robust Developmental Timing Against Pathogen Stress, Zhiji Ren, Victor R. Ambros

Victor R. Ambros

Animals maintain their developmental robustness against natural stresses through numerous regulatory mechanisms, including the posttranscriptional regulation of gene expression by microRNAs (miRNAs). Caenorhabditis elegans miRNAs of the let-7 family (let-7-Fam) function semiredundantly to confer robust stage specificity of cell fates in the hypodermal seam cell lineages. Here, we show reciprocal regulatory interactions between let-7-Fam miRNAs and the innate immune response pathway in C. elegans. Upon infection of C. elegans larvae with the opportunistic human pathogen Pseudomonas aeruginosa, the developmental timing defects of certain let-7-Fam miRNA mutants are enhanced. This enhancement is mediated by the p38 MAPK innate immune pathway acting …

Drosophila Let-7 Microrna Is Required For Remodeling Of The Neuromusculature During Metamorphosis, Nicholas S. Sokol, Peizhang Xu, Yuh-Nung Jan, Victor R. Ambros

Victor R. Ambros

The Drosophila let-7-Complex (let-7-C) is a polycistronic locus encoding three ancient microRNAs: let-7, miR-100, and fly lin-4 (miR-125). We find that the let-7-C locus is principally expressed in the pupal and adult neuromusculature. let-7-C knockout flies appear normal externally but display defects in adult behaviors (e.g., flight, motility, and fertility) as well as clear juvenile features in their neuromusculature. We find that the function of let-7-C to ensure the appropriate remodeling of the abdominal neuromusculature during the larval-to-adult transition is carried out predominantly by let-7 alone. This heterochronic role of let-7 is likely just one of the ways in which …

Effect Of Life History On Microrna Expression During C. Elegans Development, Xantha Karp, Molly Hammell, Maria C. Ow, Victor R. Ambros

Victor R. Ambros

Animals have evolved mechanisms to ensure the robustness of developmental outcomes to changing environments. MicroRNA expression may contribute to developmental robustness because microRNAs are key post-transcriptional regulators of developmental gene expression and can affect the expression of multiple target genes. Caenorhabditis elegans provides an excellent model to study developmental responses to environmental conditions. In favorable environments, C. elegans larvae develop rapidly and continuously through four larval stages. In contrast, in unfavorable conditions, larval development may be interrupted at either of two diapause stages: The L1 diapause occurs when embryos hatch in the absence of food, and the dauer diapause occurs …

Micrornas: Genetically Sensitized Worms Reveal New Secrets, Victor Ambros

Victor R. Ambros

Why do many microRNA gene mutants display no evident phenotype? Multiply mutant worms that are selectively impaired in genetic regulatory network activities have been used to uncover previously unknown functions for numerous Caenorhabditis elegans microRNAs.

Prb/Cki Pathways At The Interface Of Cell Cycle And Development, Victor Ambros

Victor R. Ambros

Comment on: The cyclin-dependent kinase inhibitors, cki-1 and cki-2, act in overlapping but distinct pathways to control cell-cycle quiescence during C. elegans development. Buck SH, et al. Cell Cycle 2009; 8:2613-20.

Yeast Integral Membrane Proteins Apq12, Brl1, And Brr6 Form A Complex Important For Regulation Of Membrane Homeostasis And Nuclear Pore Complex Biogenesis, Museer A. Lone, Aaron E. Atkinson, Christine A. Hodge, Stéphanie Cottier, Fernando Martínez-Montañés, Shelley Maithel, Laurent Mène-Saffrané, Cole Cole, Roger Schneiter

Dartmouth Scholarship

Proper functioning of intracellular membranes is critical for many cellular processes. A key feature of membranes is their ability to adapt to changes in environmental conditions by adjusting their composition so as to maintain constant biophysical proper- ties, including fluidity and flexibility. Similar changes in the biophysical properties of membranes likely occur when intracellular processes, such as vesicle formation and fusion, require dramatic changes in membrane curvature. Similar modifications must also be made when nuclear pore complexes (NPCs) are constructed within the existing nuclear membrane, as occurs during in- terphase in all eukaryotes. Here we report on the role of …