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https://www.readbyqxmd.com/read/28718764/crispr-mediated-genetic-interaction-profiling-identifies-rna-binding-proteins-controlling-metazoan-fitness
#1
Adam D Norris, Xicotencatl Gracida, John A Calarco
Genetic interaction screens have aided our understanding of complex genetic traits, diseases, and biological pathways. However, approaches for synthetic genetic analysis with null-alleles in metazoans have not been feasible. Here, we present a CRISPR/Cas9-based Synthetic Genetic Interaction (CRISPR-SGI) approach enabling systematic double-mutant generation. Applying this technique in Caenorhabditis elegans, we comprehensively screened interactions within a set of 14 conserved RNA binding protein genes, generating all possible single and double mutants...
July 18, 2017: ELife
https://www.readbyqxmd.com/read/28648677/cell-type-specific-transcriptome-profiling-in-c-elegans-using-the-translating-ribosome-affinity-purification-technique
#2
Xicotencatl Gracida, John A Calarco
Organs and specific cell types execute specialized functions in multicellular organisms, in large part through customized gene expression signatures. Thus, profiling the transcriptomes of specific cell and tissue types remains an important tool for understanding how cells become specialized. Methodological approaches to detect gene expression differences have utilized samples from whole animals, dissected tissues, and more recently single cells. Despite these advances, there is still a challenge and a need in most laboratories to implement less invasive yet powerful cell-type specific transcriptome profiling methods...
June 23, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28271995/correction-%C3%A2-neuroendocrine-modulation-sustains-the-c-elegans-forward-motor-state
#3
Maria Lim, Jyothsna Chitturi, Valeriya Laskova, Jun Meng, Daniel Findeis, Anne Wiekenberg, Ben Mulcahy, Linjiao Luo, Yan Li, Yangning Lu, Wesley Hung, Yixin Qu, Chiyip Ho, Douglas Holmyard, Ni Ji, Rebecca D McWhirter, Aravinthan Dt Samuel, David M Miller, Ralf Schnabel, John A Calarco, Mei Zhen
No abstract text is available yet for this article.
March 8, 2017: ELife
https://www.readbyqxmd.com/read/28145493/serotonin-dependent-kinetics-of-feeding-bursts-underlie-a-graded-response-to-food-availability-in-c-elegans
#4
Kyung Suk Lee, Shachar Iwanir, Ronen B Kopito, Monika Scholz, John A Calarco, David Biron, Erel Levine
Animals integrate physiological and environmental signals to modulate their food uptake. The nematode C. elegans, whose food uptake consists of pumping bacteria from the environment into the gut, provides excellent opportunities for discovering principles of conserved regulatory mechanisms. Here we show that worms implement a graded feeding response to the concentration of environmental bacteria by modulating a commitment to bursts of fast pumping. Using long-term, high-resolution, longitudinal recordings of feeding dynamics under defined conditions, we find that the frequency and duration of pumping bursts increase and the duration of long pauses diminishes in environments richer in bacteria...
February 1, 2017: Nature Communications
https://www.readbyqxmd.com/read/27941796/dynamics-of-baf-polycomb-complex-opposition-on-heterochromatin-in-normal-and-oncogenic-states
#5
Cigall Kadoch, Robert T Williams, Joseph P Calarco, Erik L Miller, Christopher M Weber, Simon M G Braun, John L Pulice, Emma J Chory, Gerald R Crabtree
The opposition between Polycomb repressive complexes (PRCs) and BAF (mSWI/SNF) complexes has a critical role in both development and disease. Mutations in the genes encoding BAF subunits contribute to more than 20% of human malignancies, yet the underlying mechanisms remain unclear, owing largely to a lack of assays to assess BAF function in living cells. To address this, we have developed a widely applicable recruitment assay system through which we find that BAF opposes PRC by rapid, ATP-dependent eviction, leading to the formation of accessible chromatin...
February 2017: Nature Genetics
https://www.readbyqxmd.com/read/27855782/neuroendocrine-modulation-sustains-the-c-elegans-forward-motor-state
#6
Maria A Lim, Jyothsna Chitturi, Valeriya Laskova, Jun Meng, Daniel Findeis, Anne Wiekenberg, Ben Mulcahy, Linjiao Luo, Yan Li, Yangning Lu, Wesley Hung, Yixin Qu, Chi-Yip Ho, Douglas Holmyard, Ni Ji, Rebecca McWhirter, Aravinthan Dt Samuel, David M Miller, Ralf Schnabel, John A Calarco, Mei Zhen
Neuromodulators shape neural circuit dynamics. Combining electron microscopy, genetics, transcriptome profiling, calcium imaging, and optogenetics, we discovered a peptidergic neuron that modulates C. elegans motor circuit dynamics. The Six/SO-family homeobox transcription factor UNC-39 governs lineage-specific neurogenesis to give rise to a neuron RID. RID bears the anatomic hallmarks of a specialized endocrine neuron: it harbors near-exclusive dense core vesicles that cluster periodically along the axon, and expresses multiple neuropeptides, including the FMRF-amide-related FLP-14...
November 18, 2016: ELife
https://www.readbyqxmd.com/read/27256389/regulation-of-tissue-specific-alternative-splicing-c-elegans-as-a-model-system
#7
Xicotencatl Gracida, Adam D Norris, John A Calarco
Alternative pre-mRNA splicing serves as an elegant mechanism for generating transcriptomic and proteomic diversity between cell and tissue types. In this chapter, we highlight key concepts and outstanding goals in studies of tissue and cell-specific alternative splicing. We place particular emphasis on the use of C. elegans as a tractable model organism for in vivo studies of alternative splicing between tissues and also at single cell resolution. We describe our current understanding of tissue and cell-specific regulation in the animal, and emerging techniques that will allow for future mechanistic studies as well as systems level investigations of spatio-temporal splicing under laboratory conditions and in response to environmental stimuli...
2016: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/26926116/revisiting-the-local-structure-in-ge-sb-te-based-chalcogenide-superlattices
#8
Barbara Casarin, Antonio Caretta, Jamo Momand, Bart J Kooi, Marcel A Verheijen, Valeria Bragaglia, Raffaella Calarco, Marina Chukalina, Xiaoming Yu, John Robertson, Felix R L Lange, Matthias Wuttig, Andrea Redaelli, Enrico Varesi, Fulvio Parmigiani, Marco Malvestuto
The technological success of phase-change materials in the field of data storage and functional systems stems from their distinctive electronic and structural peculiarities on the nanoscale. Recently, superlattice structures have been demonstrated to dramatically improve the optical and electrical performances of these chalcogenide based phase-change materials. In this perspective, unravelling the atomistic structure that originates the improvements in switching time and switching energy is paramount in order to design nanoscale structures with even enhanced functional properties...
2016: Scientific Reports
https://www.readbyqxmd.com/read/26232410/efficient-genome-editing-in-caenorhabditis-elegans-with-a-toolkit-of-dual-marker-selection-cassettes
#9
Adam D Norris, Hyun-Min Kim, Mónica P Colaiácovo, John A Calarco
Use of the CRISPR/Cas9 RNA-guided endonuclease complex has recently enabled the generation of double-strand breaks virtually anywhere in the C. elegans genome. Here, we present an improved strategy that makes all steps in the genome editing process more efficient. We have created a toolkit of template-mediated repair cassettes that contain an antibiotic resistance gene to select for worms carrying the repair template and a fluorescent visual marker that facilitates identification of bona fide recombinant animals...
October 2015: Genetics
https://www.readbyqxmd.com/read/25274815/eol-1-the-homolog-of-the-mammalian-dom3z-regulates-olfactory-learning-in-c-elegans
#10
Yu Shen, Jiangwen Zhang, John A Calarco, Yun Zhang
Learning is an essential function of the nervous system. However, our understanding of molecular underpinnings of learning remains incomplete. Here, we characterize a conserved protein EOL-1 that regulates olfactory learning in Caenorhabditis elegans. A recessive allele of eol-1 (enhanced olfactory learning) learns better to adjust its olfactory preference for bacteria foods and eol-1 acts in the URX sensory neurons to regulate learning. The mammalian homolog of EOL-1, Dom3Z, which regulates quality control of pre-mRNAs, can substitute the function of EOL-1 in learning regulation, demonstrating functional conservation between these homologs...
October 1, 2014: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/24910101/a-pair-of-rna-binding-proteins-controls-networks-of-splicing-events-contributing-to-specialization-of-neural-cell-types
#11
Adam D Norris, Shangbang Gao, Megan L Norris, Debashish Ray, Arun K Ramani, Andrew G Fraser, Quaid Morris, Timothy R Hughes, Mei Zhen, John A Calarco
Alternative splicing is important for the development and function of the nervous system, but little is known about the differences in alternative splicing between distinct types of neurons. Furthermore, the factors that control cell-type-specific splicing and the physiological roles of these alternative isoforms are unclear. By monitoring alternative splicing at single-cell resolution in Caenorhabditis elegans, we demonstrate that splicing patterns in different neurons are often distinct and highly regulated...
June 19, 2014: Molecular Cell
https://www.readbyqxmd.com/read/23979579/heritable-custom-genomic-modifications-in-caenorhabditis-elegans-via-a-crispr-cas9-system
#12
Yonatan B Tzur, Ari E Friedland, Saravanapriah Nadarajan, George M Church, John A Calarco, Monica P Colaiácovo
We adapted the CRISPR-Cas9 system for template-mediated repair of targeted double-strand breaks via homologous recombination in Caenorhabditis elegans, enabling customized and efficient genome editing. This system can be used to create specific insertions, deletions, and base pair changes in the germline of C. elegans.
November 2013: Genetics
https://www.readbyqxmd.com/read/23817069/heritable-genome-editing-in-c-elegans-via-a-crispr-cas9-system
#13
Ari E Friedland, Yonatan B Tzur, Kevin M Esvelt, Monica P Colaiácovo, George M Church, John A Calarco
We report the use of clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated endonuclease Cas9 to target genomic sequences in the Caenorhabditis elegans germ line using single-guide RNAs that are expressed from a U6 small nuclear RNA promoter. Our results demonstrate that targeted, heritable genetic alterations can be achieved in C. elegans, providing a convenient and effective approach for generating loss-of-function mutants.
August 2013: Nature Methods
https://www.readbyqxmd.com/read/23358458/-cryptic-exons-reveal-some-of-their-secrets
#14
John A Calarco
By regulating the inclusion of 'cryptic' exons in messenger RNA in nerve cells, NOVA proteins are able to influence the abundance of the corresponding proteins.
2013: ELife
https://www.readbyqxmd.com/read/22936897/emerging-roles-of-alternative-pre-mrna-splicing-regulation-in-neuronal-development-and-function
#15
Adam D Norris, John A Calarco
Alternative pre-mRNA splicing has the potential to greatly diversify the repertoire of transcripts in multicellular organisms. Increasing evidence suggests that this expansive layer of gene regulation plays a particularly important role in the development and function of the nervous system, one of the most complex organ systems found in nature. In this review, we highlight recent studies that continue to emphasize the influence and contribution of alternative splicing regulation to various aspects of neuronal development in addition to its role in the mature nervous system...
2012: Frontiers in Neuroscience
https://www.readbyqxmd.com/read/22749401/tissue-specific-alternative-splicing-remodels-protein-protein-interaction-networks
#16
Jonathan D Ellis, Miriam Barrios-Rodiles, Recep Colak, Manuel Irimia, Taehyung Kim, John A Calarco, Xinchen Wang, Qun Pan, Dave O'Hanlon, Philip M Kim, Jeffrey L Wrana, Benjamin J Blencowe
Alternative splicing plays a key role in the expansion of proteomic and regulatory complexity, yet the functions of the vast majority of differentially spliced exons are not known. In this study, we observe that brain and other tissue-regulated exons are significantly enriched in flexible regions of proteins that likely form conserved interaction surfaces. These proteins participate in significantly more interactions in protein-protein interaction (PPI) networks than other proteins. Using LUMIER, an automated PPI assay, we observe that approximately one-third of analyzed neural-regulated exons affect PPIs...
June 29, 2012: Molecular Cell
https://www.readbyqxmd.com/read/22284174/releasing-the-inner-inhibition-for-axon-regeneration
#17
COMMENT
Michelle D Po, John A Calarco, Mei Zhen
The adult mammalian central nervous system exhibits restricted regenerative potential. Chen et al. (2011) and El Bejjani and Hammarlund (2012) used Caenorhabditis elegans to uncover intrinsic factors that inhibit regeneration of axotomized mature neurons, opening avenues for potential therapeutics.
January 26, 2012: Neuron
https://www.readbyqxmd.com/read/21415141/networking-in-a-global-world-establishing-functional-connections-between-neural-splicing-regulators-and-their-target-transcripts
#18
REVIEW
John A Calarco, Mei Zhen, Benjamin J Blencowe
Recent genome-wide analyses have indicated that almost all primary transcripts from multi-exon human genes undergo alternative pre-mRNA splicing (AS). Given the prevalence of AS and its importance in expanding proteomic complexity, a major challenge that lies ahead is to determine the functional specificity of isoforms in a cellular context. A significant fraction of alternatively spliced transcripts are regulated in a tissue- or cell-type-specific manner, suggesting that these mRNA variants likely function in the generation of cellular diversity...
May 2011: RNA
https://www.readbyqxmd.com/read/21177968/genome-wide-analysis-of-alternative-splicing-in-caenorhabditis-elegans
#19
Arun K Ramani, John A Calarco, Qun Pan, Sepand Mavandadi, Ying Wang, Andrew C Nelson, Leo J Lee, Quaid Morris, Benjamin J Blencowe, Mei Zhen, Andrew G Fraser
Alternative splicing (AS) plays a crucial role in the diversification of gene function and regulation. Consequently, the systematic identification and characterization of temporally regulated splice variants is of critical importance to understanding animal development. We have used high-throughput RNA sequencing and microarray profiling to analyze AS in C. elegans across various stages of development. This analysis identified thousands of novel splicing events, including hundreds of developmentally regulated AS events...
February 2011: Genome Research
https://www.readbyqxmd.com/read/20445623/deciphering-the-splicing-code
#20
Yoseph Barash, John A Calarco, Weijun Gao, Qun Pan, Xinchen Wang, Ofer Shai, Benjamin J Blencowe, Brendan J Frey
Alternative splicing has a crucial role in the generation of biological complexity, and its misregulation is often involved in human disease. Here we describe the assembly of a 'splicing code', which uses combinations of hundreds of RNA features to predict tissue-dependent changes in alternative splicing for thousands of exons. The code determines new classes of splicing patterns, identifies distinct regulatory programs in different tissues, and identifies mutation-verified regulatory sequences. Widespread regulatory strategies are revealed, including the use of unexpectedly large combinations of features, the establishment of low exon inclusion levels that are overcome by features in specific tissues, the appearance of features deeper into introns than previously appreciated, and the modulation of splice variant levels by transcript structure characteristics...
May 6, 2010: Nature
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