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John calarco

Jyothsna Chitturi, Wesley Hung, Anas M Abdel Rahman, Min Wu, Maria A Lim, John Calarco, Renee Baran, Xun Huang, James W Dennis, Mei Zhen
UBR1 is an E3 ubiquitin ligase best known for its ability to target protein degradation by the N-end rule. The physiological functions of UBR family proteins, however, remain not fully understood. We found that the functional loss of C. elegans UBR-1 leads to a specific motor deficit: when adult animals generate reversal movements, A-class motor neurons exhibit synchronized activation, preventing body bending. This motor deficit is rescued by removing GOT-1, a transaminase that converts aspartate to glutamate...
April 2018: PLoS Genetics
John A Calarco, Adam D Norris
Genetic interaction screens are a powerful methodology to establish novel roles for genes and elucidate functional connections between genes. Such studies have been performed to great effect in single-cell organisms such as yeast and E. coli (Schuldiner et al ., 2005; Butland et al ., 2008; Costanzo et al ., 2010), but similar large-scale interaction studies using targeted reverse-genetic deletions in multi-cellular organisms have not been feasible. We developed a CRISPR/Cas9-based method for deleting genes in C...
March 5, 2018: Bio-protocol
Xicotencatl Gracida, Michael F Dion, Gareth Harris, Yun Zhang, John A Calarco
Neuromodulatory cells transduce environmental information into long-lasting behavioral responses. However, the mechanisms governing how neuronal cells influence behavioral plasticity are difficult to characterize. Here, we adapted the translating ribosome affinity purification (TRAP) approach in C. elegans to profile ribosome-associated mRNAs from three major tissues and the neuromodulatory dopaminergic and serotonergic cells. We identified elc-2, an Elongin C ortholog, specifically expressed in stress-sensing amphid neuron dual ciliated sensory ending (ADF) serotonergic sensory neurons, and we found that it plays a role in mediating a long-lasting change in serotonin-dependent feeding behavior induced by heat stress...
December 12, 2017: Cell Reports
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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
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