Distinct gating mechanism of SOC channel involving STIM–Orai coupling and an intramolecular interaction of Orai in Caenorhabditis elegans

Store-operated calcium entry (SOCE), an important mechanism of Ca2+ signaling in a wide range of cell types, is mediated by stromal interaction molecule (STIM), which senses the depletion of endoplasmic reticulum Ca2+ stores and binds and activates Orai channels in the plasma membrane. This inside-out mechanism of Ca2+ signaling raises an interesting question about the evolution of SOCE: How did these two proteins existing in different cellular compartments evolve to interact with each other? We investigated the gating mechanism of Caenorhabditis elegans Orai channels. Our analysis revealed a mechanism of Orai gating by STIM binding to the intracellular 2–3 loop of Orai in C. elegans that is radically different from Orai gating by STIM binding to the N and C termini of Orai in mammals. In addition, we found that the conserved hydrophobic amino acids in the 2–3 loop of Orai1 are important for the oligomerization and gating of channels and are regulated via an intramolecular interaction mechanism mediated by the N and C termini of Orai1. This study identifies a previously unknown SOCE mechanism in C. elegans and suggests that, while the STIM–Orai interaction is conserved between invertebrates and mammals, the gating mechanism for Orai channels differs considerably.

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A Deficiency Screen for Zygotic Loci Required for Establishment and Patterning of the Epidermis in Caenorhabditis elegans

Genetics ◽

10.1093/genetics/146.1.185 ◽

1997 ◽

Vol 146 (1) ◽

pp. 185-206 ◽

Cited By ~ 1

Author(s):

Rebecca M Terns ◽

Peggy Kroll-Conner ◽

Jiangwen Zhu ◽

Sooyoun Chung ◽

Joel H Rothman

Keyword(s):

Caenorhabditis Elegans ◽

Cell Types ◽

Epidermal Cells ◽

Epidermal Differentiation ◽

Apparent Effect ◽

Internal Organs ◽

C Elegans ◽

Seam Cell ◽

Genomic Regions ◽

Caenorhabditis Elegans Genome

To identify genomic regions required for establishment and patterning of the epidermis, we screened 58 deficiencies that collectively delete at least ∼67% of the Caenorhabditis elegans genome. The epidermal pattern of deficiency homozygous embryos was analyzed by examining expression of a marker specific for one of the three major epidermal cell types, the seam cells. The organization of the epidermis and internal organs was also analyzed using a monoclonal antibody specific for epithelial adherens junctions. While seven deficiencies had no apparent effect on seam cell production, 21 were found to result in subnormal, and five in excess numbers of these cells. An additional 23 deficiencies blocked expression of the seam cell marker, in some cases without preventing cell proliferation. Two deficiencies result in multinucleate seam cells. Deficiencies were also identified that result in subnormal numbers of epidermal cells, hyperfusion of epidermal cells into a large syncytium, or aberrant epidermal differentiation. Finally, analysis of internal epithelia revealed deficiencies that cause defects in formation of internal organs, including circularization of the intestine and bifurcation of the pharynx lumen. This study reveals that many regions of the C. elegans genome are required zygotically for patterning of the epidermis and other epithelia.

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Mapping calcium dynamics in a developing tubular structure

10.1101/2020.10.16.342535 ◽

2020 ◽

Author(s):

Jorgen Hoyer ◽

Morsal Saba ◽

Daniel Dondorp ◽

Kushal Kolar ◽

Riccardo Esposito ◽

...

Keyword(s):

Cell Types ◽

Feedback Mechanism ◽

Adult Brain ◽

Actin Dynamics ◽

Cytoskeletal Network ◽

Notochord Cell ◽

Wide Range ◽

Store Operated Calcium Entry ◽

And Function ◽

Cell Intercalation

AbstractCalcium is a ubiquitous and versatile second messenger that plays a central role in the development and function of a wide range of cell types, tissues and organs. Despite significant recent progress in the understanding of calcium (Ca2+) signalling in organs such as the developing and adult brain, we have relatively little knowledge of the contribution of Ca2+ to the development of tubes, structures widely present in multicellular organisms. Here we image Ca2+ dynamics in the developing notochord of Ciona intestinalis. We show that notochord cells exhibit distinct Ca2+ dynamics during specific morphogenetic events such as cell intercalation, cell elongation and tubulogenesis. We used an optogenetically controlled Ca2+ actuator to show that sequestration of Ca2+ results in defective notochord cell intercalation, and pharmacological inhibition to reveal that stretch-activated ion channels (SACs), inositol triphosphate receptor (IP3R) signalling, Store Operated Calcium Entry (SOCE), Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) and gap junctions are required for regulating notochord Ca2+ activity during tubulogenesis. Cytoskeletal rearrangements drive the cell shape changes that accompany tubulogenesis. In line with this, we show that Ca2+ signalling modulates reorganization of the cytoskeletal network across the morphogenetic events leading up to and during tubulogenesis of the notochord. We additionally demonstrate that perturbation of the actin cytoskeleton drastically remodels Ca2+ dynamics, suggesting a feedback mechanism between actin dynamics and Ca2+ signalling during notochord development. This work provides a framework to quantitatively define how Ca2+ signalling regulates tubulogenesis using the notochord as model organ, a defining structure of all chordates.

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Tissue-Specific Transcription Footprinting Using RNA PoI DamID (RAPID) in Caenorhabditis elegans

Genetics ◽

10.1534/genetics.120.303774 ◽

2020 ◽

Vol 216 (4) ◽

pp. 931-945 ◽

Cited By ~ 1

Author(s):

Georgina Gómez-Saldivar ◽

Jaime Osuna-Luque ◽

Jennifer I. Semple ◽

Dominique A. Glauser ◽

Sophie Jarriault ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Rna Polymerase ◽

Cell Types ◽

Neuroendocrine Cells ◽

Cell Physiology ◽

Specific Gene ◽

Cell Content ◽

Tissue Specific ◽

C Elegans ◽

Active Transcription

Differential gene expression across cell types underlies development and cell physiology in multicellular organisms. Caenorhabditis elegans is a powerful, extensively used model to address these biological questions. A remaining bottleneck relates to the difficulty to obtain comprehensive tissue-specific gene transcription data, since available methods are still challenging to execute and/or require large worm populations. Here, we introduce the RNA Polymerase DamID (RAPID) approach, in which the Dam methyltransferase is fused to a ubiquitous RNA polymerase subunit to create transcriptional footprints via methyl marks on the DNA of transcribed genes. To validate the method, we determined the polymerase footprints in whole animals, in sorted embryonic blastomeres and in different tissues from intact young adults by driving tissue-specific Dam fusion expression. We obtained meaningful transcriptional footprints in line with RNA-sequencing (RNA-seq) studies in whole animals or specific tissues. To challenge the sensitivity of RAPID and demonstrate its utility to determine novel tissue-specific transcriptional profiles, we determined the transcriptional footprints of the pair of XXX neuroendocrine cells, representing 0.2% of the somatic cell content of the animals. We identified 3901 candidate genes with putatively active transcription in XXX cells, including the few previously known markers for these cells. Using transcriptional reporters for a subset of new hits, we confirmed that the majority of them were expressed in XXX cells and identified novel XXX-specific markers. Taken together, our work establishes RAPID as a valid method for the determination of RNA polymerase footprints in specific tissues of C. elegans without the need for cell sorting or RNA tagging.

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Inducible Systemic RNA Silencing in Caenorhabditis elegans

Molecular Biology of the Cell ◽

10.1091/mbc.e03-01-0858 ◽

2003 ◽

Vol 14 (7) ◽

pp. 2972-2983 ◽

Cited By ~ 86

Author(s):

Lisa Timmons ◽

Hiroaki Tabara ◽

Craig C. Mello ◽

Andrew Z. Fire

Keyword(s):

Rna Interference ◽

Caenorhabditis Elegans ◽

Rna Silencing ◽

Molecular Mechanisms ◽

Normal Animal ◽

Cell Types ◽

Animal Cells ◽

Tissue Specific ◽

C Elegans ◽

Systemic Rna

Introduction of double-stranded RNA (dsRNA) can elicit a gene-specific RNA interference response in a variety of organisms and cell types. In many cases, this response has a systemic character in that silencing of gene expression is observed in cells distal from the site of dsRNA delivery. The molecular mechanisms underlying the mobile nature of RNA silencing are unknown. For example, although cellular entry of dsRNA is possible, cellular exit of dsRNA from normal animal cells has not been directly observed. We provide evidence that transgenic strains of Caenorhabditis elegans transcribing dsRNA from a tissue-specific promoter do not exhibit comprehensive systemic RNA interference phenotypes. In these same animals, modifications of environmental conditions can result in more robust systemic RNA silencing. Additionally, we find that genetic mutations can influence the systemic character of RNA silencing in C. elegans and can separate mechanisms underlying systemic RNA silencing into tissue-specific components. These data suggest that trafficking of RNA silencing signals in C. elegans is regulated by specific physiological and genetic factors.

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ATM Induces Cell Death with Autophagy in Response to H2O2 Specifically in Caenorhabditis elegans Nondividing Cells

Oxidative Medicine and Cellular Longevity ◽

10.1155/2018/3862070 ◽

2018 ◽

Vol 2018 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Takahito Moriwaki ◽

Akira Yamasaki ◽

Qiu-Mei Zhang-Akiyama

Keyword(s):

Cell Death ◽

Caenorhabditis Elegans ◽

Somatic Cells ◽

Dead Cell ◽

C Elegans ◽

Cell Staining ◽

Dna Damaging Agents ◽

Gfp Reporter ◽

Wide Range ◽

Reporter Assays

Introduction. Ataxia-telangiectasia-mutated (ATM) kinase is a master regulator of the DNA damage response and is directly activated by reactive oxygen species (ROSs) in addition to DNA double-stranded breaks. However, the physiological function of the response to ROSs is not understood. Purpose. In the present study, we investigated how ATM responds to ROSs in Caenorhabditis elegans (C. elegans). Materials and Methods. First, we measured sensitivities of larvae to DNA-damaging agents and ROSs. Next, we analyzed the drug sensitivities of fully matured adult worms, which consist of nondividing somatic cells. Dead cell staining with acridine orange was performed to visualize the dead cells. In addition, we performed GFP reporter assays of lgg-1, an autophagy-related gene, to determine the types of cell death. Results. atm-1(tm5027) larvae showed a wide range of sensitivities to both DNA-damaging agents and ROSs. In contrast, fully matured adult worms, which consist of nondividing somatic cells, showed sensitivity to DNA-damaging agent, NaHSO3, but they showed resistance to H2O2. Dead cell staining and GFP reporter assays of lgg-1 suggest that C. elegans ATM-1 induces the cell death with autophagy in intestinal cells in response to H2O2. Conclusion. We revealed that ATM induces cell death in response to H2O2.

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Characterization of the phototoxicity, chemigenetic profile, and mutational signatures of the chemotherapeutic CX-5461 in Caenorhabditis elegans

10.1101/2019.12.20.884981 ◽

2019 ◽

Author(s):

Frank B. Ye ◽

Akil Hamza ◽

Tejomayee Singh ◽

Stephane Flibotte ◽

Philip Hieter ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Cancer Therapeutics ◽

Copy Number Variations ◽

Factors Affecting ◽

C Elegans ◽

Dna Damaging Agents ◽

Wide Range ◽

Anti Cancer ◽

Exogenous Factors

ABSTRACTNew anti-cancer therapeutics require extensive in vivo characterization to identify endogenous and exogenous factors affecting efficacy, to measure toxicity and mutagenicity, and to determine genotypes resulting in therapeutic sensitivity or resistance. We used Caenorhabditis elegans as a platform with which to characterize properties of anti-cancer therapeutic agents in vivo. We generated a map of chemigenetic interactions between DNA damage response mutants and common DNA damaging agents. We used this map to investigate the properties of the new anti-cancer therapeutic CX-5461. We phenocopied the photoreactivity observed in CX-5461 clinical trials and found that CX-5461 generates reactive oxygen species when exposed to UVA radiation. We demonstrated that CX-5461 is a mutator, resulting in both large copy number variations and a high frequency of single nucleotide variations (SNVs). CX-5461-induced SNVs exhibited a distinct mutational signature. Consistent with the wide range of CX-5461-induced mutation types, we found that multiple repair pathways were needed for CX-5461 tolerance. Together, the data from C. elegans demonstrate that CX-5461 is a multimodal DNA damaging agent with strong similarity to ellipticines, a class of antineoplastic agents, and to anthracycline-based chemotherapeutics.

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A lineage-resolved molecular atlas of C. elegans embryogenesis at single-cell resolution

Science ◽

10.1126/science.aax1971 ◽

2019 ◽

Vol 365 (6459) ◽

pp. eaax1971 ◽

Cited By ~ 65

Author(s):

Jonathan S. Packer ◽

Qin Zhu ◽

Chau Huynh ◽

Priya Sivaramakrishnan ◽

Elicia Preston ◽

...

Keyword(s):

Nervous System ◽

Caenorhabditis Elegans ◽

Single Cell ◽

Molecular Basis ◽

Cell Types ◽

Embryonic Cells ◽

Cell Type ◽

C Elegans ◽

Exact Position ◽

Sister Cells

Caenorhabditis elegans is an animal with few cells but a wide diversity of cell types. In this study, we characterize the molecular basis for their specification by profiling the transcriptomes of 86,024 single embryonic cells. We identify 502 terminal and preterminal cell types, mapping most single-cell transcriptomes to their exact position in C. elegans’ invariant lineage. Using these annotations, we find that (i) the correlation between a cell’s lineage and its transcriptome increases from middle to late gastrulation, then falls substantially as cells in the nervous system and pharynx adopt their terminal fates; (ii) multilineage priming contributes to the differentiation of sister cells at dozens of lineage branches; and (iii) most distinct lineages that produce the same anatomical cell type converge to a homogenous transcriptomic state.

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Bacterial second messenger 3′,5′-cyclic diguanylate attracts Caenorhabditis elegans and suppresses its immunity

Communications Biology ◽

10.1038/s42003-020-01436-9 ◽

2020 ◽

Vol 3 (1) ◽

Author(s):

Joseph Angeloni ◽

Yuqing Dong ◽

Zeneng Wang ◽

Min Cao

Keyword(s):

Caenorhabditis Elegans ◽

High Rate ◽

Bacterial Invasion ◽

Signal Molecules ◽

Cyclic Diguanylate ◽

C Elegans ◽

Low Concentrations ◽

Wide Range ◽

Interkingdom Communication

AbstractCyclic di-nucleotides are important secondary signaling molecules in bacteria that regulate a wide range of processes. In this study, we found that Caenorhabditis elegans can detect and are attracted to multiple signal molecules produced by Vibrio cholerae, specifically the 3′,5′-cyclic diguanylate (c-di-GMP), even though this bacterium kills the host at a high rate. C-di-GMP is sensed through C. elegans olfactory AWC neurons, which then evokes a series of signal transduction pathways that lead to reduced activity of two key stress response transcription factors, SKN-1 and HSF-1, and weakened innate immunity. Taken together, our study elucidates the role of c-di-GMP in interkingdom communication. For C. elegans, bacterial c-di-GMP may serve as a cue that they can use to detect food. On the other hand, preexposure to low concentrations of c-di-GMP may impair their immune response, which could facilitate bacterial invasion and survival.

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Roles of Actin in the Morphogenesis of the Early Caenorhabditis elegans Embryo

International Journal of Molecular Sciences ◽

10.3390/ijms21103652 ◽

2020 ◽

Vol 21 (10) ◽

pp. 3652

Author(s):

Dureen Samandar Eweis ◽

Julie Plastino

Keyword(s):

Cell Division ◽

Caenorhabditis Elegans ◽

Asymmetric Cell Division ◽

Cell Types ◽

Actin Dynamics ◽

Actin Binding ◽

Asymmetric Cell Divisions ◽

C Elegans ◽

Asymmetric Divisions ◽

Different Cell Types

The cell shape changes that ensure asymmetric cell divisions are crucial for correct development, as asymmetric divisions allow for the formation of different cell types and therefore different tissues. The first division of the Caenorhabditis elegans embryo has emerged as a powerful model for understanding asymmetric cell division. The dynamics of microtubules, polarity proteins, and the actin cytoskeleton are all key for this process. In this review, we highlight studies from the last five years revealing new insights about the role of actin dynamics in the first asymmetric cell division of the early C. elegans embryo. Recent results concerning the roles of actin and actin binding proteins in symmetry breaking, cortical flows, cortical integrity, and cleavage furrow formation are described.

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The lysosomal cathepsin protease CPL-1 plays a leading role in phagosomal degradation of apoptotic cells in Caenorhabditis elegans

Molecular Biology of the Cell ◽

10.1091/mbc.e14-01-0015 ◽

2014 ◽

Vol 25 (13) ◽

pp. 2071-2083 ◽

Cited By ~ 16

Author(s):

Meng Xu ◽

Yubing Liu ◽

Liyuan Zhao ◽

Qiwen Gan ◽

Xiaochen Wang ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Apoptotic Cell ◽

Cell Types ◽

Apoptotic Cells ◽

C Elegans ◽

Lysosomal Protease ◽

Leading Role ◽

Lysosomal Proteases ◽

Cell Corpse

During programmed cell death, the clearance of apoptotic cells is achieved by their phagocytosis and delivery to lysosomes for destruction in engulfing cells. However, the role of lysosomal proteases in cell corpse destruction is not understood. Here we report the identification of the lysosomal cathepsin CPL-1 as an indispensable protease for apoptotic cell removal in Caenorhabditis elegans. We find that loss of cpl-1 function leads to strong accumulation of germ cell corpses, which results from a failure in degradation rather than engulfment. CPL-1 is expressed in a variety of cell types, including engulfment cells, and its mutation does not affect the maturation of cell corpse–containing phagosomes, including phagosomal recruitment of maturation effectors and phagosome acidification. Of importance, we find that phagosomal recruitment and incorporation of CPL-1 occurs before digestion of cell corpses, which depends on factors required for phagolysosome formation. Using RNA interference, we further examine the role of other candidate lysosomal proteases in cell corpse clearance but find that they do not obviously affect this process. Collectively, these findings establish CPL-1 as the leading lysosomal protease required for elimination of apoptotic cells in C. elegans.

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