A Rapid Protocol for Integrating Extrachromosomal Arrays With High Transmission Rate into the &lt;em&gt;C. elegans&lt;/em&gt; Genome

Mariol, Marie-Christine; Walter, Ludivine; Bellemin, Stéphanie; Gieseler, Kathrin

doi:10.3791/50773

Cited by 48 publications

(35 citation statements)

References 13 publications

(18 reference statements)

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“…ceh-17:: GCaMP6 was injected into N2 animals together with ceh-17::dsred (Suo et al, 2006) and lin-44::gfp (Murakami et al, 2001) as a coinjection marker. The resulting strain was subjected to UV irradiation to integrate the extrachromosomal array into the genome (Mariol et al, 2013). The strain carrying the integrated transgene was backcrossed to N2 animals three times and then crossed to CB4088 him-5.…”

Section: Methodsmentioning

confidence: 99%

Sexually Dimorphic Regulation of Behavioral States by Dopamine in Caenorhabditis elegans

et al. 2019

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Sex differences in behavior allow animals to effectively mate and reproduce. However, the mechanism by which biological sex regulates behavioral states, which underlie the regulation of sex-shared behaviors, such as locomotion, is largely unknown. In this study, we studied sex differences in the behavioral states of Caenorhabditis elegans and found that males spend less time in a low locomotor activity state than hermaphrodites and that dopamine generates this sex difference. In males, dopamine reduces the low activity state by acting in the same pathway as polycystic kidney disease-related genes that function in male-specific neurons. In hermaphrodites, dopamine increases the low activity state by suppression of octopamine signaling in the sex-shared SIA neurons, which have reduced responsiveness to octopamine in males. Furthermore, dopamine promotes exploration both inside and outside of bacterial lawn (the food source) in males and suppresses it in hermaphrodites. These results demonstrate that sexually dimorphic signaling allows the same neuromodulator to promote adaptive behavior for each sex.

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Section: Methodsmentioning

confidence: 99%

Sexually Dimorphic Regulation of Behavioral States by Dopamine in Caenorhabditis elegans

et al. 2019

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“…Additionally, the following multi-copy integrated transgenes were used: adIs2122(lgg-1p::GFP::lgg-1 + rol-6 (su1006)) [102], bpIs151 (sqst-1p::sqst-1::GFP + unc-76(+)) [51], zcIs9 (P hsp-60 ::gfp::unc-54 3'UTR) [14], zcIs13 (P hsp-6 ::gfp::unc-54 3'UTR) [14], zcIs18 (P ges-1 ::gfp(cyt)) [103], bcIs79 (P let-858:: gfp mt ::let-858 3'UTR + rol-6(su1006)), bcIs78 (P myo-3 ::gfp mt ::unc-54 3'UTR + rol-6(su1006)) [46]. The strains MOC92 bicIs10(hsp-1::tagRFP::unc-54 3'UTR) and MOC119 bicIs12(ttr-45p:: tagRFP::ttr-45 3'UTR) were generated in the Casanueva lab by gonadal microinjection of plasmids pMOC1 and pMOC2, respectively followed by genome integration via UV irradiation using a Stratagene UV Crosslinker (Stratalinker) [104]. The irradiation dose was 35mJ/cm 2 corresponding to Stratalinker power set up at 350.…”

Section: General C Elegans Methods and Strainsmentioning

confidence: 99%

Autophagy compensates for defects in mitochondrial dynamics

et al. 2020

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Compromising mitochondrial fusion or fission disrupts cellular homeostasis; however, the underlying mechanism(s) are not fully understood. The loss of C. elegans fzo-1 MFN results in mitochondrial fragmentation, decreased mitochondrial membrane potential and the induction of the mitochondrial unfolded protein response (UPR mt). We performed a genome-wide RNAi screen for genes that when knocked-down suppress fzo-1 MFN (lf)-induced UPR mt. Of the 299 genes identified, 143 encode negative regulators of autophagy, many of which have previously not been implicated in this cellular quality control mechanism. We present evidence that increased autophagic flux suppresses fzo-1 MFN (lf)-induced UPR mt by increasing mitochondrial membrane potential rather than restoring mitochondrial morphology. Furthermore, we demonstrate that increased autophagic flux also suppresses UPR mt induction in response to a block in mitochondrial fission, but not in response to the loss of spg-7 AFG3L2 , which encodes a mitochondrial metalloprotease. Finally, we found that blocking mitochondrial fusion or fission leads to increased levels of certain types of triacylglycerols and that this is at least partially reverted by the induction of autophagy. We propose that the breakdown of these triacylglycerols through autophagy leads to elevated metabolic activity, thereby increasing mitochondrial membrane potential and restoring mitochondrial and cellular homeostasis.

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“…Wild-type: (N2 Bristol); Transgenic strains: NM2415 (lin-15B(n765); jsIs68[Prab-3::GFP::rab-3 + lin-15(+)]), LC108 (vIs69 [pCFJ90(Pmyo-2::mCherry + Punc-119::sid-1)]) OH441(punc-119::GFP); Mutant strains: RB652 (puf-7(ok361)) and JK3231(puf-8(q725)) were obtained from the Caenorhabditis Genetics Center (University of Minnesota, Minneapolis, MN). The transgenic strain CQ574 (lin-15B(n765); jsIs682 ]; wqIs3 [Prab3::mCherry]) was generated by UV integration (Mariol et al, 2013) of NM2415 (lin-15B(n765); jsIs68[Prab-3::GFP::rab-3 + lin-15(+)]) animals microinjected with a Prab3::mCherry transgenic construct, followed by 3 rounds of outcrossing with wild-type (N2 Bristol) worms.…”

Section: Experimental Model and Subject Details C Elegans Geneticsmentioning

confidence: 99%

Profiling of presynaptic mRNAs reveals a role for axonal PUMILIOs in associative memory formation

Arey

Kaletsky

Murphy

2019

Preprint

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Presynaptic protein synthesis is important in the adult central nervous system; however, the set of mRNAs localized to presynaptic areas has yet to be identified in any organism. We differentially labeled somatic and synaptic compartments nervous system-wide in adult C. elegans and isolated synaptic regions for deep sequencing. Analysis of the synaptic transcriptome reveals that synaptic transcripts are predicted to have specialized functions in neurons. Differential expression analysis identified 543 genes enriched in synaptic regions relative to somatic regions, with synaptic functions conserved in higher organisms. We find that mRNAs for pumilio RNA-binding proteins are abundant in synaptic regions, which we confirmed through high-sensitivity in situ hybridization. We identified a new role for the PUM2 orthologs puf-7/8 as repressors of memory formation through regulation of the mitochondrial dynamics regulator, mff-1. Identification of presynaptic mRNAs provides insight into mechanisms that regulate synaptic function and behavior.

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A Rapid Protocol for Integrating Extrachromosomal Arrays With High Transmission Rate into the <em>C. elegans</em> Genome

Cited by 48 publications

References 13 publications

Sexually Dimorphic Regulation of Behavioral States by Dopamine in Caenorhabditis elegans

Sexually Dimorphic Regulation of Behavioral States by Dopamine in Caenorhabditis elegans

Autophagy compensates for defects in mitochondrial dynamics

Profiling of presynaptic mRNAs reveals a role for axonal PUMILIOs in associative memory formation

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