Muscle contraction phenotypic analysis enabled by optogenetics reveals functional relationships of sarcomere components in Caenorhabditis elegans

Hwang, Hyundoo; Barnes, Dawn E.; Matsunaga, Yohei; Benian, Guy M.; Ono, Shoichiro; Lu, Hang

doi:10.1038/srep19900

Cited by 34 publications

(29 citation statements)

References 31 publications

(43 reference statements)

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“…The results indicated no significant difference in the contraction and relaxation rates of both experimental conditions. Moreover, our data matched the contraction and relaxation rates reported by optogenetics for unexposed worms [31] . This showed the potential use of our technique in studying the overall muscle kinetics of wild-type and potentially other worm strains.…”

Section: Effect Of Ef Direction On the Egg-laying Behavioursupporting

confidence: 87%

Electric Egg-Laying: Effect of Electric Field in a Microchannel onC. elegansEgg-Laying Behavior

Youssef

Archonta

Kubiseski

et al. 2020

Preprint

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In this paper, the novel effect of electric field (EF) on adult C. elegans egg-laying in a microchannel is discovered and correlated with neural and muscular activities. The quantitative effects of worm aging and EF strength, direction, and exposure duration on egg-laying is studied phenotypically using egg-count, body length, head movement, and transient neuronal activity readouts. Electric egg-laying rate increases significantly when worms face the anode and the response is EF-dependent, i.e. stronger (6V/cm) and longer EF (40s) exposure result in a shorter egg laying response duration. Worm aging significantly deteriorates the electric egg-laying behaviour with 88% decrease in the egg-count from Day-1 to Day-4 post young-adult stage. Fluorescent imaging of intracellular calcium dynamics in the main parts of the egg-laying neural circuit demonstrate the involvement and sensitivity of the serotonergic hermaphrodite specific neurons (HSNs), vulva muscles, and ventral cord neurons to the EF. HSN mutation also results in a reduced rate of electric egg-laying allowing the use of this technique for cellular screening and mapping of the neural basis of electrosensation in C. elegans. This novel assay can be parallelized and performed in a high-throughput manner for drug and gene screening applications.

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Section: Effect Of Ef Direction On the Egg-laying Behavioursupporting

confidence: 87%

Electric Egg-Laying: Effect of Electric Field in a Microchannel onC. elegansEgg-Laying Behavior

Youssef

Archonta

Kubiseski

et al. 2020

Preprint

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“…Kinetic analysis of muscle contraction and relaxation was performed using a microfluidic device to trap worms and an optical system for light illumination and imaging as described previously [Stirman et al, ] with an optimization for analysis of muscle contractility phenotypes as described [Hwang et al, ]. Briefly, two‐layer microfluidic devices made of polydimethylsiloxane (PDMS; Sylgard 184, Dow‐Corning Corp., Midland, MI) with 16 straight microchannels with membrane valves at the entrance and the exit were used to trap worms.…”

Section: Methodsmentioning

confidence: 99%

“…Expression of channelrhodopsin‐2 did not alter sarcomere organization and worm motility (our unpublished data). Recently, we utilized this method to characterize 16 muscle‐affecting mutants and verified that this is an excellent non‐invasive method for quantitative analysis of muscle contractility [Hwang et al, ].…”

Section: Troponin I N‐terminal Extension Is Required For Regulation Omentioning

confidence: 99%

“…Previous phenotypic analysis of unc‐27 mutants suggest that they are constitutively contracted under normal culture conditions [Burkeen et al, ]. In the optogenetic experiments, reduction of the body area upon light stimulation was slightly less in unc‐27 than in wild‐type, suggesting that the unc‐27 mutant had been partially pre‐contracted before light stimulation [Hwang et al, ]. Basal contraction levels can potentially influence the rates of light‐induced contraction and relaxation.…”

Section: Troponin I N‐terminal Extension Is Required For Regulation Omentioning

confidence: 99%

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Molecular evolution of troponin I and a role of its N‐terminal extension in nematode locomotion

et al. 2016

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Summary The troponin complex, composed of troponin T (TnT), troponin I (TnI), and troponin C (TnC), is the major calcium-dependent regulator of muscle contraction, which is present widely in both vertebrates and invertebrates. Little is known about evolutionary aspects of troponin in the animal kingdom. Using a combination of data mining and functional analysis of TnI, we report evidence that an N-terminal extension of TnI is present in most of bilaterian animals as a functionally important domain. Troponin components have been reported in species in most of representative bilaterian phyla. Comparison of TnI sequences shows that the core domains are conserved in all examined TnIs, and that N- and C-terminal extensions are variable among isoforms and species. In particular, N-terminal extensions are present in all protostome TnIs and chordate cardiac TnIs but lost in a subset of chordate TnIs including vertebrate skeletal-muscle isoforms. Transgenic rescue experiments in C. elegans striated muscle show that the N-terminal extension of TnI (UNC-27) is required for coordinated worm locomotion but not in sarcomere assembly and single muscle-contractility kinetics. These results suggest that N-terminal extensions of TnIs are retained from a TnI ancestor as a functional domain.

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“…Severe lev-11 mutants are paralyzed and arrested at an embryonic stage (Williams and Waterston 1994). Some of lev-11 mutants are resistant to levamisole, an agonist for acetylcholine receptor that induces muscle contraction (Lewis et al 1980a; Lewis et al 1980b), because slow muscle relaxation occurs while cholinergic neurons are active (Hwang et al 2016). Splicing of two mutually exclusive exon 7s is differentially regulated in the head and body regions of the body wall muscle cells to express LEV-11O in the head muscles and LEV-11A in the rest of the body wall muscles, where these isoforms differentially regulate muscle contractility (Barnes et al 2018).…”

Section: Introductionmentioning

confidence: 99%

Alternative splicing of the Caenorhabditis elegans lev‐11 tropomyosin gene is regulated in a tissue‐specific manner

2018

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Tropomyosin isoforms contribute to generation of functionally divergent actin filaments. In the nematode Caenorhabditis elegans, multiple isoforms are produced from lev-11, the single tropomyosin gene, by combination of two separate promoters and alternative pre-mRNA splicing. In this study, we report that alternative splicing of lev-11 is regulated in a tissue-specific manner so that a particular tropomyosin isoform is expressed in each tissue. Reverse-transcription polymerase chain reaction analysis of lev-11 mRNAs confirms five previously reported isoforms (LEV-11A, LEV-11C, LEV-11D, LEV-11E and LEV-11O) and identifies a new sixth isoform LEV-11T. Using transgenic alternative-splicing reporter minigenes, we find distinct patterns of preferential exon selections in the pharynx, body wall muscles, intestine and neurons. The body wall muscles preferentially process splicing to produce high-molecular-weight isoforms, LEV-11A, LEV-11D and LEV-11O. The pharynx specifically processes splicing to express a low-molecular-weight isoform LEV-11E, whereas the intestine and neurons process splicing to express another low-molecular-weight isoform LEV-11C. The splicing pattern of LEV-11T was not predominant in any of these tissues, suggesting that this is a minor isoform. Our results suggest that regulation of alternative splicing is an important mechanism to express proper tropomyosin isoforms in particular tissue and/or cell types in C. elegans.

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Muscle contraction phenotypic analysis enabled by optogenetics reveals functional relationships of sarcomere components in Caenorhabditis elegans

Cited by 34 publications

References 31 publications

Electric Egg-Laying: Effect of Electric Field in a Microchannel onC. elegansEgg-Laying Behavior

Electric Egg-Laying: Effect of Electric Field in a Microchannel onC. elegansEgg-Laying Behavior

Molecular evolution of troponin I and a role of its N‐terminal extension in nematode locomotion

Alternative splicing of the Caenorhabditis elegans lev‐11 tropomyosin gene is regulated in a tissue‐specific manner

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