Cassada RC (1991) International C. elegans Meeting "I. TS EMB-MUTANT ANALYSIS EXTENDED. II. RECONSIDERATION OF TS MUTANTS FOR DEFINING GENE FUNCTION & SATURATING GENETICALLY."

Cassada RC

[International C. elegans Meeting, 1991]

i. Early TSP's before fertilization were defined by shifting single ts emb adults to 25 C; laying and hatching kinetics were followed, a la Vanderslice & Hirsh (1976), also downshifting to determine TSP- begin. The number of fertilized eggs in the gonad vs. age was determined, as well as brood size. ii. ts emb adults from 16 C were allowed to lay eggs at various temps to obtain: * critical temp. (50% lethality) If all alleles have same crit. temp., maybe gene product is not ts, rather function is needed only above a certain temp., e.g. if another product (isozyme?) is naturally ts. Then the null phenotype will be ts, and ts alleles 25x more frequent than if missense. Instead, alleles of the highly mutable emb-9 have different crit. temps. (ditto emb-6). ** a maximum ts-phenotyPe, constant at all temps. above a minimum may be the null phenotype. Perdurance is less a problem than for non-ts alleles! *** neomorphs at intermediate temps. (For most embs, any Acc phenotype had a lower crit. temp.) iii. For paternal-effect tests, the male ts phenotype and TSP were defined for several embs. Also chromosome loss generated males (Him- effect) among leakies, making one emb appear hermaphrodite specific. iv. Just for fun, N2 and Freiburg (RC301) were crossed to look for growth of Fl's and segregants ar 28 C. One might expect growth if different genes limited the two strains, but no such luck, so their last common ancestor probably didn't grow at 28 C. II. DISCUSSION INVITED ON POTENTIAL & PROBLEMS OF TS MUTANTS: Are most ts embs just for housekeeping? Is polyphasy interesting? It can only be analyzed with ts mutants and pulses. How to identify interesting genes and genetically saturate? Design (localized?) screens to detect and study leakier mutants, not just tightest ones.

van Auken KM et al. (1995) International C. elegans Meeting "TOWARDS ANALYSIS OF THE NOB-1 PHENOTYPE"

van Auken KM, Wood WB, Edgar LG

[International C. elegans Meeting, 1995]

nob-1(III) is defined by three alleles: ct223 and ct351, which result in L1 lethality with the Nob phenotype (severely deformed posterior), and ct230, which results in viable animals with lumpy, misshapen tails (see Edgar et al., 1993 Meeting Abstracts). Staining of late stage Nob-1 embryos with the hypodermal-specific antibody MH27 reveals disorganized and misshapen posterior ventral and lateral hypodermis. Examination of the seam cell-specific marker wIs3, shows variable loss of expression suggesting that some posterior seam cells are either missing or fail to differentiate properly. To identify possible missing cells, we analyzed the complete embryonic lineages that give rise to posterior hypodermis (ABarpp, ABplap, ABprap, Cpa, Caa with the exception of the late divisions that give rise to V5 and Q), as well as the E and early MS lineages, but could find no lineage defects. Possibly, therefore, the Nob-1 phenotype results from failed differentiation of some posterior cells rather than from defects in posterior lineages. Phenotypic analysis of viable ct223/ct230 and ct230/ct230 animals reveals additional post-embryonic defects, which we are still characterizing. Males display crumpled spicules and loss of ray sensilla, while hermaphrodites are variably Egl. Both sexes have phasmid-filling defects and are constipated. These observations suggest possible differentiation defects in B, V5, V6, T and neuronal (HSN and PHA/B) lineages. Finally, since nob-1 resides in a gap on the physical map, we are making use of the 75% viabiity of heteroallelic animals to perform non-complementation screens with the mutator strain, RW7097, to isolate a Tc1 insertion. To date, we have screened approximately 20,000 haploid chromosomes with no luck, although we have introduced new DNA polymorphisms into the region, which we are in the process of mapping.

Swierczek, Nicholas et al. (2009) International Worm Meeting "High-throughput mostly-automated quantification of subtle behaviors."

Kerr, Rex A., Swierczek, Nicholas, Rankin, Catharine, Giles, Andrew

[International Worm Meeting, 2009]

We have designed and built a tool, the Multi-Worm Tracker (MWT), that enables analysis of subtle behavioral defects at speeds suitable for forward genetic screening. The Multi-Worm Tracker images a plate of worms with a 4 megapixel camera running at 10 Hz, delivers stimuli at user-specified intervals, and uses real-time image analysis to extract and save key parameters for each worm. We typically track dozens of worms on a single 5 cm plate; this allows rapid screening of baseline behaviors such as speed and turning frequency; of stimulus-driven behaviors such as reversals induced by tap to the plate; and of adaptations to repeated stimuli. Due to the low magnification, small-scale features such as shape of body bends cannot be reliably quantified; these require a high-magnification single-worm tracker. We are using the MWT to quantify in detail the tap habituation behavior of C. elegans. For instance, we have recorded from tens of thousands of wild-type worms to generate highly reliable statistics, and carefully sampled inter-stimulus intervals from seconds through minutes. We are also preparing to screen for tap habituation mutants, which would be highly impractical without an automated system. We demonstrate that we can detect and cluster existing mutants both for baseline behavior and tap habituation, and use analysis of wild-type variability to design an effective protocol for conducting a large-scale screen. (With luck, we will present preliminary results from a pilot screen.) Although the MWT was designed with tap habituation in mind, the system is generally useful for rapid quantification of behavior. We therefore have taken a number of steps to enable other labs to set up similar systems. The MWT software is open source (but requires commercial run-time libraries) and after the meeting will be available at http://sourceforge.net/projects/mwt; parts lists and installation instructions are also available. We also provide post-acquisition analysis software that plots or saves summary data with more sophistication than is possible online, and allows browsing of the data in 2D map format for easy manual validation of the results. We have conducted proof-of-principle experiments for foraging and chemotaxis; we hope that other labs will further develop these assays using the MWT or a similar system.

Asim A Beg et al. (2002) West Coast Worm Meeting "A GABA-gated non-selective cation channel in C. elegans."

Asim A Beg, Erik M Jorgensen

[West Coast Worm Meeting, 2002]

-aminobutyric acid A receptors are members of the ligand-gated ion channel superfamily that mediate inhibitory neurotransmission in both vertebrates and invertebrates. GABA binding to these receptors causes the opening of an anion-selective channel that predominantly conducts chloride ions, thereby hyperpolarizing cells. However, there are numerous mechanisms by which GABA can serve as an excitatory neurotransmitter. To date, the molecular identity of an excitatory ionotropic GABA receptor has remained elusive.

Sarah Straud Anselmo et al. (2002) West Coast Worm Meeting "Developing a screen for HERG blocking drugs using the C. elegans pharynx"

Sarah Straud Anselmo, Leon Avery

[West Coast Worm Meeting, 2002]

The HERG (human ether-a-go-go related gene) potassium channel is expressed in the human heart and is responsible for proper repolarization of cardiac muscle at the end of the action potential. Compromising HERG function gives rise to Long QT Syndrome, a disorder in which one is predisposed to ventricular fibrillation and sudden death. Many drugs, including Seldane (an anti-histamine) and Propulsid (an acid reflux inhibitor), block HERG as an unwanted side effect and consequently have been taken off of the market.

Schulze, Jens et al. (2011) International Worm Meeting "Plectus - a stepping stone in embryonic cell lineage evolution of nematodes."

Schulze, Jens, Schierenberg, Einhard, Uenk, Jana

[International Worm Meeting, 2011]

The outstanding feature of Caenorhabditis elegans is its invariant cell lineage generating blastomeres of fixed identity. In contrast, the early embryo of the basal nematode Enoplus brevis shows only symmetric early cell divisions and the only identifiable lineage generates the gut (Voronov and Panchin, 1998; Development 15:143-150). How an organism with invariant development like C. elegans can evolve from an ancestor with an indeterminate mode remains a central question. Our lineage analysis of Plectus sambesii offers an important piece of the puzzle. Unexpectedly, cell arrangements in the AB lineage of this species are rather variable but merge into six different stereotyped patterns which all are compatible with normal development. However, one of the six patterns is prevalent and shows the same early cell arrangement as found in C elegans. This observation can be interpreted as an evolutionary trend from an indeterminate to an invariant pattern of cells. Furthermore, our cell lineage analysis of hypodermal and pharyngeal cells reveals that AB cell fates in all six stereotyped patterns are determined in a position-dependent rather than a lineage-dependent manner resulting in a single distinct right-handed juvenile pattern. This differs fundamentally from the lineage-dependent C. elegans mechanism where a left-handed situs inversus can be generated experimentally by switching positions of AB cells in the early embryo (Wood, 1991; Nature 349:536-38).

Marc Hammarlund et al. (2002) West Coast Worm Meeting "Mapping a new class of Ric mutants using an improved SNP mapping technique"

Tracey Harrach, Erik M Jorgensen, Marc Hammarlund

[West Coast Worm Meeting, 2002]

In an effort to identify additional molecules that function in synaptic transmission we have conducted several large screens for Aldicarb resistance. Aldicarb is an inhibitor of acetylcholinesterase and mutants that secrete less neurotransmitter are resistant to this drug. The mutants obtained from these screens fall into two classes: those that are uncoordinated, and those that are not uncoordinated.

Laskova, Valeriya et al. (2013) International Worm Meeting "Mapping the entire connectome of C. elegans L1 larvae."

Zhen, Mei, Kasthuri, Bobby, Shalek, Richard, Lichtman, Jeff, Samuel, Aravi, Pfister, Hanspeter, Laskova, Valeriya, Kaynig-Fittkau, Verena, Wen, Quan, Berger, Daniel, Guan, Asuka

[International Worm Meeting, 2013]

To investigate the poorly understood mechanisms of development and function of the nervous system, connections between neurons must be deciphered first. The adult nervous system of C. elegans was mapped to near completion by Dr. John White and his colleagues in 1970s. However, neuronal wiring in C. elegans larvae differs from that of an adult, since it undergoes multiple rounds of neuronal birth, apoptosis and rewiring during development. We utilize an Automatic Tape-Collecting Ultramicrotome (ATUM) to section an entire L1 stage animal into thousands cross-sections, followed by the automated imaging with a scanning electron microscope (SEM) to map an entire neuronal wiring diagram with synaptic resolution. The acquired wiring diagram will guide and be validated by calcium imaging to map the functional connection of the juvenile motor circuit.

Cynthia Wagner et al. (2006) Development & Evolution Meeting "The Role of gak-1 in Meiosis"

Gabrielle Howard, Cynthia Wagner, Rachel Webster, Judith Yanowitz, Dana Hubbard

[Development & Evolution Meeting, 2006]

We performed an RNAi feeding screen to identify chromatin associated gene products, that when mutant, result in an increase in recombination. We screened greater than 300 genes and identified 20 genes that give between a two-fold and four-fold enhancement of recombination. We are in the process of characterizing one of these genes, gak-1. gak-1 has previously been reported to function in meiotic chromosome segregation, including an increase in X-chromosome nondisjunction, which results in a him phenotype. We will present our cytological analysis on gak-1 mutants as well as detailed information on recombination effects in gak-1 mutants.

Kikuchi, Taisei et al. (2021) International Worm Meeting "An alternative ERGO-1 pathway in a sibling species of C. elegans, C. inopinata"

Sugimoto, Asako, Kikuchi, Taisei, Tanaka, Ryusei, Yoshida, Akemi, Sun, Simo, Hunt, Vicky

[International Worm Meeting, 2021]

The recent discovery of Caenorhabditis inopinata, a sibling species of C. elegans, provides an opportunity to investigate the conservation and diversification of small RNA (sRNA) pathways in two closely related species. We report that the microRNA, PIWI and male-expressed ALG-3/4 siRNA pathways are highly conserved in C. inopinata and C. elegans. However, C. inopinata has lost key components of the female-expressed ERGO-1 siRNA pathway associated with the clearance of transcripts. C. inopinata do not express the ERGO-1 associated 26G primary siRNAs and have similar sRNA expression profiles to C. elegans ergo-1 and eri-6 mutants. However, expression of the downstream secondary 22G siRNAs is retained in C. inopinata. We have identified a class of 22U siRNAs that are candidates for an alternative ERGO-1 siRNA. The 22U loci overlap with 22Gs and target similar genes but, unlike C. elegans 26Gs, are not dicer-processed. We also identified an Argonaute protein with an expression profile similar to the C. elegans ERGO-1, as a candidate for an alternative ERGO-1 Argonaute protein. Our results indicate that there has been a partial loss of the ERGO-1 pathway in C. inopinata and an alternative ERGO-1 pathway has evolved.