Two K2P Channels, TWK-46 and TWK-26, Do Not Affect C. elegans Egg-Laying Behavior

Sep 10, 2026

Two-pore domain potassium (K2P) channels regulate resting membrane potential and influence diverse physiological processes across species. The C. elegans genome contains 46 K2P-encoding genes, but most functions remain unclear. This study examined twk-26 and twk-46 in the egg-laying neural circuit using CRISPR/Cas9-generated null mutants. Multiple behavioral assays detected no significant differences in egg-laying output or timing between mutants and wild-type worms.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages7

Reading Time18 min

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Abstract

Experimental Approach

CRISPR/Cas9 null mutants

Results: Egg-laying behavior assays

Eggs laid on and off food

Unlaid eggs in uterus

Timing of egg-laying events

Serotonin responsiveness

Background and Circuit Rationale

1/27/2025-Open Access Two K2P Channels, TWK-46 and TWK-26 do not affect C. elegans EggLaying Behavior Li Chen1, Isabel Beets1, William Schafer1,2§ 1Biology, KU Leuven, Leuven, Flanders, Belgium 2Neurobiology, MRC Laboratory of Molecular Biology, Cambridge, England, United Kingdom § To whom correspondence should be [addressed: william.schafer@kuleuven.be](addressed: william.schafer@kuleuven.be) Abstract Two-pore domain potassium channels, also known as K2P channels, play vital roles in maintaining the resting membrane potential in excitable cells, affecting a variety of physiological processes across species. The Caenorhabditis elegans (C. elegans) genome contains 46 different K2P-encoding genes, yet most of their functions remain unknown. Here, we have investigated the possible roles of two C. elegans K2P channel genes – twk-26 and twk-46 – that are expressed in the egg-laying neural circuit by characterizing the egg-laying behavior of null mutants generated by CRISPR/Cas9 gene editing. However, using a variety of assays, we did not observe significant differences in egg-laying behavior between twk-26 and twk-46 mutants and wild-type worms. 1/27/2025-Open Access Figure 1. Characterization of egg-laying behavior in two C. elegans K2P channel mutants: 1/27/2025-Open Access A. Graphic representation of CRISPR full knockout alleles of twk-26(ibt22) and twk-46(ibt17) (scale bar: 100bp) . B. Quantification of the number of eggs laid on food within 2 hours (N2 : n=69; twk-26: n=70; twk-46: n=61 animals) and off food within 1 hour (N2 : n=55; twk-26: n=63; twk-46: n=62 animals) . Statistical significance was evaluated using a one-way ANOVA test, ns: p>0.05. C. Quantification of the number of unlaid eggs within uterus (N2 : n=72; twk-26: n=71; twk-46: n=70 animals) . Statistical significance (one-way ANOVA test), ns: p>0.05. D. Quantitative analysis of the timing of egg-laying events. Individual animals were tracked on food for 6 hours, and the intervals between egg-laying events were measured. The left panel represents the histogram of log intervals of 10 individual animals of each genotype, with the maximum likelihood estimation (Waggoner et al., 1998; Zhou et al, 1998) of intra-and inter-cluster time constants as well as three parameters of the two-state egg-laying model (p: probability of egg-laying events during active egg-laying phase, λ1: rate constant of inactive egg-laying phase, and λ2: rate constant of active egg-laying phase) . The blue and green dashed lines indicate the intra-and inter-cluster time constants, respectively. The right panel plots the inter- and inter-cluster time constants determined by the same method per individual animal. Estimated parameters were not statistically different between genotypes according to Brown-Forsythe and Welch ANOVA, with Dunnett's post-hoc test. E. Quantification of the number of laid eggs in response to different exogenous serotonin concentrations within 1 hour in the absence of food (n= 15 animals for each genotype) . Significance was tested using an Aligned Ranked two-way ANOVA test, ns: p>0.05. For all graphs, error bars represent standard error of the mean (SEM) . Description Two-pore domain potassium channels are a class of 4-transmembrane spanning K+ channels, with two pore-forming domainsin each subunit (Natale et al., 2021) . In mammals, some K2P genes have been discovered as novel anesthesia targets due to their activation by certain anesthetics and broad expression in the central nervous system (Steinberg et al., 2015; Talley et al., 2001) . Surprisingly, the most diversified K2P gene family has been found in the model organism C. elegans (Hobert, 2013), which has a very compact nervous system with only 302 neurons in hermaphrodites (White et al., 1986) . Different studies have indicated that K2P genes are involved in regulating various activities in C. elegans. For instance, twk-40 impacts both defecation and locomotion by regulating the excitabi

FAQs

Why study K2P channels twk-26 and twk-46 in C. elegans egg-laying?

K2P channels help set resting membrane potential, yet most C. elegans K2P gene functions are unknown. The study targets twk-26 and twk-46 because they are expressed in the egg-laying neural circuit.

How were twk-26 and twk-46 functions tested?

Null mutants were generated with CRISPR/Cas9 gene editing, and egg-laying behavior was assessed using multiple assays to compare mutants with wild-type worms.

What were the main findings about egg-laying behavior in twk-26 and twk-46 mutants?

Across the tested assays, egg-laying behavior and event timing showed no significant differences between twk-26 and twk-46 mutants and wild-type animals.

Sep 10, 2026

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