Mathers PH et al. (1997) Nature "The Rx homeobox gene is essential for vertebrate eye development."

Mahon KA, Grinberg A, Mathers PH, Jamrich M

[Nature, 1997]

Development of the vertebrate eye requires a series of steps including specification of the anterior neural plate, evagination of the optic vesicles from the ventral forebrain, and the cellular differentiation of the lens and retina. Homeobox-containing genes, especially the transcription regulator Pax6, play a critical role in vertebrate and invertebrate eye formation. Mutations in Pax6 function result in eye malformations known as Aniridia in humans and Small eye syndrome in mice. The Drosophila homologue of Pax6, eyeless, is also necessary for correct invertebrate eye development, and its misexpression leads to formation of ectopic eyes in Drosophila. Here we show that a conserved vertebrate homeobox gene, Rx, is essential for normal eye development, and that its misexpression has profound effects on eye morphology. Xenopus embryos injected with synthetic Rx RNA develop ectopic retinal tissue and display hyperproliferation in the neuroretina. Mouse embryos carrying a null allele of this gene do not form optic cups and so do not develop eyes. The Rx gene family plays an important role in the establishment and/or proliferation of retinal progenitor cells.

Solorzano E et al. (2011) BMC Evol Biol "Shifting patterns of natural variation in the nuclear genome of caenorhabditis elegans."

Bergeron RD, Sung W, Okamoto K, Thomas WK, Solorzano E, Datla P

[BMC Evol Biol, 2011]

BACKGROUND: Genome wide analysis of variation within a species can reveal the evolution of fundamental biological processes such as mutation, recombination, and natural selection. We compare genome wide sequence differences between two independent isolates of the nematode Caenorhabditis elegans (CB4856 and CB4858) and the reference genome (N2). RESULTS: The base substitution pattern when comparing N2 against CB4858 reveals a transition over transversion bias (1.32:1) that is not present in CB4856. In CB4856, there is a significant bias in the direction of base substitution. The frequency of A or T bases in N2 that are G or C bases in CB4856 outnumber the opposite frequencies for transitions as well as transversions. These differences were not observed in the N2/CB4858 comparison. Similarly, we observed a strong bias for deletions over insertions in CB4856 (1.44: 1) that is not present in CB4858. In both CB4856 and CB4858, there is a significant correlation between SNP rate and recombination rate on the autosomes but not on the X chromosome. Furthermore, we identified numerous significant hotspots of variation in the CB4856-N2 comparison.In both CB4856 and CB4858, based on a measure of the strength of selection (ka/ks), all the chromosomes are under negative selection and in CB4856, there is no difference in the strength of natural selection in either the autosomes versus X or between any of the chromosomes. By contrast, in CB4858, ka/ks values are smaller in the autosomes than in the X chromosome. In addition, in CB4858, ka/ks values differ between chromosomes. CONCLUSIONS: The clear bias of deletions over insertions in CB4856 suggests that either the CB4856 genome is becoming smaller or the N2 genome is getting larger. We hypothesize the hotspots found represent alleles that are shared between CB4856 and CB4858 but not N2. Because the ka/ks ratio in the X chromosome is higher than the autosomes on average in CB4858, purifying selection is reduced on the X chromosome.

Ju J et al. (2023) Nucleic Acids Res "Structure of the Caenorhabditis elegans m6A methyltransferase METT10 that regulates SAM homeostasis."

Tomita K, Ju J, Kuroyanagi H, Aoyama T, Yashiro Y, Yamashita S

[Nucleic Acids Res, 2023]

In Caenorhabditis elegans, the N6-methyladenosine (m6A) modification by METT10, at the 3'-splice sites in S-adenosyl-l-methionine (SAM) synthetase (sams) precursor mRNA (pre-mRNA), inhibits sams pre-mRNA splicing, promotes alternative splicing coupled with nonsense-mediated decay of the pre-mRNAs, and thereby maintains the cellular SAM level. Here, we present structural and functional analyses of C. elegans METT10. The structure of the N-terminal methyltransferase domain of METT10 is homologous to that of human METTL16, which installs the m6A modification in the 3'-UTR hairpins of methionine adenosyltransferase (MAT2A) pre-mRNA and regulates the MAT2A pre-mRNA splicing/stability and SAM homeostasis. Our biochemical analysis suggested that C. elegans METT10 recognizes the specific structural features of RNA surrounding the 3'-splice sites of sams pre-mRNAs, and shares a similar substrate RNA recognition mechanism with human METTL16. C. elegans METT10 also possesses a previously unrecognized functional C-terminal RNA-binding domain, kinase associated 1 (KA-1), which corresponds to the vertebrate-conserved region (VCR) of human METTL16. As in human METTL16, the KA-1 domain of C. elegans METT10 facilitates the m6A modification of the 3'-splice sites of sams pre-mRNAs. These results suggest the well-conserved mechanisms for the m6A modification of substrate RNAs between Homo sapiens and C. elegans, despite their different regulation mechanisms for SAM homeostasis.

Kimura A et al. (2000) J Biol Chem "Both PCE-1/RX and OTX/CRX interactions are necessary for photoreceptor-specific gene expression."

Wawrousek EF, Kimura A, Nakamura M, Singh D, Shinohara T, Kikuchi M

[J Biol Chem, 2000]

RX, a homeodomain-containing protein essential for proper eye development (Mathers, P. H. Grinberg, A., Mahon, K. A., and Jamrich, M. (1997) Nature 387, 603-607), binds to the photoreceptor conserved element-1 (PCE-1/Ret 1) in the photoreceptor cell-specific arrestin promoter and stimulates gene expression. RX is found in many retinal cell types including photoreceptor cells. Another homeodomain-containing protein, CRX, which binds to the OTX element to stimulate promoter activity, is found exclusively in photoreceptor cells (Chen, S., Wang, Q. L., Nie, Z., Sun, H., Lennon, G., Copeland, N. G., Gillbert, D. J. Jenkins, N. A., and Zack, D. J. (1997) Neuron 19, 1017-1030; Furukawa, T., Morrow, E. M., and Cepko, C. L. (1997) Cell 91, 531-541). Binding assay and cell culture studies indicate that both PCE-1 and OTX elements and at least two different regulatory factors RX and CRX are necessary for high level, photoreceptor cell-restricted gene expression. Thus, photoreceptor specificity can be achieved by multiple promoter elements interacting with a combination of both photoreceptor-specific regulatory factors and factors present in closely related cell lineages.

Rose JK et al. (2022) Front Behav Neurosci "Neuroligin Plays a Role in Ethanol-Induced Disruption of Memory and Corresponding Modulation of Glutamate Receptor Expression."

Liang J, Lin CHS, Rose JK, Parvand M, Butterfield M, Rankin CH

[Front Behav Neurosci, 2022]

Exposure to alcohol causes deficits in long-term memory formation across species. Using a long-term habituation memory assay in Caenorhabditis elegans, the effects of ethanol on long-term memory (> 24 h) for habituation were investigated. An impairment in long-term memory was observed when animals were trained in the presence of ethanol. Cues of internal state or training context during testing did not restore memory. Ethanol exposure during training also interfered with the downregulation of AMPA/KA-type glutamate receptor subunit (GLR-1) punctal expression previously associated with long-term memory for habituation in C. elegans. Interestingly, ethanol exposure alone had the opposite effect, increasing GLR-1::GFP punctal expression. Worms with a mutation in the C. elegans ortholog of vertebrate neuroligins (nlg-1) were resistant to the effects of ethanol on memory, as they displayed both GLR-1::GFP downregulation and long-term memory for habituation after training in the presence of ethanol. These findings provide insights into the molecular mechanisms through which alcohol consumption impacts memory.

Russell GJ et al. (1989) Biochim Biophys Acta "Colchicine binding in the free-living nematode Caenorhabditis elegans."

Lacey E, Russell GJ

[Biochim Biophys Acta, 1989]

The [3H]colchicine-binding activity of a crude supernatant of the free-living nematode Caenorhabditis elegans was resolved into a non-saturable component and a tubulin-specific component after partial purification of tubulin by polylysine affinity chromatography. The two fractions displayed opposing thermal dependencies of [3H]colchicine binding, with non-saturable binding increasing, and tubulin binding decreasing, at 4 degrees C. Binding of [3H]colchicine to C.elegans tubulin at 37 degrees C is a pseudo-first-order rate process with a long equilibration time. The affinity of C. elegans tubulin for [3H]colchicine is relatively low (Ka = 1.7 x 10(5) M(-1)) and is characteristic of the colchicine binding affinities observed for tubulins derived from parasitic nematodes. [3H]Colchicine binding to C. elegans tubulin was inhibited by unlabelled colchicine, podophyllotoxin and mebendazole, and was enhanced by vinblastine. The inhibition of [3H]colchicine binding by mebendazole was 10-fold greater for C. elegans tubulin than for ovine brain tubulin. The inhibition of [3H]colchicine binding to C. elegans tubulin by mebendazole is consistent with the recognised anthelmintic action of the benzimidazole carbamates. These data indicate that C. elegans is a useful model for examining the interactions between microtubule inhibitors and the colchicine binding site of nematode tubulin.

Miyazaki H et al. (2012) Biophys J "CLC anion channel regulatory phosphorylation and conserved signal transduction domains."

Parton A, Kim S, Yamada T, Miyazaki H, Beth AH, Morrison R, Strange K

[Biophys J, 2012]

The signaling mechanisms that regulate CLC anion channels are poorly understood. Caenorhabditis elegans CLH-3b is a member of the CLC-1/2/Ka/Kb channel subfamily. CLH-3b is activated by meiotic cell-cycle progression and cell swelling. Inhibition is brought about by GCK-3 kinase-mediated phosphorylation of S742 and S747 located on a 176 amino acid disordered domain linking CBS1 and CBS2. Much of the inter-CBS linker is dispensable for channel regulation. However, deletion of a 14 amino acid activation domain encompassing S742 and S747 inhibits channel activity to the same extent as GCK-3. The crystal structure of CmCLC demonstrated that CBS2 interfaces extensively with an intracellular loop connecting membrane helices H and I, the C-terminus of helix D, and a short linker connecting helix R to CBS1. Point mutagenesis of this interface identified two highly conserved aromatic amino acid residues located in the H-I loop and the first -helix (1) of CBS2. Mutation of either residue to alanine rendered CLH-3b insensitive to GCK-3 inhibition. We suggest that the dephosphorylated activation domain normally interacts with CBS1 and/or CBS2, and that conformational information associated with this interaction is transduced through a conserved signal transduction module comprising the H-I loop and CBS2 1.

Jentsch TJ et al. (2018) Physiol Rev "CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease."

Jentsch TJ, Pusch M

[Physiol Rev, 2018]

CLC anion transporters are found in all phyla and form a gene family of eight members in mammals. Two CLC proteins, each of which completely contains an ion translocation parthway, assemble to homo- or heteromeric dimers that sometimes require accessory -subunits for function. CLC proteins come in two flavors: anion channels and anion/proton exchangers. Structures of these two CLC protein classes are surprisingly similar. Extensive structure-function analysis identified residues involved in ion permeation, anion-proton coupling and gating and led to attractive biophysical models. In mammals, ClC-1, -2, -Ka/-Kb are plasma membrane Cl^- channels, whereas ClC-3 through ClC-7 are 2Cl^-/H⁺-exchangers in endolysosomal membranes. Biological roles of CLCs were mostly studied in mammals, but also in plants and model organisms like yeast and Caenorhabditis elegans. CLC Cl^- channels have roles in the control of electrical excitability, extra- and intracellular ion homeostasis, and transepithelial transport, whereas anion/proton exchangers influence vesicular ion composition and impinge on endocytosis and lysosomal function. The surprisingly diverse roles of CLCs are highlighted by human and mouse disorders elicited by mutations in their genes. These pathologies include neurodegeneration, leukodystrophy, mental retardation, deafness, blindness, myotonia, hyperaldosteronism, renal salt loss, proteinuria, kidney stones, male infertility, and osteopetrosis. In this review, emphasis is laid on biophysical structure-function analysis and on the cell biological and organismal roles of mammalian CLCs and their role in disease.

Nathalie Strutz et al. (2001) International C. elegans Meeting "Identification of functional ion channel domains of C. elegans glutamate receptor subunits"

Nathalie Strutz, David Madsen, Michael Hollmann, Andres V Maricq

[International C. elegans Meeting, 2001]

Glutamate receptors are not only abundant in the nervous system of vertebrates but also of invertebrates. In Caenorhabditis elegans , 10 putative ionotropic glutamate receptor subunits have been identified so far (Brockie et al. 2001, J. Neurosci. 21, 1510). Two of these, GLR-1 and GLR-2, have been expressed in Xenopus oocytes and failed to show ligand-activated currents. Therefore, we set out to investigate if the pore-forming domains of these two subunits are intrinsically functional ion conducting domains. We have previously shown that domain transplantation between different subtypes of glutamate receptors can generate functional chimeras that allow the characterization of those domains. To test for pore function, we transplanted the pore-forming region of GLR-1 or GLR-2 into either rat GluR1, as a representative of the AMPA receptor family, or rat GluR6, as a representative of the KA receptor family. The resulting chimeras GluR1-PGLR1, GluR1-PGLR2, GluR6-PGLR1, and GluR6-PGLR2, as well as the respective wildtypes GluR1, GluR6, GLR-1, and GLR-2 were expressed and characterized in Xenopus oocytes. Surprisingly, GluR1-PGLR1, GluR1-PGLR2, and GluR6-PGLR2 produced ligand-activated currents, despite the GLR-1 and GLR-2 wildtype receptor's apparent lack of ion channel function. The maximal current amplitudes of GluR6-PGLR2 were comparable to wildtype GluR6. GluR1-PGLR1 and GluR1-PGLR2 showed reduced current amplitudes in comparison to wildtype GluR1. The ion channel properties of the chimeras classify GLR-1 and GLR-2 into the group of non-NMDA receptor subunits, consistent with conclusions derived from sequence homology data.

Nehrke K et al. (2002) J Biol Chem "The NHX family of Na+-H+ exchangers in Caenorhabditis elegans."

Nehrke K, Melvin JE

[J Biol Chem, 2002]

Na-H exchangers prevent cellular acidification by catalyzing the electroneutral exchange of extracellular sodium for an intracellular proton. To date, seven Na-H exchangers have been identified in mammals, and although several members of this family have been extensively studied and characterized, it is clear that there are major gaps in our understanding with respect to the remaining family members. In order to initiate the study of Na-H exchangers in a genomically-defined and genetically-tractable model system, we have cloned the complete cDNAs and analyzed splice site variation for nine putative homologs from the nematode C. elegans, which we have called nhx-1 through -9. The expression patterns and cellular distributions of the nhx proteins were determined using transcriptional and translational promoter-transgene fusion constructs to Green Fluorescent Protein (GFP). Four of the putative exchangers were expressed at the cell surface, while five of the exchangers were associated with the membranes of intracellular organelles. Individual isoforms were expressed exclusively in the intestine, seam cells, hypodermal cells of the main body syncytium, and the excretory cell, all of which are polarized epithelial cells, suggesting a role for these proteins in epithelial membrane transport processes in the nematode. Other isoforms were found to express either ubiquitously, or in a pan-neural pattern, suggesting a more conserved role in cell pH regulation or neuronal function. Finally, we show that recombinant nhx-4, the ubiquitous nematode Na-H exchanger, mediates Na+-dependent pH recovery following intracellular acidification. Nhx-4 activity has a Ka for Na+ of approximately 32 mM, is not Cl--dependent, and is relatively insensitive to the amiloride analog EIPA.