scholarly journals Caenorhabditis elegans lin-13, a Member of the LIN-35 Rb Class of Genes Involved in Vulval Development, Encodes a Protein With Zinc Fingers and an LXCXE Motif

Genetics ◽  
2000 ◽  
Vol 155 (3) ◽  
pp. 1127-1137 ◽  
Author(s):  
Alicia Meléndez ◽  
Iva Greenwald
Keyword(s):  
Caenorhabditis Elegans ◽  
Zinc Fingers ◽  
Null Alleles ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Vulval Development ◽  
Vulval Precursor Cells ◽  
Class B ◽  
Genes Encoding ◽  
Lxcxe Motif

Abstract The SynMuv genes appear to be involved in providing a signal that inhibits vulval precursor cells from adopting vulval fates in Caenorhabditis elegans. One group of SynMuv genes, termed class B, includes genes encoding proteins related to the tumor suppressor Rb and RbAp48, a protein that binds Rb. Here, we provide genetic evidence that lin-13 behaves as a class B SynMuv gene. We show that null alleles of lin-13 are temperature sensitive and maternally rescued, resulting in phenotypes ranging in severity from L2 arrest (when both maternal and zygotic activities are removed at 25°), to sterile Multivulva (when only zygotic activity is removed at 25°), to sterile non-Multivulva (when both maternal and zygotic activities are removed at 15°), to wild-type/class B SynMuv (when only zygotic activity is removed at 15°). We also show that LIN-13 is a nuclear protein that contains multiple zinc fingers and a motif, LXCXE, that has been implicated in Rb binding. These results together suggest a role for LIN-13 in Rb-mediated repression of vulval fates.

scholarly journals The multivulva phenotype of certain Caenorhabditis elegans mutants results from defects in two functionally redundant pathways.

Genetics ◽  
1989 ◽  
Vol 123 (1) ◽  
pp. 109-121 ◽  
Author(s):  
E L Ferguson ◽  
H R Horvitz
Keyword(s):  
Caenorhabditis Elegans ◽  
Cell Lineage ◽  
Gene Families ◽  
Precursor Cells ◽  
Wild Type ◽  
Vulval Development ◽  
Vulval Precursor Cells ◽  
New Mutations ◽  
Intercellular Signal

Abstract We previously identified Caenorhabditis elegans mutants in which certain of the six vulval precursor cells adopt fates normally expressed by other vulval precursor cells. These mutants define genes that appear to function in the response to an intercellular signal that induces vulval development. The multivulva (Muv) phenotype of one such mutant, CB1322, results from an interaction between two unlinked mutations, lin-8(n111) II and lin-9(n112) III. In this paper, we identify 18 new mutations, which are alleles of eight genes, that interact with either lin-8(n111) or lin-9(n112) to generate a Muv phenotype. None of these 20 mutations alone causes any vulval cell lineage defects. The "silent Muv" mutations fall into two classes; hermaphrodites carrying a mutation of each class are Muv, while hermaphrodites carrying two mutations of the same class have a wild-type vulval phenotype. Our results indicate that the Muv phenotype of these mutants results from defects in two functionally-redundant pathways, thereby demonstrating that redundancy can occur at the level of gene pathways as well as at the level of gene families.

Reversal of cell fate determination in Caenorhabditis elegans vulval development

Development ◽  
1996 ◽  
Vol 122 (8) ◽  
pp. 2507-2515 ◽  
Author(s):  
S. Euling ◽  
V. Ambros
Keyword(s):  
Caenorhabditis Elegans ◽  
Cell Fate ◽  
Larval Stage ◽  
Cell Types ◽  
Wild Type ◽  
Vulval Development ◽  
Fate Determination ◽  
Developmental Arrest ◽  
Dauer Larva ◽  
Vulval Precursor Cells

In Caenorhabditis elegans, the fates of the multipotent vulval precursor cells (VPCs) are specified by intercellular signals. The VPCs divide in the third larval stage (L3) of the wild type, producing progeny of determined cell types. In lin-28 mutants, vulva development is similar to wild-type vulva development except that it occurs precociously, in the second larval stage (L2). Consequently, when lin-28 hermaphrodites temporarily arrest development at the end of L2 in the dauer larva stage, they have partially developed vulvae consisting of VPC progeny. During post-dauer development, these otherwise determined VPC progeny become reprogrammed back to the multipotent, signal-sensitive state of VPCs. Our results indicate that VPC fate determination by intercellular signals is reversible by dauer larva developmental arrest and post-dauer development.

Genetic analysis of small nuclear RNAs in Saccharomyces cerevisiae: viable sextuple mutant

1988 ◽  
Vol 8 (8) ◽  
pp. 3150-3159
Author(s):  
R Parker ◽  
T Simmons ◽  
E O Shuster ◽  
P G Siliciano ◽  
C Guthrie
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcriptional Start Site ◽  
Single Copy ◽  
Gene Replacement ◽  
Null Alleles ◽  
Wild Type ◽  
Small Nuclear Rnas ◽  
Apparent Correlation ◽  
Genes Encoding ◽  
Sm Antigen

Saccharomyces cerevisiae contains at least 24 distinct small nuclear RNAs (snRNAs), several of which are known to be essential for viability and to participate in the splicing of pre-mRNAs; the RNAs in this subset contain binding sites for the Sm antigen, a hallmark of metazoan snRNAs involved in mRNA processing. In contrast, we showed previously that the single-copy genes for three other snRNAs (snR3, snR4, and snR10) are not required for viability, although cells lacking snR10 are growth impaired at low temperature. None of these RNAs associates with the Sm antigen. To assess this apparent correlation, we cloned and sequenced the genes encoding three additional non-Sm snRNAs. Comparison of these genes with nine additional yeast snRNA genes revealed a highly conserved TATA box located 92 +/- 8 nucleotides 5' of the transcriptional start site. By using the technique of gene replacement with null alleles, each of these three single copy genes was shown to be completely dispensable. We constructed multiple mutants to test the hypothesis that, individually, each of these snRNAs is nonessential because the snRNAs play functionally overlapping roles. A mutant lacking five snRNAs (snR3, snR4, snR5, snR8, snR9) was indistinguishable from the wild type, and growth of the sextuple mutant was no more impaired than that in strains lacking only snR10. This widespread dispensability of snRNAs was completely unexpected and forces us to reconsider the possible roles of these ubiquitous RNAs.

The terminal differentiation factor LIN-29 is required for proper vulval morphogenesis and egg laying in Caenorhabditis elegans

Development ◽  
1997 ◽  
Vol 124 (21) ◽  
pp. 4333-4342 ◽  
Author(s):  
J.C. Bettinger ◽  
S. Euling ◽  
A.E. Rougvie
Keyword(s):  
Caenorhabditis Elegans ◽  
Terminal Differentiation ◽  
Egg Laying ◽  
Small Subset ◽  
Wild Type ◽  
Vulval Development ◽  
Genetic Mosaics ◽  
Hypodermal Cell ◽  
Heterochronic Gene ◽  
Final Molt

Caenorhabditis elegans vulval development culminates during exit from the L4-to-adult molt with the formation of an opening through the adult hypodermis and cuticle that is used for egg laying and mating. Vulva formation requires the heterochronic gene lin-29, which triggers hypodermal cell terminal differentiation during the final molt. lin-29 mutants are unable to lay eggs or mate because no vulval opening forms; instead, a protrusion forms at the site of the vulva. We demonstrate through analysis of genetic mosaics that lin-29 is absolutely required in a small subset of lateral hypodermal seam cells, adjacent to the vulva, for wild-type vulva formation and egg laying. However, lin-29 function is not strictly limited to the lateral hypodermis. First, LIN-29 accumulates in many non-hypodermal cells with known roles in vulva formation or egg laying. Second, animals homozygous for one lin-29 allele, ga94, have the vulval defect and cannot lay eggs, despite having a terminally differentiated adult lateral hypodermis. Finally, vulval morphogenesis and egg laying requires lin-29 activity within the EMS lineage, a lineage that does not generate hypodermal cells.

scholarly journals FLR-4, a Novel Serine/Threonine Protein Kinase, Regulates Defecation Rhythm in Caenorhabditis elegans

2005 ◽  
Vol 16 (3) ◽  
pp. 1355-1365 ◽  
Author(s):  
Masaya Take-uchi ◽  
Yuri Kobayashi ◽  
Koutarou D. Kimura ◽  
Takeshi Ishihara ◽  
Isao Katsura
Keyword(s):  
Caenorhabditis Elegans ◽  
Protein Kinase ◽  
Temperature Shift ◽  
Kinase Domain ◽  
Temperature Sensitive ◽  
Missense Mutations ◽  
Wild Type ◽  
Hydrophobic Region ◽  
Pharyngeal Muscles ◽  
Unique Structural Feature

The defecation behavior of the nematode Caenorhabditis elegans is controlled by a 45-s ultradian rhythm. An essential component of the clock that regulates the rhythm is the inositol trisphosphate receptor in the intestine, but other components remain to be discovered. Here, we show that the flr-4 gene, whose mutants exhibit very short defecation cycle periods, encodes a novel serine/threonine protein kinase with a carboxyl terminal hydrophobic region. The expression of functional flr-4::GFP was detected in the intestine, part of pharyngeal muscles and a pair of neurons, but expression of flr-4 in the intestine was sufficient for the wild-type phenotype. Furthermore, laser killing of the flr-4–expressing neurons did not change the defecation phenotypes of wild-type and flr-4 mutant animals. Temperature-shift experiments with a temperature-sensitive flr-4 mutant suggested that FLR-4 acts in a cell-functional rather than developmental aspect in the regulation of defecation rhythms. The function of FLR-4 was impaired by missense mutations in the kinase domain and near the hydrophobic region, where the latter allele seemed to be a weak antimorph. Thus, a novel protein kinase with a unique structural feature acts in the intestine to increase the length of defecation cycle periods.

scholarly journals Mitochondrial Dysfunction Confers Resistance to Multiple Drugs in Caenorhabditis elegans

2010 ◽  
Vol 21 (6) ◽  
pp. 956-969 ◽  
Author(s):  
Iryna O. Zubovych ◽  
Sarah Straud ◽  
Michael G. Roth
Keyword(s):  
Drug Resistance ◽  
Caenorhabditis Elegans ◽  
Mitochondrial Protein ◽  
Drug Resistant ◽  
Wild Type ◽  
C Elegans ◽  
Mitochondrial Inhibitors ◽  
Mitochondrial Superoxide ◽  
Genes Encoding ◽  
Multiple Drugs

In a previous genetic screen for Caenorhabditis elegans mutants that survive in the presence of an antimitotic drug, hemiasterlin, we identified eight strong mutants. Two of these were found to be resistant to multiple toxins, and in one of these we identified a missense mutation in phb-2, which encodes the mitochondrial protein prohibitin 2. Here we identify two additional mutations that confer drug resistance, spg-7 and har-1, also in genes encoding mitochondrial proteins. Other mitochondrial mutants, isp-1, eat-3, and clk-1, were also found to be drug-resistant. Respiratory complex inhibitors, FCCP and oligomycin, and a producer of reactive oxygen species (ROS), paraquat, all rescued wild-type worms from hemiasterlin toxicity. Worms lacking mitochondrial superoxide dismutase (MnSOD) were modestly drug-resistant, and elimination of MnSOD in the phb-2, har-1, and spg-7 mutants enhanced resistance. The antioxidant N-acetyl-l-cysteine prevented mitochondrial inhibitors from rescuing wild-type worms from hemiasterlin and sensitized mutants to the toxin, suggesting that a mechanism sensitive to ROS is necessary to trigger drug resistance in C. elegans. Using genetics, we show that this drug resistance requires pkc-1, the C. elegans ortholog of human PKCε.

scholarly journals An efficient genome editing strategy to generate putative null mutants in Caenorhabditis elegans using CRISPR/Cas9

2018 ◽  
Author(s):  
Han Wang ◽  
Heenam Park ◽  
Jonathan Liu ◽  
Paul W. Sternberg
Keyword(s):  
Dna Repair ◽  
Caenorhabditis Elegans ◽  
Genome Editing ◽  
Gene Function ◽  
Unique Sequence ◽  
Target Site ◽  
Null Alleles ◽  
Wild Type ◽  
C Elegans ◽  
Null Mutants

AbstractNull mutants are essential for analyzing gene function. Here, we describe a simple and efficient method to generate Caenorhabditis elegans null mutants using CRISPR/Cas9 and short single stranded DNA oligo repair templates to insert a universal 43-nucleotide-long stop knock-in (STOP-IN) cassette into the early exons of target genes. This cassette has stop codons in all three reading frames and leads to frameshifts, which will generate putative null mutations regardless of the reading frame of the insertion position in exons. The STOP-IN cassette also contains an exogenous Cas9 target site that allows further genome editing and provides a unique sequence that simplifies the identification of successful insertion events via PCR. As a proof of concept, we inserted the STOP-IN cassette right at a Cas9 target site in aex-2 to generate new putative null alleles by injecting preassembled Cas9 ribonucleoprotein and a short synthetic single stranded DNA repair template containing the STOP-IN cassette and two 35-nucleotide-long homology arms identical to the sequences flanking the Cas9 cut site. We showed that these new aex-2 alleles phenocopied an existing loss-of-function allele of aex-2. We further showed that the new aex-2 null alleles could be reverted back to the wild-type sequence by targeting exogenous Cas9 cut site included in the STOP-IN cassette and providing a single stranded wild-type DNA repair oligo. We applied our STOP-IN method to generate new putative null mutants for additional 20 genes, including three pharyngeal muscle-specific genes (clik-1, clik-2, and clik-3), and reported a high insertion rate (46%) based on the animals we screened. We showed that null mutations of clik-2 cause recessive lethality with a severe pumping defect and clik-3 null mutants have a mild pumping defect, while clik-1 is dispensable for pumping. We expect that the knock-in method using the STOP-IN cassette will facilitate the generation of new null mutants to understand gene function in C. elegans and other genetic model organisms.SummaryWe report a simple and efficient CRISPR/Cas9 genome editing strategy to generate putative null C. elegans mutants by inserting a small universal stop knock-in (STOP-IN) cassette with stop codons in three frames and frameshifts. The strategy is cloning-free, with the mixture consisting of preassembled Cas9 ribonucleoprotein and single stranded repair DNA oligos directly injected into gonads of adult C. elegans. The universal STOP-IN cassette also contains a unique sequence that simplifies detection of successful knock-in events via PCR and an exogenous Cas9 target sequence that allows further genome editing.

scholarly journals Rules of Nonallelic Noncomplementation at the Synapse in Caenorhabditis elegans

Genetics ◽  
2001 ◽  
Vol 158 (1) ◽  
pp. 209-220 ◽  
Author(s):  
Karen J Yook ◽  
Stephen R Proulx ◽  
Erik M Jorgensen
Keyword(s):  
Caenorhabditis Elegans ◽  
Gene Product ◽  
Genetic Interaction ◽  
Synaptic Proteins ◽  
Null Alleles ◽  
Nematode Caenorhabditis Elegans ◽  
Recessive Mutations ◽  
Genes Encoding ◽  
Physical Interactions ◽  
Encoding Gene

Abstract Nonallelic noncomplementation occurs when recessive mutations in two different loci fail to complement one another, in other words, the double heterozygote exhibits a phenotype. We observed that mutations in the genes encoding the physically interacting synaptic proteins UNC-13 and syntaxin/UNC-64 failed to complement one another in the nematode Caenorhabditis elegans. Noncomplementation was not observed between null alleles of these genes and thus this genetic interaction does not occur with a simple decrease in dosage at the two loci. However, noncomplementation was observed if at least one gene encoded a partially functional gene product. Thus, this genetic interaction requires a poisonous gene product to sensitize the genetic background. Nonallelic noncomplementation was not limited to interacting proteins: Although the strongest effects were observed between loci encoding gene products that bind to one another, interactions were also observed between proteins that do not directly interact but are members of the same complex. We also observed noncomplementation between genes that function at distant points in the same pathway, implying that physical interactions are not required for nonallelic noncomplementation. Finally, we observed that mutations in genes that function in different processes such as neurotransmitter synthesis or synaptic development complement one another. Thus, this genetic interaction is specific for genes acting in the same pathway, that is, for genes acting in synaptic vesicle trafficking.

scholarly journals fog-1, a regulatory gene required for specification of spermatogenesis in the germ line of Caenorhabditis elegans.

Genetics ◽  
1990 ◽  
Vol 125 (1) ◽  
pp. 29-39 ◽  
Author(s):  
M K Barton ◽  
J Kimble
Keyword(s):  
Caenorhabditis Elegans ◽  
Germ Cell ◽  
Germ Line ◽  
Regulatory Gene ◽  
Sensitive Period ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Male Germ Line ◽  
Somatic Tissues ◽  
Lines Of Evidence

Abstract In wild-type Caenorhabditis elegans, the XO male germ line makes only sperm and the XX hermaphrodite germ line makes sperm and then oocytes. In contrast, the germ line of either a male or a hermaphrodite carrying a mutation of the fog-1 (feminization of the germ line) locus is sexually transformed: cells that would normally make sperm differentiate as oocytes. However, the somatic tissues of fog-1 mutants remain unaffected. All fog-1 alleles identified confer the same phenotype. The fog-1 mutations appear to reduce fog-1 function, indicating that the wild-type fog-1 product is required for specification of a germ cell as a spermatocyte. Two lines of evidence indicate that a germ cell is determined for sex at about the same time that it enters meiosis. These include the fog-1 temperature sensitive period, which coincides in each sex with first entry into meiosis, and the phenotype of a fog-1; glp-1 double mutant. Experiments with double mutants show that fog-1 is epistatic to mutations in all other sex-determining genes tested. These results lead to the conclusion that fog-1 acts at the same level as the fem genes at the end of the sex determination pathway to specify germ cells as sperm.

GENETIC STUDIES OF UNUSUAL LOCI THAT AFFECT BODY SHAPE OF THE NEMATODE CAENORHABDITIS ELEGANS AND MAY CODE FOR CUTICLE STRUCTURAL PROTEINS

Genetics ◽  
1986 ◽  
Vol 113 (3) ◽  
pp. 621-639
Author(s):  
Meredith Kusch ◽  
R S Edgar
Keyword(s):  
Caenorhabditis Elegans ◽  
Body Shape ◽  
Structural Proteins ◽  
Null Alleles ◽  
Wild Type ◽  
Nematode Caenorhabditis Elegans ◽  
Genetic Studies ◽  
Wild Type Phenotype ◽  
Mutant Phenotypes ◽  
Collagen Genes

ABSTRACT In Caenorhabditis elegans, four loci (sqt-1, sqt-2, sqt-3 and rol-8) in which mutations affect body shape and cuticle morphology have unusual genetic properties. (1) Mutant alleles of sqt-1 can interact to produce animals with a variety of mutant phenotypes: left roller, right roller, dumpy and long. At least three mutant phenotypes are specified by mutations in the sqt-3 locus. (2) Most alleles at these loci are either dominant or cryptic dominant (i.e., are dominant only in certain genetic backgrounds). (3) Most alleles of these loci exhibit codominance. (4) Two putative null alleles of the sqt-1 locus produce a wild-type phenotype. (5) Many alleles of these genes demonstrate unusual intergenic interactions that are not the result of simple epistasis: animals doubly heterozygous for mutations at two loci often display unexpected and unpredictable phenotypes. We suggest that these genetic properties might be expected of genes, such as the collagen genes, the products of which interact to form the animal's cuticle, and which are member genes of a gene family.

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