Kraemer BC et al. (2006) Hum Mol Genet "Molecular pathways that influence human tau-induced pathology in Caenorhabditis elegans."

Kraemer BC, Burgess JK, Chen JH, Thomas JH, Schellenberg GD

[Hum Mol Genet, 2006]

Mutations in the gene encoding tau cause frontotemporal dementia with parkinsonism - chromosome 17 type (FTDP-17). In FTDP-17, Alzheimer''s disease (AD), and other tauopathies, aggregated hyper-phosphorylated tau forms the neurofibrillary tangles characteristic of these disorders. We previously reported a C. elegans model for tauopathies using human normal and FTDP-17 mutant tau as transgenes. Neuronal transgene expression caused insoluble phosphorylated tau accumulation, neurodegeneration, and uncoordinated (Unc) movement. Here we describe a genome-wide RNA mediated interference (RNAi) screen for genes that modify the tau induced Unc phenotype. We tested RNAi sequences for 16,757 genes and found 75 that enhanced the transgene induced Unc phenotype. Forty-six of these genes have sequence similarity to known human genes and fall into a number of broad classes including kinases, chaperones, proteases, and phosphatases. The remaining 29 modifiers have sequence similarity only with other nematode genes. To determine if the enhancers are specific for the tau-induced Unc behavior, we exposed several non-tau Unc mutants to tau RNAi enhancer clones. Fifteen enhancers modified phenotypes in multiple Unc mutants, while 60 modified only the Unc phenotype in the tau transgenic lines. We also introduced the tau transgene into the background of genetic loss of function mutations for a subset of the enhancer genes. Tau transgenic animals homozygous for loss of these enhancer genes exhibited increased impaired motility relative to the tau transgene line alone. This work uncovers novel candidate genes that prevent tau toxicity, as well as genes previously implicated in tau-mediated neurodegeneration.

Fields CA et al. (1992) Worm Breeder's Gazette "DEAD and DEAH Box RNA Helicase Genes Identified by EST Sequencing"

Fields CA, McCombie WR

[Worm Breeder's Gazette, 1992]

The DEAD-box family of proteins includes RNA-dependent ATPases involved in processing both structural and messenger RNAs (Linder et al., Nature 337 (1989) 121-122). A number of DEAD-box proteins have been identified as translational initiation factors, including the mouse eIF-4 A1protein (Nielsen et al., NAR 13 (1985) 6867-6880)) and the recently-characterized ME31 Bprotein of Drosophila (de Valoir et al., Proc. Natl. Acad. Sci. USA 88 (1991) 2113-2117). The predicted translation product of EST D18 R(McCombie et al., this issue) is very similar to the C-terminal ends of these proteins, as well as other members of the DEAD-box family, suggesting that D18 Rtags an initiation-factor gene. EST D33 Falso matches these proteins. The DEAH-box family includes putative RNA helicases involved in spliceosome assembly, and hence presumably in splice-site selection (Burgess et al., Cell 60 (1990) 705-717). The maleless gene of Drosophila, which regulates dosage compensation in males by binding multiple sites on the X chromosome, is also a DEAH-box protein (Kuroda et al., Cell 66 (1991) 935-947). Two ESTs with high similarity to yeast PRP16 (Burgess et al.), maleless, and other members of the DEAH-box family have been sequenced. EST H51 Rtags the DEAH-box region of the protein, the putative RNA helicase domain, which is highly conserved between members of the family. EST H57 Rtags a region similar to the C-terminal ends of PRP2 (Chen, J. and Lin. R. Nucl. Acids Res. 18 (1990) 6447) and PRP16 ;the maleless protein is significantly less similar to the PRP proteins in this region than is the C. elegans protein. We do not yet know whether H51 Rand H57 Rtag the same gene. [See Figure 1]

Tanner K et al. (1988) Worm Breeder's Gazette "further analysis of ct31 X, a locus affecting sex determination and dosage compensation."

Tanner K, Wood WB, Burgess SM, Trent C

[Worm Breeder's Gazette, 1988]

We described earlier the isolation of ct31 as a gamma-ray-induced, semi-dominant, feminizing mutation that suppresses the masculinization of XX animals caused by the her-1(gf) allele n695, and reported that ct31 increases the penetrance of the X-linked hypomorph lin-15(n765), suggesting that ct31 might cause a decrease in X-chromosome expression (Burgess et al., WBG May, 1986). ct31 XX animals are Dpy, Egl hermaphrodites; XO animals are Dpy, nonfertile, partially feminized males. Here we present evidence that ct31 lowers expression of the X- linked act-4 gene and that it is probably a gain-of-function mutation involving duplication of a region of the X chromosome. Assay of act-4 mRNA in ct31 and N2 L1 hermaphrodites using an RNA dot blot assay ( Donahue et al., PNAS 84:7600,1987) showed that the level of this transcript in ct31 animals is only 46% + 4% of the level in N2 animals. This finding supports the genetic evidence cited above that ct31 causes a decrease in expression of X-linked genes. Three-factor mapping data places ct31 in the lin-2 - unc-9 interval on X. Deficiencies of this region have not been obtained, but it is covered by the duplication mnDp10. XX hermaphrodites of genotype mnDp10/+; ct31 lin15, which have two mutant and one wild-type copy of the ct31 locus, are more severely Dpy and Egl and generally sicker and more slow-growing than ct31 homozygotes. When the former animals are allowed to self-fertilize, one quarter of the progeny arrest as L1 larvae. We surmise that these arrested larvae are the class carrying two copies of mnDp10 (ct31/ct31/+/+). Since the ct31 phenotype appears to be enhanced by additional copies of the wildtype locus, these results are consistent with ct31 being a gain-of-function mutation. However, given the large size of mnDp10, and the fact that it carries at least one other locus known to affect X expression (sdc- 1), more definitive evidence on this point must await isolation and characterization of deficiencies in the region. The high reversion rate of ct31 described earlier (Burgess et al., 1986) suggested that it might involve an unstable chromosomal rearrangement. In an attempt to test this suggestion, we used a probe derived from the cloned myo-2 gene, which is located on the physical map at a position that should be near the ct31 locus, to carry out quantitative Southern blot analyses. The copy number of the myo-2 gene in the ct31 strain is clearly higher than in N2 or in the her-1(n695) parent strain of ct31. The elevated copy number of myo-2 appears identical to that measured in a strain homozygous for mnDp10. Seven independent nonDpy ct31 revertants do not show the elevated myo-2 copy number, correlating the visible ct31 phenotype and the apparent duplication. Preliminary experiments using as probes cosmid clones from the myo-2 contig ( obtained from A. Coulson and J. Sulston) indicate that at least 40 kb including and to the left of the myo-2 gene is duplicated in the ct31 mutant. Experiments are in progress to characterize further the extent and nature of the rearrangement.

Manser J et al. (1985) Worm Breeder's Gazette "Autosomal Extragenic Suppressors Of her-1(n695) V"

Wood WB, Manser J, Trent C

[Worm Breeder's Gazette, 1985]

We are studying five extragenic mutations that suppress the Tra phenotype of her-1(n695) V (see preceding abstract): ct27 III (gamma- ray induced), n1091 III and ct49 III (allelic, EMS-induced; ct49 was isolated by Sean Burgess), and n1092 V and n1102 V (allelic, EMS- induced). ct27 III has a semi-dominant dpy phenotype in XX animals; XO animals are less severely Dpy and are abnormal males. Both ct27; n695 and ct27/+;n695 XX animals exhibit strong suppression of the n695 Tra phenotype. Because ct27 results in a phenotype somewhat similar to that caused by mutations in the chauvinist dpy genes (dpy's 21, 26, 27, 28) it may suppress n695 with an effect on X-chromosome expression. n1091 III has no apparent effect on XX or XO animals by itself, while ct49 III XX and XO animals are Dpy h we have not eliminated the possibility that this phenotype results from a second closely linked mutation). n1091;n695 XX animals are well suppressed; n1091/+;n695 XX animals are not. n1092 V and n1102 V map near unc-76 (about 5% from her-1). Neither appears to have any effect on XX or XO animals by itself. For n1092 and n1102, both n695 sup and n695 sup/n695 + XX animals show strong suppression. n695n1O91 XO, n695n1O92 XO, and n695n11O2 XO animals are all normal males (that is, they are apparently identical to n695 XO animals). This suggests that the EMS-induced suppressor mutations on III and V affect a process that is active in XX, but not XO, animals. For example, the genes identified by these mutations could be involved in turning off her-1 expression in XX animals.

Orbidans, Helen E. et al. (2013) International Worm Meeting "Widespread pleiotropic Bateson-Dobzhansky-Muller incompatibilities between C. elegans isolates."

Harvey, Simon C., Snoek, L. Basten, Kammenga, Jan E., Stastna, Jana, Orbidans, Helen E.

[International Worm Meeting, 2013]

Bateson-Dobzhansky-Muller (BDM) incompatibilities are a result of deleterious interactions between alleles that are neutral or advantageous in their own genetic backgrounds. Both outbreeding depression and a specific incompatibility causing embryonic lethality have been identified within C. elegans. We therefore hypothesised that alleles producing BDM incompatibilities would also be present. Identifying the underlying loci producing such negative epistatic effects within a species is important as it will allow comparison to the loci and alleles that generate isolation between species, i.e. it addresses the role of BDM incompatibilities in driving speciation. To identify genomic regions showing BDM incompatibilities we undertook screens for regions that disrupted the normal process of egg-laying, a complex, highly regulated and coordinated phenotype. Screens were undertaken in recombinant inbred lines (RILs) and a genome-wide panel of nearly isogenic lines (NILs) both produced from the isolates CB4856 and N2. These RIL and NIL analyses identify a number of quantitative trait loci (QTLs) that show synthetic effects on egg-laying, i.e. the disruption in egg-laying is not seen in the parental isolates and is a consequence of negative interactions between CB4856 and N2 alleles. Analysis of these QTLs shows that they also affect other life history traits, affecting lifespan and the internal hatching of progeny (bagging). Further analysis indicates that this approach identifies only a subset of the incompatibilities between CB4856 and N2, that these incompatibilities are a consequence of complex interactions between multiple loci, and that they interact with the stress response. LBS and JK were funded by NWO-ALW NEMADAPT (project 855.01.151) and the Graduate School Production Ecology & Resource Conservation.

Bosenberg M et al. (1995) International C. elegans Meeting "LAG-1 Interacts with the GLP-1 ANK REPEATS."

Roehl H, Kimble JE, Bosenberg M

[International C. elegans Meeting, 1995]

glp-1 and lin-12 encode homologous transmembrane proteins that transduce a variety of signals required for proper development in C. elegans. To learn how signals are transduced by GLP-1, we have examined whether proteins implicated in GLP-1 signaling can interact with the cytoplasmic domain of GLP-1. One such candidate gene is lag-1, whose single mutant phenotype is similar to that of lin-12 glp-1 double mutants and its gene product is highly homologous to two DNA binding proteins: D. melanogaster Supressor of Hairless and human RBP-Jk (abstract by Christensen et al.). The possibility of a direct interaction between GLP-1 and LAG-1 was examined using the yeast two-hybrid system. Proteins that interact in this assay reconstitute the ability of two domains of Gal4 to activate transcription, which is monitored with a Gal4-dependent b-galactosidase reporter. A vector encoding a translational fusion between the Gal4 DNA binding domain and LAG-1 and one encoding a fusion of the Gal4 activation domain with the GLP-1 ankyrin repeats were prepared and cotransformed into a reporter strain. b-gal activity was induced greater than 30 fold relative to control cotransformations of either construct with unrelated proteins, indicating a specific interaction between LAG-1 and the GLP-1 ankyrin repeats. Additional experiments indicate that LAG-1 can also interact with itself and with D. melanogaster Hairless. We are testing whether in vitro translated LAG-1 and the GLP-1 ankyrin repeats interact and will be examining whether full-length GLP-1 interacts with LAG-1 in tissue culture cells.

Tanner K et al. (1990) Worm Breeder's Gazette "Large Duplications Including sdc-2 Cause Feminization and Reduced Viability of XO Animals"

Wood WB, Tanner K

[Worm Breeder's Gazette, 1990]

Three independently isolated X-linked suppressors of the dominant masculinizing her-1 allele n695 (Burgess et al., WBG 9.2, 1986) are large X-chromosome duplications (Tanner et al. C. elegans Meeting Abstracts 1989) that result in a high incidence of XO lethality with the surviving XO worms variably transformed toward hermaphrodite. The endpoints of these duplications have now been more precisely mapped by quantitative Southern blotting and each has been shown to include the sdc-2 locus. All three, ct28, ct30, and ct31 have left endpoints between xol-1 and lin-14. On the right ct30 terminates between myo-2 and unc-3, while ct28 and ct31 terminate between unc-3 and let-2. Thus ct30 can be estimated to duplicate between 14% and 51% of the X chromosome; ct31 and ct28 between 24% and 54%. ct31 has been shown genetically not to include Unc-15, and therefore does not include sdc- 1. All three duplications result in virtually indistinguishable phenotypes although they probably duplicate different amounts of the X chromosome. XO animals, carrying one copy of the duplication, are non- mating, slow-growing, and variably feminized. Nomarski observation of 140 ct31 XO and ct31 lon-2(e678) XO animals showed that 77% had defective tails, 18% had a mid-ventral hypodermal protrusion, 16% had a sac-like or a two-armed gonad, and 8% contained oocytes. Immunofluorescent staining of 193 ct31 XO animals with an anti- vitellogenin antibody indicated 11% (21 animals) produced yolk protein. Three ct31 XO animals have been observed to contain developing embryos. Homozygous ct28, ct30. and ct31 XX animals carrying two copies of the duplication are slow-growing Dumpy hermaphrodites. This phenotype is very different from that of animals carrying mnDp10, another large homozygous-viable X-chromosome duplication ( between 22% and 38% of the X chromosome), that includes much of the same region as ct28, ct30, and ct31 but does not include sdc-2. XO animals homozygous for mnDp10 are healthy mating males, although their fertility is reduced. XX animals homozygous for mnDp10 are perhaps very slightly dumpy but grow normally. These observations and the apparently different sizes of ct28, ct30, and ct31 suggest that their resulting phenotypes are not due simply to increased X-chromosome dosage, but rather to the duplication of specific sequences. In addition to feminizing XO animals, the ct31 duplication results in significant XO lethality. The ratio of XO to XX cross progeny counted in two separate crosses was 226/399, indicating 43% XO lethality. This lethality is XO specific since the self progeny of ct31 homozygous hermaphrodites show only 5% lethality. Similar experiments with ct28 and ct30 are in progress. We suggest that the feminization and reduced viability of XO animals containing these duplications is due primarily to increased copy number of sdc-2. Loss-of-function alleles of sdc-2 result in masculinization of XX animals, XX lethality and increased X-chromosome expression (Genetics 122; 579-593, 1989). If sdc-2 is a developmental switch gene, the gain-of-function phenotype should be hermaphroditization of XO animals, XO lethality and reduced X chromosome expression, as predicted by Nusbaum and Meyer (ibid.). Previous results from this lab indicate that the X-linked duplications affect X-chromosome expression, suggesting effects on dosage compensation as well as sexual phenotype. The ct31 duplication increases the penetrance of the hypomorphic lin-15 allele n765, consistent with a general decrease in X expression caused by ct31 ( Burgess et al. WBG 9.2, 1986). RNA dot-blot analyses indicated a slightly higher level (1.24x) of myo-2 mRNA in ct31 than in wild type. Since the ct31 strain has four copies of myo-2 compared to two copies in N2, this result may still reflect a reduction of X-chromosome expression. Further analysis of dosage compensation effects of ct28, ct30 and ct31 is in progress, as are experiments on the results of specifically varying sdc-2(+) dosage in the presence of these duplications.

Phuong Anh T Nguyen et al. (2005) International Worm Meeting "Suppressor analysis of the dietary ubiquinone requirement of clk-1 mutants."

Robyn Branicky, Siegfried Hekimi, Phuong Anh T Nguyen

[International Worm Meeting, 2005]

Ubiquinone (UQ or Coenzyme Q) is a lipid found in all cellular membranes. It is involved in multiple cellular processes, either directly or through its effect on the redox status of the cell. clk-1 encodes a highly conserved hydroxylase that is required for UQ biosynthesis (1). clk-1 mutants are devoid of UQ, and instead accumulate the UQ precursor, demethoxyubiquinone (DMQ) (2). Under normal culture conditions, clk-1 mutants are viable and healthy, presumably because they contain DMQ (3), which can partially replace UQ, and because they can use dietary UQ obtained from their bacterial food source (4). However, clk-1 mutants have slow development, reproduction and behaviors, a decreased brood size and an extended lifespan (5). Furthermore, in the absence of dietary UQ the mutants transiently arrest development, and grow up to become sterile adults after resuming development (6). Notably, animals carrying the missense allele clk-1(e2519), which produce a full length mutant protein (7), have a much less severe phenotype when compared to animals carrying the null mutation clk-1(qm30) (5), even though both mutants are completely devoid of UQ (2, 4). This suggests that, in addition to its function in UQ biosynthesis, CLK-1 could have another function that would not be entirely abolished by the e2519 mutation. In order to understand the pleiotropic effect of clk-1 mutations as well as the difference in severity of the two mutant alleles, we have been carrying out several suppressor screens on different aspects of the phenotypes produced by clk-1 mutations (8, 9). We have recently carried out screens for suppressors of the slow growth phenotype of clk-1 mutants on UQ+ bacteria as well as for suppressors of the growth arrest and sterility on UQ- bacteria. To date we have isolated nine mutants. All mutations are dominant, e2519-specific, suppress the dietary dependence, and map to more than 4 distinct loci. We will be presenting our progress in the phenotypic and molecular characterization of these mutants. (1).J. J. Ewbank et al., 1997. Science 275, 980-3. (2).H. Miyadera et al. , 2001. J Biol Chem 276, 7713-6. (3).A. K. Hihi et al., 2002. J Biol Chem 277, 2202-6. (4).T. Jonassen et al., 2001. PNAS 98, 421-6. (5).A. Wong et al., 1995. Genetics 139, 1247-59. (6).J. Burgess et al., 2003. J Biol Chem 278, 49555-62. (7).A. K. Hihi et al., 2003. J Biol Chem 278, 41013-8. (8).R. Branicky et al., 2001. Genetics 159, 997-1006. (9).Y. Shibata et al., 2003. Science 302, 1779-82.

Yu, Xinwei et al. (2021) International Worm Meeting "Fast deep learning correspondence for neuron tracking and identification in C. elegans using semi-synthetic training"

Sharma, Anuj K., Linderman, Scott W., Randi, Francesco, Yu, Xinwei, Leifer, Andrew M., Creamer, Matthew S.

[International Worm Meeting, 2021]

The nervous system of the nematode C.elegans is well characterized, such that each of the 302 neurons is named and has stereotyped locations across animals(Witvliet et al., 2020). The capability to find corresponding neurons across animals is essential to investigate neural coding and neural dynamics across animals. Despite the worm's overall stereotypy, the variability in neurons' spatial arrangement is sufficient to make predicting neural correspondence a challenge. We present an automated method to track and identify neurons in C. elegans, called "fast Deep Learning Correspondence" or fDLC, based on the transformer network architecture(Vaswani et al., 2017). The transformer has shown great success in natural language processing tasks by modeling the dependencies between words in a sentence. We reasoned this same architecture would be well-suited to extract spatial relationships between neurons in order to build a representation that facilitates finding correspondence to neurons in a template worm. The model is trained once on empirically derived semi-synthetic data and then predicts neural correspondence across held-out real animals via transfer learning. The same pre-trained model both tracks neurons across time and identifies corresponding neurons across individuals. Performance is evaluated against hand-annotated datasets, including NeuroPAL(Yemini et al., 2020). Using only position information, the method achieves 80.0% accuracy at tracking neurons within an individual and 65.8% accuracy at identifying neurons across individuals. Accuracy is even higher on a published dataset(Chaudhary et al., 2021). Accuracy reaches 76.5% when using color information from NeuroPAL. Unlike previous methods, fDLC does not require straightening or transforming the animal into a canonical coordinate system. The method is fast and predicts correspondence in 10 ms making it suitable for future real-time applications. Witvliet D, Mulcahy B, Mitchell JK, Meirovitch Y, Berger DR, Wu Y, et al.Connectomes across development reveal principles of brain maturation in C.elegans. bioRxiv. 2020;doi:10.1101/2020.04.30.066209. Vaswani A, Shazeer N, Parmar N, Uszkoreit J, Jones L, Gomez AN, et al.Attention is All You Need; 2017.Available from:https://arxiv.org/pdf/1706.03762.pdf. Yemini E, Lin A, Nejatbakhsh A, Varol E, Sun R, Mena GE, et al. NeuroPAL: A multicolor atlas for whole-brain neuronal identification in C. Elegans. Cell.2020;doi:10.1016/j.cell.2020.12.012. Chaudhary S, Lee SA, Li Y, Patel DS, Lu H. Graphical-model framework for automated annotation of cell identities in dense cellular images. eLife.2021;10:e60321. doi:10.7554/eLife.60321.

Villeneuve AM (1992) Worm Breeder's Gazette "Pairing of Homologous Chromosomes Inhibits Intrachromosomal Recombination"

Villeneuve AM

[Worm Breeder's Gazette, 1992]

ctDp31 is a large tandem chromosomal duplication of approximately 40% of the X chromosome (Tanner and Wood, 1991 Meeting Abstracts). Hermaphrodites homozygous for ctDp31 are small, Dpy, slow-growing and often Egl, and males hemizygous for ctDp31 are small, Dpy, slow-growing, and have abnormal gonads and genitalia, rendering them sterile. These phenotypes revert spontaneously to wild-type at a frequency of 104 - 10-5, concomitant with loss of the duplicated DNA (Tanner et al., WBG 10:3). Such revertants could be due either to unequal crossing over between homologous chromosomes, or to intrachromosomal recombination. Using the reversion of ctD31 as an assay, I have demonstrated that the absenee of a homologous chromosome that can act as a pairing partner dramatically increases the frequency of intrachromosomal recombination [See Figure 1]. This was shown in two ways, by measuring the reversion rate of ctDp31 in the germlines of her-1 ;ctDp3110 XO hermaphrodites and in the germlines of him-10 ;ctDp31 XX hermaphrodites. (him-10 is a mitotic mutant that undergoes premeiotic loss of X chromosomes, producing some XO germ cell nuclei in an XX germline [A.V., WBG, this issue].) Lack of a homolog during meiosis resulted in a 250 - >1000 fold increase in the frequency of intrachromosomal recombination. This indicates that the pairing of homologous chromosomes during meiosis strongly inhibits intrachromosomal recombination events. The meiotic behavior of this large tandem Dp reinforces the view that chromosome pairing initiates at a chromosome end and proceeds unidirectionally. This type of mechanism would preclude misalignment of the duplicated region despite the huge extent of homology. Further evidence that the two ctDp31 homologs are fully paired is provided by the fact that ctDp31 XX hermaphrodites produce >10-fold fewer spontaneous XO males than do wild-type hermaphrodites, implying a decrease in the incidence of non-recombinant X chromosomes. The ctDp31 reversion assay can be used to identify mutations that reduce recombination between homologs but still permit intrachromosomal recombination. This was first shown in the Wood lab, where it was found that him-8 (e1489)caused a >100 fold increase in ctDp31 reversion (Burgess et al., 1987 Meeting Abstracts). I have shown that two new alleles of him-8 (obtained in my general screen for mutants with increased non-disjunction) also cause increased ctDp31 reversion. Observation of increased levels of intrachromosomal recombination in this assay suggest that the him-8 mutations are defective in some aspect of homolog pairing, consistent with the altered distribution of interchromosomal recombination events observed in him-8 mutants by Broverman and Meneely (1991 Meeting Abstracts). Not all defects in the homolog pairing process are expected to share this property however. For example, most models for homolog recognition, a crucial phase in the pairing process, involve a mechanism to search for DNA sequence homology; such a homology search would be required for both inter- and intrachromosomal recombination events. [See Figure]