Larson, Braden John et al. (2019) International Worm Meeting "Redundant Mechanisms of X-Chromosome Repression in the C. elegans Male Germline."

Larson, Braden John, Strome, Susan

[International Worm Meeting, 2019]

Organization of the genome into domains of euchromatin and heterochromatin is a conserved and essential feature of all eukaryotes. Heterochromatin plays important roles in repression of transcription, chromosome segregation, and maintenance of genome integrity. Heterochromatin can be divided into constitutive heterochromatin and facultative heterochromatin, which can be distinguished by their associated histone modifications. Methylation of lysine 27 on histone H3 (H3K27me) is associated with facultative heterochromatin, while methylation of lysine 9 on histone H3 (H3K9me) is associated with constitutive heterochromatin. In mouse, fungus, and Drosophila, these two marks largely anticorrelate. However, in C. elegans, H3K27me and H3K9me show a surprising positive correlation, suggesting a species-specific difference in the organization of heterochromatin. In C. elegans, H3K27me represses transcription of the X chromosome in the germline, and its loss leads to a maternal-effect sterile (Mes) phenotype. Intriguingly, Gaydos et al. (Science, 2014) showed that mes mutant males that inherited their single X chromosome from the father are usually fertile and that the fertility of those males is dependent on H3K9me. Together, these observations suggest a potential redundant function of H3K27me and H3K9me in repressing the single X chromosome in the male germline to promote fertility in subsequent generations. We are investigating the potential redundant functions of H3K27me and H3K9me in the C. elegans male germline. First, by generating chimeric animals whose germlines inherit only paternal chromosomes, we have shown that mes mutant XX hermaphrodites are rendered fertile when both of their X chromosomes, which lack H3K27me, are inherited from the father. Furthermore, as in males, fertility is dependent on H3K9me. Second, we are comparing misexpression of genes and repetitive elements in hermaphrodite and male germlines lacking either H3K27me or H3K9me or lacking both marks, to investigate cross-talk between those two marks and differential responses of hermaphrodite (XX) and male (XO) germlines. Lastly, we are examining other species of Caenorhabditis. Early results suggest that C. briggsae accomplishes X-chromosome repression differently than C. elegans.

Larson, Braden John et al. (2021) International Worm Meeting "Redundant Mechanisms of X Chromosome Repression in the C. elegans Male Germline"

Larson, Braden John, Strome, Susan, Egelhofer, Thea

[International Worm Meeting, 2021]

Organization of the genome into domains of euchromatin and heterochromatin is a conserved feature of all eukaryotes and precise regulation of these domains is important for organism health and development. Proper formation of heterochromatin is crucial for transcriptional repression, chromosome segregation, and maintenance of genome integrity. Heterochromatin can be categorized as either facultative or constitutive. These two types of heterochromatin are often distinguished by their associated histone modifications: methylation of lysine 27 or lysine 9 on histone H3. H3K27me is associated with facultative heterochromatin, and its domains are found throughout genomes, often with developmentally regulated genes. H3K9me is associated with constitutive heterochromatin, and is generally concentrated in gene-poor, repeat-rich regions such as pericentric regions. Thus, anticorrelation of H3K27me and H3K9me domains is observed in many model organisms. However, in C. elegans, H3K27me and H3K9me domains show a surprising amount of positive correlation, suggesting a species-specific mechanism for organizing facultative and constitutive heterochromatin. In the C. elegans germline, H3K27me is enriched on the X chromosome, and its loss leads to sterility in the F2 generation. Interestingly, H3K27me(-) F2 males that inherit a paternal X chromosome (Xp) and no maternal X chromosome (Xm) are usually fertile. The fertility of these males is dependent on H3K9me, which is enriched on the single X chromosome in the male germline. These observations suggest a potential redundant function of H3K27me and H3K9me in repressing the X chromosome in the male germline to promote fertility in subsequent generations. We are investigating the organization and functions of H3K27me and H3K9me in the C. elegans male germline. By generating chimeric animals whose germlines inherit only paternal chromosomes (XpXp), we've shown that a double dose of paternal chromosomes lacking H3K27me can also support fertile hermaphrodite germline development in an H3K9me-dependent manner. We are comparing misexpression of genes and repetitive elements in hermaphrodite and male germlines lacking H3K27me, H3K9me, or both. Lastly, we are using CUT&RUN to examine the distribution of H3K27me in germ cells and to test whether H3K9me regulates this distribution, as it does in Mouse and Neurospora.

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.

Phuongnhung Nguyen et al. (2006) Neuronal Development, Synaptic Function, and Behavior Meeting "Dissecting Genetically the Neural Networks that Coordinate Locomotion"

Bill Walthall, Gennady Cymbalyuk, Phuongnhung Nguyen

[Neuronal Development, Synaptic Function, and Behavior Meeting, 2006]

The networks of nerves and muscles responsible for forward and backward movements in C. elegans offer unique advantages for investigating the functional relationships among cells in integrated cellular networks. The first advantage is that John Sulston and his colleagues identified the interneurons, motor neurons and muscle cells that compose the networks (Sulston, et al., 1983). Second, John White and colleagues determined the synaptic patterns that interconnect the network (White et al., 1986). Finally, the C. elegans community has generated mutations that cause specific alterations in the cellular networks. A model based on the connectivity pattern and laser ablation results suggests that distinct sets of interneurons and excitatory motor neurons are dedicated to forward and backward movement, respectively. These two circuits converge upon two classes of inhibitory motor neurons and four longitudinal strands of body wall muscles and create antiphasic contractile muscular waves that travel along the dorsal and ventral axes of the body. We will report results from the analysis of movies showing locomotion patterns of animals. These movies allows us to quantitatively characterize the traveling wave in terms of amplitudes, dorsal and ventral deflections, frequency and velocity of the wave progression and the forward and backward velocity of the animal's progression. We will compare wild-type locomotion characteristics with those exhibited by three uncoordinated mutants (unc-4, unc-55, and cnd-1) that are known to make specific alterations in the network that result in movement defects that are predicted by the model. The connectivity model can also be used to predict epistatic relationships for double mutant combinations (for example unc-55 mutants exhibit a ventral asymmetric pattern of backward movement, whereas unc-4 exhibits a dorsal asymmetric pattern of backward movement). We find that unc-4 masks the defect of unc-55 mutants, which is predicted by the model neural circuit. However the model's predictions for forward movement are not as accurate. We will discuss possible explanations for the resiliency of forward movement and the vulnerability of backward movement to genetic perturbations.

Consortium Wormbase et al. (2000) West Coast Worm Meeting "From ACeDB to a more complete and usable database"

Consortium Wormbase, Foltz NL, Schwarz EM, Sternberg PW

[West Coast Worm Meeting, 2000]

We briefly describe the current status and plans for WormBase, initially an extension of the existing ACeDB database with a new user interface. The WormBase consortium includes the team that developed ACeDB (Richard Durbin and colleagues at the Sanger Centre; Jean Thierry-Mieg and colleagues at Montpellier); Lincoln Stein and colleagues at Cold Spring Harbor, who developed the current web interface for WormBase; and John Spieth and colleagues at the Genome Sequencing Center at Washington University, who along with the Sanger Centre team, continue to annotate the genomic sequence. The Caltech group will curate new data including cell function in development, behavior and physiology, gene expression at a cellular level, and gene interactions. Data will be extracted from the literature, as well as by community submission. We look forward to providing the C. elegans and broader research community easy access to vast quantities of high quality data on C. elegans. Also, we welcome your suggestions and criticism at any time. WormBase can be accessed at www.wormbase.org.

Ken-ichi Oshio et al. (2002) Japanese Worm Meeting "Revision of synaptic connectivity database"

Eizo Akiyama, Yuishi Iwasaki, Yuko Osana, Ken-ichi Oshio, Sohei Gomi, Satoru Morita, Kotaro Oka, Kazumi Omata

[Japanese Worm Meeting, 2002]

The synaptic connectivity of C. elegans is well known from observations of the somatic system by White et al. and those of the pharyngeal system by Albertson et al. So far, three databases were constructed for computational usage by Achacoso et al. and Durbin, and recently in WormBase. However, they lack some data such as those in tables of White's paper and those in figures of Albertson's book. Our database (K. Oshio, S. Morita, Y. Osana and K. Oka: Technical Report of CCEP, Keio Future No.1, 1998) includes all data described in White's paper and Albertson's book. Unfortunately, some mistakes were found in the database through private communications with John White who is the author of White's paper and with the users of the database. Thus we have been proceeding with the revision to make it perfect one. We are planning to complete the revision in September 2002. The database should be worthwhile not only for neurophysiological studies, but also for post-genomic interests mediating genomes and behavior.

Kevin Eliceiri et al. (2006) Neuronal Development, Synaptic Function, and Behavior Meeting "A Swept-Field Confocal for Cell Biology Studies"

Kevin Eliceiri, Mike Szulczewski

[Neuronal Development, Synaptic Function, and Behavior Meeting, 2006]

We describe a novel instrument, The Swept-Field Confocal Microscope (SFC) that combines high resolution pinhole imaging with slit imaging. Unlike spinning disk confocal systems that have their pinhole apertures embedded in a spinning disc, the SFC's 32 pinhole array remains stationary. Galvonometric and piezo controlled mirrors sweep the image of the pinholes across the sample. The emission photons are de-scanned and focused through a complementary set of pinholes onto a CCD camera. This results in a high resolution image that can be collected at up to 100 frames/second in the pinhole mode and greater than 1000 frames per second in the slit mode. The ability for the scientist to match the optical recording with the temporal biological fluorescence response has great promise for cell and developmental biology studies where live dynamic events must be captured quickly in high resolution. In this poster we describe the optical path of the microscope and present some early data in collaboration with John White's group at the UW-Madison of using the SFC to image GFP constructs in C .elegans to investigate cell division in the early embryo.

Tylon Stephney et al. (2007) International Worm Meeting "WormImage: An online photo album for C. elegans electron microscopy."

Huawei Weng, Maurice Volaski, David H. Hall, Toni Long, Kevin Fisher, Meng Xu, Tylon Stephney

[International Worm Meeting, 2007]

The Hall lab is the home for the physical EM data for C. elegans that was collected by many scientists working at several key laboratories over the past 40 years. We have almost all of the annotated prints, negatives, data notebooks and often the original thin sections and blocks from the MRC, Missouri, Caltech and AECOM collections, including the work of John White, John Sulston, Sam Ward, and many others. This archive includes wild type and mutant data, and covers both males and hermaphrodites, and all ages from embryo to adulthood, and some aging animals. We are scanning much of this physical print data to produce a digital archive of C. elegans anatomy. We have generated about 3 terabytes of digital images, and there is much more scanning to be done. We have designed a simple online photo album, WormImage, to share some of this archive as small thumbnail versions that can quickly transit across the Internet to all users, for free. Bandwidth issues limit our ability to ship full size scans electronically, but we also supply users with higher resolution scans upon request, by ftp or on DVDs. WormImage (www.wormimage.org) is an online database developed for remote retrieval of this digitized microscopy data. Currently this database contains about 20,000 different digital images. Most scans have been taken from the workprints, to preserve original hand annotations which mark identified cell types. The WormImage database is updated weekly to provide more images. The user can search this database by animal name, age, sex, or by regional information (head, midbody, tail) or by tissue types to identify potential images of interest. The user can quickly survey many animals in small thumbnail images, or concentrate on details of a few images expanded to larger size. The program was recently revised to make it easy for the user to click through all the thumbnails for one animal in serial order. Hall has been reviewing the original annotations and writing brief summaries for each animal, also offered on the website. These Color Code summaries help to translate the shorthand markings on the original prints into the familiar cell names for many structures. During 2007, WormImage will begin showing images of key mutant phenotypes and aging animals. We are grateful to Demian Nave and Art Wetzel at the Pittsburgh Supercomputing Center for their help in providing a mirror site. This work is funded by NIH RR 12596.

Karla Opperman et al. (2007) International Worm Meeting "A structure-function analysis of the C. elegans L1CAMs."

Lihsia Chen, Xuelin Wang, Karla Opperman, Shan Zhou

[International Worm Meeting, 2007]

Cell adhesion molecules play important roles during development and in maintaining tissue integrity. L1CAMs are a family of immunoglobulin (Ig)-like cell adhesion molecules that are important in vertebrate nervous system development. L1CAMs are conserved in C. elegans, and are encoded by the lad-1/sax-7 and lad-2 genes. SAX-7 and LAD-2 have distinct functions in the nervous system despite overlapping neuronal expression. SAX-7 is required to maintain neuronal positioning while LAD-2 participates in axon pathfinding. We are performing a structure-function analysis to determine how both proteins mediate their functions. Because the cytoplasmic tails between both proteins are distinct, we are focusing our analysis on how the cytoplasmic tail regulates L1CAM functions. Indeed, while the extracellular domains of SAX-7 and LAD-2 are conserved, the LAD-2 cytoplasmic tail is completely divergent and does not contain any of motifs conserved in SAX-7 and vertebrate L1CAMs. These motifs include the ankyrin-binding motif, which links L1CAMs to the spectrin-actin cytoskeleton, the PDZ-binding motif, and conserved tyrosine residues that are phosphorylated. Our preliminary results reveal that both conserved motifs, as well as phosphorylation one of the tyrosine residues, which regulates ankyrin binding, all contribute to the function of SAX-7 in neuronal position maintenance. (1) Chen L, Ong B, Bennett V. J Cell Biol. 2001 Aug 20;154(4):841-55. (2) Wang X, Kweon J, Larson S, Chen L. Dev Biol. 2005 Aug 15;284(2):273-91. (3) Sasakura H, Inada H, Kuhara A, Fusaoka E, Takemoto D, Takeuchi K, Mori I. EMBO J. 2005 Apr 6;24(7):1477-88. Epub 2005 Mar 17.

Chan S et al. (1991) International C. elegans Meeting "SEEKING UNC-40."

Culotti JG, Su M-W, Chan S, Hedgecock EM

[International C. elegans Meeting, 1991]

unc-40 is required with unc-6 to guide ventral migrations on the nematode epidermis (Hedgecock et al., Neuron 4, 61-85, 1990). We have positioned unc-40 genetically by 3-factor crosses with dpy-5 and bli-4 as flanking markers and we have isolated 5 new alleles of unc-40 (2 spontaneous, 2 gamma, and 1 EMS) making a total of 12. DNAs from cosmids covering this region (obtained from Alan Coulson and John Sulston) are being used to probe DNA and RNA from the mutants and DNA from strains carrying duplications that break to either side of unc-40 (obtained from Anne Rose's laboratory). We are also injecting cosmid DNAs into gonads to look for rescue by germline transformation. The brood sizes of various unc-40 alleles are very small, so we have resorted to injecting the wild type in order to create a series of transformed lines, each carrying an extrachromosomal array of a different cosmid. Each array will be passed into appropriately marked unc-40 animals to ask if any can rescue the mutant phenotype. Mosaic experiments are also underway to determine whether unc-40 acts in migrating cells.