Lee, Garrett et al. (2019) International Worm Meeting "Understanding the development, plasticity, and function of synaptic asymmetry in C. elegans."

Tang, Leo Tsz-Ho, Emmons, Scott, Buelow, Hannes, Lee, Garrett, Cook, Steven

[International Worm Meeting, 2019]

Lateralization of the nervous system is conserved across the animal kingdom from invertebrates to humans. The human brain asymmetrically processes faculties across the left and right hemispheres, and impaired functional lateralization has been implicated in depression, schizophrenia, and autism spectrum disorders. However, whether asymmetric connectivity exists in anatomically symmetric structures and what its function may be remains largely unknown. Serial electron micrograph (EM) reconstruction of the C. elegans connectome revealed that the chemosensory ASE neurons exhibit left-right asymmetry in their connection to the odorsensory AWC neurons. While morphologically symmetric, ASEL and ASER exhibit differential gene expression and function. ASEL primarily senses Na+ while ASER responds to Cl-. To confirm the EM data as well as study the connection's development, maintenance, plasticity, and function, we created a fluorescent reporter of this connection using in vivo Biotin Labeling of Intercellular Contacts (iBLINC). iBLINC analysis confirmed that ASEL tends to form more synaptic contacts onto AWC than ASER. Analysis of connectivity over development revealed that the asymmetry is established by L3 and maintained through adulthood. Genetic conversion of both ASE neurons to an ASEL fate reversed the asymmetry. In contrast, neither changing both ASE neurons to the ASER fate nor symmetrizing the postsynaptic AWC neurons displayed altered synaptic lateralization from wildtype. These findings suggest that pre-synaptic identity contributes to establishment of the asymmetric ASE/AWC connection. Interestingly, the left-right lateralization is changed when C. elegans are exposed to different salt environments, suggesting that the lateral connectivity is plastic. Finally, we have identified several genes that are required to establish or maintain the ASE/AWC synaptic asymmetry. In analyzing this connection, we aim to understand fundamental aspects of the formation and function of asymmetric connectivity.

Lee, Garrett et al. (2021) International Worm Meeting "Understanding the development, plasticity, and function of synaptic asymmetry in C. elegans"

Emmons, Scott, Cook, Steven, Tang, Leo, Buelow, Hannes, Lee, Garrett

[International Worm Meeting, 2021]

Lateralization of the nervous system is conserved across the animal kingdom from invertebrates to humans. The human brain asymmetrically processes faculties across the left and right hemispheres, and impaired functional lateralization has been implicated in depression, schizophrenia, and autism spectrum disorders. However, whether asymmetric connectivity exists in anatomically symmetric structures and what its function may be remains largely unknown. Serial electron micrograph (EM) reconstruction of the C. elegans connectome revealed that the chemosensory ASE neurons exhibit left-right asymmetry in their connection to the odorsensory AWC neurons. While morphologically symmetric, ASEL and ASER exhibit differential gene expression and function. ASEL primarily mediates attraction to increases in Na+ concentration, while ASER triggers avoidance of decreases in Cl- concentration. To confirm the EM data as well as study the connection's development, maintenance, plasticity, and function, we created a fluorescent reporter of this connection using in vivo Biotin Labeling of Intercellular Contacts (iBLINC). iBLINC analysis confirmed that ASEL tends to form more synaptic contacts onto AWC than ASER. Analysis of connectivity over development revealed that the asymmetry is established by L3 and maintained through adulthood. Genetic conversion of both ASE neurons to an ASEL fate reversed the asymmetry. In contrast, neither changing both ASE neurons to the ASER fate nor symmetrizing the postsynaptic AWC neurons displayed altered synaptic lateralization from wildtype. These findings suggest that pre-synaptic identity contributes to establishment of the asymmetric ASE/AWC connection. Interestingly, the left-right lateralization is reversed when C. elegans are exposed to increased NaCl environments. Synaptic number changes progressively over the course of twelve hours and in a dose-dependent manner, suggesting that the lateral connectivity is plastic. Finally, we have identified several genes that are required to establish or maintain the ASE/AWC synaptic asymmetry. Future experiments will characterize the functional significance of left-right asymmetry using Ca2+ imaging. In analyzing this connection, we aim to understand fundamental aspects concerning the formation and function of asymmetric connectivity.

Abate JP et al. (2009) Cell Metab "Life is short, if sweet."

Blackwell TK, Abate JP

[Cell Metab, 2009]

Insulin is essential for glucose homeostasis, but reducing its activity delays the aging process in model organisms. In this issue of Cell Metabolism, Lee et al. (2009) show how these effects of insulin signaling intersect when glucose is fed to C. elegans.

Lee C et al. (2017) Bio Protoc "Single-molecule RNA Fluorescence in situ Hybridization (smFISH) in Caenorhabditis elegans."

Sorensen EB, Lee C, Seidel HS, Lynch TR, Crittenden SL, Kimble J

[Bio Protoc, 2017]

Single-molecule RNA fluorescence <i>in situ</i> hybridization (smFISH) is a technique to visualize individual RNA molecules using multiple fluorescently-labeled oligonucleotide probes specific to the target RNA ( Raj <i>et al.</i>, 2008 ; Lee <i>et al.</i>, 2016a ). We adapted this technique to visualize RNAs in the <i>C. elegans</i> whole adult worm or its germline, which enabled simultaneous recording of nascent transcripts at active transcription sites and mature mRNAs in the cytoplasm ( Lee <i>et al.</i>, 2013 and 2016b). Here we describe each step of the smFISH procedure, reagents, and microscope settings optimized for <i>C. elegans</i> extruded gonads.

Dawes-Hoang RE et al. (2003) Dev Cell "Bringing classical embryology to C elegans gastrulation."

Zallen JA, Wieschaus EF, Dawes-Hoang RE

[Dev Cell, 2003]

In a recent paper, Lee and Goldstein develop an explant assay that recapitulates key aspects of gastrulation in C. elegans and permits classical embryological manipulations. The resulting detailed analysis of cell behavior will ultimately extend to broader issues, such as, whether morphogenesis can be described as the sum of single-cell events or if unique phenomena emerge at the multicellular level.

Tang, Leo et al. (2021) International Worm Meeting "Insulin-like signaling regulates left/right asymmetric synaptic connection"

Lee, Garrett, Emmons, Scott, Cook, Steven, Tang, Leo, Buelow, Hannes

[International Worm Meeting, 2021]

Lateral specialization of the central nervous system is a well-established feature across species, yet the underlying mechanism through which functional asymmetry arises is largely unknown. EM reconstruction of the C. elegans connectome found that ASE to AWC synapses were stronger on the left than the right. To corroborate and elucidate this asymmetric connection, we generated a reporter strain labelling ASE-&gt;AWC connections using in vivo Biotin Labelling of Intercellular Contact (iBLINC). While we observed the same left-sided bias of this connection, we also discovered that the asymmetric fates of the ASEs are a necessary but not sufficient factor in establishing this left-side bias. Furthermore, we found that ins-6/insulin-like is involved in the establishment of this asymmetric connection. Using a fosmid-based ins-6::GFP reporter, we observed that ins-6 expression in ASJ also exhibits left-sided bias. The asymmetry of ASE to AWC iBLINC signal is abolished in cell-specific knock-out animals of ins-6 in ASJ but not in ASI. Moreover, genetically removing ins-6 in ASJL by use of tbx-37p::Cre reversed the asymmetry of the ASE to AWC connection. Meanwhile, removing the putative ins-6 receptor daf-2 in ASEL but not ASER symmetrized the ASE to AWC connection. Finally, we observed that mutation on an antagonistic insulin, ins-22, partially suppressed the phenotype of ins-6 mutants. These results taken together suggest that the left-side bias of ASE to AWC connection is controlled by insulin signaling, where asymmetrically expressed insulin-like molecules from ASJs act locally to regulate connectivity of the ASE&gt;AWC synaptic connection. We aim to further investigate the effect of insulin signaling on the plasticity of ASE to AWC connection and general synapse dynamics. We also aim to characterize the previously unreported asymmetric gene expression in ASJ

Bauer, Jack et al. (2021) MicroPubl Biol

Labb, Jean-Claude, Lacroix, La, Bauer, Jack

[MicroPubl Biol, 2021]

To perturb actomyosin function in the primordial germ line, we first monitored germ line organization in L1-stage animals bearing temperature-sensitive (ts) alleles in genes encoding actomyosin regulators and that were reported to interfere with cytokinesis during embryogenesis (Davies et al. 2014). Previous work demonstrated that the initial stages of germline expansion occur normally in cyk-4(ts) and zen-4(ts) animals raised at restrictive temperature from the L1 stage (Lee et al. 2018). We found that primordial germ line organization in cyk-1(ts), nmy-2(ts), cyk-4(ts) or zen-4(ts) L1 larvae maintained at restrictive temperature for 12h was no different than control (Figure 1A-B). Furthermore, the first primordial germ cell (PGC) division occurred normally upon feeding these animals at restrictive temperature with typical bacterial food (E. coli OP50). As noted previously (Lee et al. 2018), germ line disorganization and sterility were observed in all cases when animals reached adulthood (Figure 1B).

Mondol V et al. (2012) Curr Top Dev Biol "Let's make it happen: the role of let-7 microRNA in development."

Mondol V, Pasquinelli AE

[Curr Top Dev Biol, 2012]

Noncoding RNAs have emerged as an integral part of posttranscriptional gene regulation. Among that class of RNAs are the microRNAs (miRNAs), which posttranscriptionally regulate target mRNAs containing complementary sequences. The broad presence of miRNAs in lower eukaryotes, plants, and mammals highlights their importance throughout evolution. MiRNAs have been shown to regulate many pathways, including development, and disruption of miRNA function can lead to disease (Ivey and Srivastava, 2010; Jiang et al., 2009). Although the first miRNA genes were discovered in the nematode, Caenorhabditis elegans, almost 20 years ago, the field of miRNA research began when they were found in multiple organisms a little over a decade ago (Lagos-Quintana et al., 2001; Lau et al., 2001; Lee and Ambros, 2001; Lee et al., 1993; Pasquinelli et al., 2000; Wightman et al., 1993). Here, we review one of the first characterized miRNAs, let-7, and describe its role in development and the intricacies of its biogenesis and function.

Ho F et al. (1992) Worm Breeder's Gazette "The Second Approach to Aligning the Genetic and Physical Maps on Chromosome I: PCR mapping of Potential Deletions"

Ho F, Janmaat A, Ha TT, Melathopoulos A, Liu T, Rose AM, Salmon R

[Worm Breeder's Gazette, 1992]

For the right portion of the chromosome, where we don't have a high density of mapped mutants, we are using the PCR mapping protocol of Barstead and Waterston (in 1991 C Mot Cyto 20:69) to position gamma-induced lethals (potential deletions). Using hIn1 as a balancer (Zetka and Rose, 1992 Genetics), we recovered several lethals linked to unc-101 and lev-11 which were induced with 1500 rads of gamma radiation. Primers were kindly provided by Junho Lee in Paul Sternberg's lab and Yuko Kozono in Bob Waterstons's lab. As controls for the PCR reactions, we are using a deletion outside the hIn1 region, hDf8 ,and primers from bli-4 (Ken Peters, unpublished) and AHH sequences (Prasad, Starr and Rose, in press). hDf8 deletes AHH, but not bli-4 .We have tested nine of our gamma induced lethals to date and one of these hDf14 (h1277 )is lacking DNA for the primer set INT 7-10 from unc-101 sequences provided by Junho Lee.

Miranda MC et al. (2024) MicroPubl Biol "Investigating the effects of antipsychotic drugs as a treatment for improving the activity of the unc-33 /Dpysl2 gene in C. elegans."

Miranda MC, Hyde J, Holgado A, Salazar K, Bell B

[MicroPubl Biol, 2024]

Prenatal stress is hypothesized to contribute to the development of schizophrenia. Lee and colleagues determined that prenatal stress in rats decreases levels of Dpysl2, which is found to be inactivated in schizophrenic patients. UNC-33 , the homolog to Dpysl2 in <i>C. elegans</i> , is important for axonal outgrowth and synapse formation. Herein, we study the effects of antipsychotic drugs on developing <i>C.elegans</i> exposed to stress through high temperatures. Results indicate that the <i>unc-33</i> promoter was not impacted by antipsychotic drug treatment, but the lifespan was decreased.