Melissa Cizeron

Institute NeuroMyoGene; Lyon, France

KX89

Caenorhabditis elegans

Zhou, Xin et al. (2021) International Worm Meeting "The HSPG Syndecan is a core organizer of cholinergic synapses in C. elegans"

Cizeron, Melissa, Zhou, Xin, Romatif, Oceane, Vachon, Camille, Jospin, Maelle, Bessereau, Jean-Louis, Bulow, Hannes E.

[International Worm Meeting, 2021]

The extracellular matrix has emerged as an active component of chemical synapses regulating synaptic formation, maintenance and homeostasis. The heparan sulfate proteoglycan syndecans are known to regulate cellular and axonal migration in the brain. They are also enriched at synapses, but their synaptic functions remain more elusive. Here we show that SDN-1/Syndecan is a key organizer of the neuromuscular junctions (NMJs) and is the core component that clusters the homomeric alpha7-like nicotinic receptor ACR-16 at cholinergic NMJs. SDN-1 is concentrated at neuromuscular junctions (NMJs) by the neurally-secreted synaptic organizer MADD-4/Ce-Punctin. Punctin is secreted by cholinergic and GABAergic motoneurons into the synaptic cleft and triggers appropriate postsynaptic differentiation. We now show that Punctin has at least two parallel functions at cholinergic NMJs. First, it localizes the syndecan produced by muscle cells at postsynaptic sites. Second, it concentrates and likely activates the netrin receptor UNC-40/DCC. Those cooperatively recruit the FRM-3/FARP and LIN-2/CASK through direct interaction with the PDZ domain binding site of SDN-1 and the C-terminal P3 domain of UNC-40, respectively. The FERM-FA domain of FRM-3/FARP also engages direct interaction with SDN-1, likely with its submembrane C1 domain. The resulting CASK/FARP/Syndecan complex localizes N-AChRs at cholinergic NMJs through physical interactions. Interestingly, we were able to relocalize N-AChRs at GABAergic NMJs using a chimera containing the extracellular domain of the NLG-1/Neuroligin and the intracellular domain of SDN-1. Therefore, SDN-1 stands at the core of the cholinergic synapse organization by bridging the extracellular synaptic determinants to the intracellular synaptic scaffold that controls the postsynaptic receptor content. Surprisingly, the molecular mechanisms that control alpha7-like nicotinic receptors localization and dynamics are still largely unknown. Because all the components identified in C. elegans are evolutionarily conserved and expressed in mammalian neurons, our results provide a new framework to test if syndecan regulates the localization of nicotinic receptors in the mammalian brain.

KX54

Caenorhabditis elegans

Craig Ceol et al. (2001) International C. elegans Meeting "IDENTIFICATION AND CHARACTERIZATION OF lin-35 Rb PATHWAY GENES THAT ANTAGONIZE RAS SIGNALING DURING VULVAL INDUCTION"

Bob Horvitz, Craig Ceol

[International C. elegans Meeting, 2001]

The synthetic multivulva (synMuv) class A and class B genes define two functionally redundant pathways that antagonize Ras signaling during vulval induction. Cloned class B synMuv genes encode proteins similar to the mammalian tumor suppressor pRB and the pRB-binding heterodimeric transcription factor DP/E2F. We propose that the class B synMuv proteins act as transcriptional repressors to negatively regulate Ras signaling. Specifically, we propose that a DPL-1 DP/EFL-1 E2F heterodimer binds DNA and recruits LIN-35 Rb and other synMuv proteins involved in chromatin modification, including LET-418 Mi-2, LIN-53 RbAp48 and HDA-1 HDAC, to repress the transcription of genes required for vulval cell-fate specification. To further investigate this RB signaling pathway, we cloned additional known class B synMuv genes and performed a genetic screen to identify new class B synMuv genes. The class B synMuv genes lin-52 and lin-54 were identified by Chip Ferguson and Jeff Thomas, respectively, two former graduate students in our laboratory. We cloned both genes and found that each encodes a protein of unknown function. We obtained candidate null alleles of each gene and are using these alleles to characterize the multivulva, sterile and other abnormalities caused by lin-52 or lin-54 loss of function. We are focusing additional functional studies on lin-54 . We are currently assessing whether a human lin-54 - like gene can functionally substitute for lin-54 and are conducting a range of protein-protein interaction studies with LIN-54 and the human LIN-54-like protein. Together with graduate students Frank Stegmeier and Melissa Harrison we screened for additional class B synMuv genes. Using a lin-15A background, we screened 6,500 mutagenized haploid genomes and obtained at least 51 class B synMuv mutations. Our analyses to date have identified at least five new candidate class B synMuv genes. We will describe our genetic and molecular characterizations of these genes.

Craig Ceol et al. (2002) East Coast Worm Meeting "CHARACTERIZATION OF lin-54 AND OTHER GENES THAT NEGATIVELY REGULATE let-60 Ras SIGNALING DURING VULVAL DEVELOPMENT"

Craig Ceol, Bob Horvitz

[East Coast Worm Meeting, 2002]

The synthetic multivulva (synMuv) class A and class B genes act redundantly to negatively regulate Ras signaling during vulval development. The class B synMuv genes encode proteins that likely function to target the histone deacetylase HDA-1 to specific DNA sequences. Some of the proteins involved in this proposed targeting, including LIN-35 RB, DPL-1 DP and EFL-1 E2F, are conserved in mammals, and a similar targeting function has been described for their mammalian counterparts. To better understand this conserved pathway, we cloned the class B synMuv gene lin-54 and identified new synMuv genes in a genetic screen. lin-54 was originally identified and mapped to a small region of LGIV by Jeff Thomas, a former graduate student in our laboratory. We cloned lin-54 and found that it encodes a protein with two copies of a cysteine-rich domain. A similar domain is found in proteins in other species; however, the function of this domain is not understood. We observed partial maternal rescue of the Muv phenotype of lin-54 mutants, indicating that maternally-provided lin-54 can regulate the vulval cell fate specification decisions that occur during postembryonic development. However, the stage-specific ectopic expression of lin-54 under the control of heat shock promoters indicated that lin-54 need not function prior to vulval induction to regulate this process. In addition, we conducted in vitro protein interaction experiments using LIN-54 and other class B synMuv proteins. We will present preliminary data concerning these studies. With graduate students Frank Stegmeier and Melissa Harrison, we screened approximately 6500 haploid genomes and recovered 95 new synMuv mutations. The mutations we assigned to complementation groups fall into three general classes: 1) mutations that affect new and previously characterized class B synMuv genes, 2) mutations that affect ark-1, sli-1 and gap-1, genes that are thought to directly regulate Ras pathway components, and 3) mutations that synergize with class A and class B synMuv genes and define previously uncharacterized genes (see abstract by Ceol and Horvitz). We will present our characterization of these new mutations.