The guided migration of neurons and neuronal progenitors in the nervous systems is a complex process involving extracellular cues that act through specific receptors and downstream signals to direct cell movements. The two bilateral Q neuroblasts in C. elegans offer a simple and genetically accessible model of neuronal progenitor migration. Despite identical differentiation patterns, QR on the right migrates anteriorly, whereas QL on the left moves toward the posterior. Thus, the Q cells express a shared program of differentiation but offer opposite responses to signals that direct cell migration. This proposal is designed to dissect a novel role of the CELF1 (CUG binding protein, Elav Family 1) molecule ETR-1 in Q neuroblast migration in C. elegans. CELF1 molecules are RNA-binding proteins that mediate multiple aspects of RNA processing, including alternative splicing. In an unbiased forward genetic screen for new mutations with defects in Q neuroblast migration, we identified a novel allele of etr-1, which encodes the C. elegans version of CELF1. Previous studies showed that etr-1 loss-of-function results in embryonic lethality with severe muscle disorganization, including defects in muscle attachment. The etr-1(lq61) mutation that we identified is viable, and corresponds to a premature stop codon in an alternatively-spliced exon that is limited to a subset of etr-1 transcripts. Thus, etr-1(lq61) offers a unique opportunity to probe the function of etr-1/CELF1 and the regulation of alternative mRNA processing in both muscle cell development and Q neuroblast migration. etr-1 expression in C. elegans is limited to body wall muscles. This observation suggests that etr-1 may exert an indirect effect on Q neuroblast migration, possibly via a secreted or transmembrane factor expressed from adjacent muscle cells. Indeed, preliminary results indicate that etr-1 acts in muscles in Q migrations. Q-cell migration is highly dependent on body muscle-derived Wnt signals. It is possible, for example, that etr-1 could regulate expression of a key component of the Wnt secretory pathway. On the other hand, additional aspects of Q cell migration do not require Wnt and are therefore likely to depend on alternative extracellular cues. The existence of a diverse set of transmembrane proteins that function in Q cells to regulate their migration, (e.g., UNC-40/DCC, PTP-3/LAR, MIG-21, MIG-13) (3, 4), underscores the importance of these additional intercellular signals to Q-cell trajectory. We hypothesize that ETR-1/CELF-1 regulates expression of key signals that are produced in muscle cells to guide Q neuroblast migration.
In aim 1, we will use cell-specific knock-down and rescue to determine if ETR-1 acts in muscles for Q migrations.
In aim 2, we will use fluorescent-activated cell sorting of body wall muscles combined with RNA-seq in wild-type and etr-1 mutants to define transcripts affected by etr-1 that might regulate Q migrations. Because of our novel and innovative approach, new genes and gene interactions that regulate neuroblast migration will be discovered.
Directed neuronal migration is a key aspect of nervous system development in which neurons are guided to their targets via cues from surrounding tissues. This proposal is aimed at understanding how the CELF1 RNA binding protein regulates RNA processing (e.g. alternative splicing) involved in producing guidance cues from body wall muscles that direct neuronal migration the model organism nematode worm C. elegans.
