The Predicted RNA-Binding Protein ETR-1/CELF1 Acts in Muscles To Regulate Neuroblast Migration in Caenorhabditis elegans

This work is licensed under a Creative Commons Attribution 4.0 International License.Neuroblast migration is a critical aspect of nervous system development (e.g., neural crest migration). In an unbiased forward genetic screen, we identified a novel player in neuroblast migration, the ETR-1/CELF1 RNA binding protein. CELF1 RNA binding proteins are involved in multiple aspects of RNA processing including alternative splicing, stability, and translation. We find that a specific mutation in alternatively-spliced exon 8 results in migration defects of the AQR and PQR neurons, and not the embryonic lethality and body wall muscle defects of complete knockdown of the locus. Surprisingly, ETR-1 was required in body wall muscle cells for AQR/PQR migration (i.e., it acts cell non-autonomously). Genetic interactions indicate that ETR-1 acts with Wnt signaling, either in the Wnt pathway or in a parallel pathway. Possibly, ETR-1 is involved in the production of a Wnt signal or a parallel signal by the body wall muscles that controls AQR and PQR neuronal migration. In humans, CELF1 is involved in a number of neuromuscular disorders. If the role of ETR-1/CELF1 is conserved, these disorders might also involve cell or neuronal migration. Finally, we describe a technique of amplicon sequencing to detect rare, cell-specific genome edits by CRISPR/Cas9 in vivo (CRISPR-seq) as an alternative to the T7E1 assay.NIH P40 OD010440National Institute of General Medical Sciences (P20GM103638)Madison and Lila Self Graduate Fellowship progra

New roles for Hox and Wnt in Cell Migration

Neuron migration is a critical process during central nervous system development. In the dissertation below I report new roles for established genes such as Wnt and Hox, and describe roles for several new genes in neuron migration. To study migrating neurons I use the model organism nematode worm Caenorhabditis elegans. With only 302 neurons, C. elegans is an excellent model to examine migration of individual neurons. The QR and QL neuroblasts are two bilaterally symmetric neural progenitor cells born in the posterior region of the worm. Although born in the same area, these cells undergo opposite directions of migration. QR migrates anteriorly, and QL migrates posteriorly. In QL, detection of extracellular EGL-20/Wnt results in trasncription of mab-5/Antennapedia/Hox through canonical Wnt signalling. MAB-5 is a posterior migration determinant of the QL lineage, and expression of mab-5 is necessary and sufficient for posterior migration. lin-39/Sex combs reduced/Hox is activated through unknown mechanisms in QR and promotes anterior migration of that lineage. lin-39 and mab-5 are well studied genes than have been implicated in cell-autonomous control of Q migrations. After an introduction to Q cell migration in chapter I, chapter II describes new roles for Hox gene in Q descendant migration that may change the way we think about how Hox genes work. I show that lin-39, mab-5 and a third Hox gene egl-5/Abdominal-B/Hox act in parallel to promote migration of the Q cells, and in their absence almost no migration occurs. This in contrast to the typical opposing roles of Hox genes whereby lin-39 promotes anterior migration of QR lineage, and mab-5 promotes posterior migration of the QL lineage. I also find that mab-5, and egl-5 are able to promote Q migration through their expression in posterior body wall muscles, a non-autonomous role in migration. Again this shifts how Hox genes should be considered, as they are often thought to act solely as autonomous drivers of cell fate and to have opposing roles in differentiation. As a Hox factor mab-5 regulates many genes, but few mab-5 targets have been identified. Results in this dissertation show that mab-5 may regulate the secreted F-spondin homolog spon-1 to control migration. I demonstrate that the spon-1 promoter can be driven by MAB-5 in the body wall muscles, and that spon-1 is important in Q cell direction and extent of migration. Chapter III presents new roles for egl-20/Wnt and mab-5/Hox in inhibiting anterior migration. Detailed analysis of the timing of Q descendant migration reveals that egl-20/Wnt can act in two steps to inhibit anterior migration and promote posterior migration of the QL linage. (i) Through an acute non-canonical Wnt mechanism EGL-20 inhibits anterior migration, and (ii) later, through canonical Wnt pathway activates transcription of mab-5 which can further inhibit anterior migration and promote further migration of the QL lineage. The first characterization of the C. elegans Neurofibromatosis Type II(NF2)/Merlin homolog nfm-1 is detailed in chapter IV. Previously in the Lundquist lab an unbiased forward genetic screen identified an allele of nfm-1 is required for Q descendant migration. I show that nfm-1 promotes migration of QR and QL lineages through non-autonomous mechanisms, and can act genetically, possibly in the same pathway with the guidance cue slt-1/Slit, to promote complete migration of the QR and QL lineages. The results contained below use new techniques to study established and novel genes in nervous system development, highlighting the intricacies of developmental genetics. Many genes play multiple roles in development, demonstrated here by new roles for Hox and Wnt in Q cell migration

EGL-20/Wnt and MAB-5/Hox Act Sequentially to Inhibit Anterior Migration of Neuroblasts in C. elegans

Directed neuroblast and neuronal migration is important in the proper development of nervous systems. In C. elegans the bilateral Q neuroblasts QR (on the right) and QL (on the left) undergo an identical pattern of cell division and differentiation but migrate in opposite directions (QR and descendants anteriorly and QL and descendants posteriorly). EGL-20/ Wnt, via canonical Wnt signaling, drives the expression of MAB-5/Hox in QL but not QR. MAB-5 acts as a determinant of posterior migration, and mab-5 and egl-20 mutants display anterior QL descendant migrations. Here we analyze the behaviors of QR and QL descendants as they begin their anterior and posterior migrations, and the effects of EGL-20 and MAB-5 on these behaviors. The anterior and posterior daughters of QR (QR.a/p) after the first division immediately polarize and begin anterior migration, whereas QL.a/p remain rounded and non-migratory. After ~1 hour, QL.a migrates posteriorly over QL.p. We find that in egl-20/Wnt, bar-1/β-catenin, and mab-5/Hox mutants, QL.a/p polarize and migrate anteriorly, indicating that these molecules normally inhibit anterior migration of QL.a/p. In egl-20/Wnt mutants, QL.a/p immediately polarize and begin migration, whereas in bar-1/β- catenin and mab-5/Hox, the cells transiently retain a rounded, non-migratory morphology before anterior migration. Thus, EGL-20/Wnt mediates an acute inhibition of anterior migration independently of BAR-1/β-catenin and MAB-5/Hox, and a later, possible transcriptional response mediated by BAR-1/β-catenin and MAB-5/Hox. In addition to inhibiting anterior migration, MAB-5/Hox also cell-autonomously promotes posterior migration of QL.a (and QR.a in a mab-5 gain-of-function)

Universal point contact resistance between thin-film superconductors

A system comprising two superconducting thin films connected by a point contact is considered. The contact resistance is calculated as a function of temperature and film geometry, and is found to vanish rapidly with temperature, according to a universal, nearly activated form, becoming strictly zero only at zero temperature. At the lowest temperatures, the activation barrier is set primarily by the superfluid stiffness in the films, and displays only a weak (i.e., logarithmic) temperature dependence. The Josephson effect is thus destroyed, albeit only weakly, as a consequence of the power-law-correlated superconducting fluctuations present in the films below the Berezinskii-Kosterlitz-Thouless transition temperature. The behavior of the resistance is discussed, both in various limiting regimes and as it crosses over between these regimes. Details are presented of a minimal model of the films and the contact, and of the calculation of the resistance. A formulation in terms of quantum phase-slip events is employed, which is natural and effective in the limit of a good contact. However, it is also shown to be effective even when the contact is poor and is, indeed, indispensable, as the system always behaves as if it were in the good-contact limit at low enough temperature. A simple mechanical analogy is introduced to provide some heuristic understanding of the nearly-activated temperature dependence of the resistance. Prospects for experimental tests of the predicted behavior are discussed, and numerical estimates relevant to anticipated experimental settings are provided.Comment: 29 pages (single column format), 7 figure

Heterogeneous histories of recombination suppression on stickleback sex chromosomes

How consistent are the evolutionary trajectories of sex chromosomes shortly after they form? Insights into the evolution of recombination, differentiation, and degeneration can be provided by comparing closely related species with homologous sex chromosomes. The sex chromosomes of the threespine stickleback (Gasterosteus aculeatus) and its sister species, the Japan Sea stickleback (G. nipponicus), have been well characterized. Little is known, however, about the sex chromosomes of their congener, the blackspotted stickleback (G. wheatlandi). We used pedigrees to obtain experimentally phased whole genome sequences from blackspotted stickleback X and Y chromosomes. Using multispecies gene trees and analysis of shared duplications, we demonstrate that Chromosome 19 is the ancestral sex chromosome and that its oldest stratum evolved in the common ancestor of the genus. After the blackspotted lineage diverged, its sex chromosomes experienced independent and more extensive recombination suppression, greater X–Y differentiation, and a much higher rate of Y degeneration than the other two species. These patterns may result from a smaller effective population size in the blackspotted stickleback. A recent fusion between the ancestral blackspotted stickleback Y chromosome and Chromosome 12, which produced a neo-X and neo-Y, may have been favored by the very small size of the recombining region on the ancestral sex chromosome. We identify six strata on the ancestral and neo-sex chromosomes where recombination between the X and Y ceased at different times. These results confirm that sex chromosomes can evolve large differences within and between species over short evolutionary timescales

Searching for signatures of sexually antagonistic selection on stickleback sex chromosomes.

Intralocus sexually antagonistic selection occurs when an allele is beneficial to one sex but detrimental to the other. This form of selection is thought to be key to the evolution of sex chromosomes but is hard to detect. Here we perform an analysis of phased young sex chromosomes to look for signals of sexually antagonistic selection in the Japan Sea stickleback (Gasterosteus nipponicus). Phasing allows us to date the suppression of recombination on the sex chromosome and provides unprecedented resolution to identify sexually antagonistic selection in the recombining region of the chromosome. We identify four windows with elevated divergence between the X and Y in the recombining region, all in or very near genes associated with phenotypes potentially under sexually antagonistic selection in humans. We are unable, however, to rule out the alternative hypothesis that the peaks of divergence result from demographic effects. Thus, although sexually antagonistic selection is a key hypothesis for the formation of supergenes on sex chromosomes, it remains challenging to detect. This article is part of the theme issue 'Genomic architecture of supergenes: causes and evolutionary consequences'

Fate of the Josephson effect in thin-film superconductors

The dc Josephson effect refers to the dissipationless electrical current -- the supercurrent -- that can be sustained across a weak link connecting two bulk superconductors. This effect is a probe of the fundamental nature of the superconducting state. Here, we analyze the case of two superconducting thin films connected by a point contact. Remarkably, the Josephson effect is absent at nonzero temperature, and the resistance across the contact is nonzero. Moreover, the point contact resistance is found to vary with temperature in a nearly activated fashion, with a UNIVERSAL energy barrier determined only by the superfluid stiffness characterizing the films, an angle characterizing the geometry, and whether or not the Coulomb interaction between Cooper pairs is screened. This behavior reflects the subtle nature of the superconductivity in two-dimensional thin films, and should be testable in detail by future experiments.Comment: 16 + 8 pages. 1 figure, 1 tabl

Vortex Collisions: Crossing or Recombination?

We investigate the collision of two vortex lines moving with viscous dynamics and driven towards each other by an applied current. Using London theory in the approach phase we observe a non-trivial vortex conformation producing anti-parallel segments; their attractive interaction triggers a violent collision. The collision region is analyzed using the time-dependent Ginzburg-Landau equation. While we find vortices will always recombine through exchange of segments, a crossing channel appears naturally through a double collision process.Comment: 4 pages, 3 figure