Biology subjects

Wallingford, J. B.

Publications and source records attributed to Wallingford, J. B..

5 recordsLinked to original sources

bioRxiv2018

Mutations in Kinesin Family Member 6 Reveal Specific Role in Ependymal Cell Function and Human Neuro-Cranial Development

Cerebrospinal fluid flow is crucial for neurodevelopment and homeostasis of the ventricular system of the brain, with localized flow being established by the polarized beating of the ependymal cell (EC) cilia. Here, we report a homozygous one base-pair deletion, c.1193delT (p.Leu398Glnfs*2), in the Kinesin Family Member 6 (KIF6) gene in a child displaying neurocranial defects and intellectual disability. To test the pathogenicity of this novel human KIF6 mutation we engineered an analogous C-terminal truncating mutation in mouse. These mutant mice display severe, postnatal-onset hydrocephalus. We generated a Kif6-LacZ transgenic mouse strain and report expression specifically and uniquely within the ependymal cell (EC) layer of the brain, without labeling other multiciliated mouse tissues. Analysis of Kif6 mutant mice with scanning electron microscopy (SEM) and immunofluorescence (IF) revealed a reduction in EC cilia, without effect on other multiciliated tissues. Consistent with our findings in mice, defects of the ventricular system and EC cilia were observed in kif6 mutant zebrafish. Overall, this work describes the first clinically-defined KIF6 homozygous null mutation in human and defines KIF6 as a conserved mediator of neuro-cranial morphogenesis with a specific role in the maintenance of EC cilia in vertebrates.\n\nAUTHOR SUMMARYCerebrospinal fluid flow is crucial for neurodevelopment and homeostasis of the ventricular system of the brain. Localized flows of cerebrospinal fluid throughout the ventricular system of the brain are established from the polarized beating of the ependymal cell (EC) cilia. Here, we identified a homozygous truncating mutation in KIF6 in a child displaying neuro-cranial defects and intellectual disability. To test the function of KIF6 in vivo, we engineered mutations of Kif6 in mouse. These Kif6 mutant mice display severe hydrocephalus, coupled with a loss of EC cilia. Similarly, we observed hydrocephalus and a reduction in EC cilia in kif6 mutant zebrafish. Overall, this work describes the first clinically-defined KIF6 mutation in human, while our animal studies demonstrate the pathogenicity of mutations in KIF6 and establish KIF6 as a conserved mediator of neuro-cranial development and EC cilia maintenance in vertebrates.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC=\"FIGDIR/small/350140_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (58K):\norg.highwire.dtl.DTLVardef@d36bc6org.highwire.dtl.DTLVardef@48cbfborg.highwire.dtl.DTLVardef@147e273org.highwire.dtl.DTLVardef@641ce3_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2018

Frequency and synchrony of actomyosin oscillation during PCP-dependent convergent extension

Oscillatory actomyosin flows play a key role in single cell migration and in collective cell movements that shape invertebrates embryos, but the role of such oscillations in vertebrate morphogenesis remains poorly defined. Here, data from mathematical modeling and in vivo 4D imaging of actomyosin in the Xenopus gastrula suggest that oscillatory actomyosin contractions are a general feature of convergent extension by junction shrinking. We show that synchronous intracellular flows link two spatially distinct populations of actomyosin within individual cells, but that oscillations are asynchronous between neighboring cells that share a shrinking cell-cell junction. We also show that the core PCP protein Prickle2 displays a parallel oscillatory behavior and is required for tuning the frequency of actomyosin contractions, indicating that PCP signaling controls not only the orientation of actomyosin contractions, but also their frequency. Together, these data provide new insights into the function and control of oscillatory actomyosin contractions in collective cell movement.

developmental biology↗

bioRxiv2018

Spatial and temporal PCP protein dynamics coordinate cell intercalation during neural tube closure

Planar cell polarity (PCP) controls the convergent extension cell movements that drive axis elongation in all vertebrates. Though asymmetric localization of core PCP proteins is central to their function, we currently understand little about PCP protein localization as it relates to the subcellular behaviors that drive convergent extension. Here, we have used high magnification time-lapse imaging to simultaneously monitor cell intercalation behaviors and the localization of the PCP proteins Prickle2 and Vangl2. We observed the expected asymmetric enrichment of PCP proteins, but more interestingly, we also observed tight temporal and spatial correlation of PCP protein enrichment with contractile behavior in cell-cell junctions. These patterns of localization were associated with similar pattern of protein turnover at junctions as assessed by FRAP. In fact, dynamic enrichment of PCP proteins was linked more strongly to junction behavior than to spatial orientation. Finally, recruitment of Prickle2 and Vangl2 to cell-cell junctions was temporally and spatially coordinated with planar polarized oscillations of actomyosin enrichment, and all of these dynamic relationships were disrupted when PCP signaling was manipulated. Together, these results provide a dynamic and quantitative view of PCP protein localization during convergent extension and suggest a complex and intimate link between the dynamic localization of core PCP proteins, actomyosin assembly, and polarized junction shrinking during cell intercalation of the closing vertebrate neural tube.

developmental biology↗

bioRxiv2017

Evolutionary proteomics uncovers ciliary signaling components

Cilia are organelles specialized for movement and signaling. To infer when during animal evolution signaling pathways became associated with cilia, we characterized the proteomes of cilia from three organisms: sea urchins, sea anemones and choanoflagellates. From these ciliomes, we identified 437 high confidence ciliary candidate proteins conserved in mammals, including known regulators of Hh, GPCR and TRP channel signaling. The phylogenetic profiles of their ciliary association indicate that the Hh and GPCR pathways were linked to cilia before the origin of bilateria and TRP channels before the origin of animals. We demonstrated that some of the candidates not previously implicated in ciliary biology localized to cilia and further investigated ENKUR, a TRP channel-interacting protein that we identified in the cilia of all three organisms. In animals, ENKUR is expressed by cells with motile cilia, ENKUR localizes to cilia in diverse organisms and, in both Xenopus laevis and mice, ENKUR is required for patterning the left/right axis. Moreover, mutation of ENKUR causes situs inversus in humans. Thus, proteomic profiling of cilia from diverse eukaryotes defines a conserved ciliary proteome, reveals ancient connections to Hh, GPCR and TRP channel signaling, and uncovers a novel ciliary protein that controls vertebrate development and human disease.

evolutionary biology↗

bioRxiv2016

A synthesis of over 9,000 mass spectrometry experiments reveals the core set of human protein complexes

Macromolecular protein complexes carry out many of the essential functions of cells, and many genetic diseases arise from disrupting the functions of such complexes. Currently there is great interest in defining the complete set of human protein complexes, but recent published maps lack comprehensive coverage. Here, through the synthesis of over 9,000 published mass spectrometry experiments, we present hu.MAP, the most comprehensive and accurate human protein complex map to date, containing >4,600 total complexes, >7,700 proteins and >56,000 unique interactions, including thousands of confident protein interactions not identified by the original publications. hu.MAP accurately recapitulates known complexes withheld from the learning procedure, which was optimized with the aid of a new quantitative metric (k-cliques) for comparing sets of sets. The vast majority of complexes in our map are significantly enriched with literature annotations and the map overall shows improved coverage of many disease-associated proteins, as we describe in detail for ciliopathies. Using hu.MAP, we predicted and experimentally validated candidate ciliopathy disease genes in vivo in a model vertebrate, discovering CCDC138, WDR90, and KIAA1328 to be new cilia basal body/centriolar satellite proteins, and identifying ANKRD55 as a novel member of the intraflagellar transport machinery. By offering significant improvements to the accuracy and coverage of human protein complexes, hu.MAP (http://proteincomplexes.org) serves as a valuable resource for better understanding the core cellular functions of human proteins and helping to determine mechanistic foundations of human disease.

systems biology↗