1 / 17

Structure, function, and temporal expression of Fibrillin in Danio rerio.

Structure, function, and temporal expression of Fibrillin in Danio rerio. Christian Francom, Jeffrey Caldwell, Barbara Trask, Ph.D. Extracellular Matrix. Functions Structure—tense or elastic Cellular Adhesion Cellular Motility Molecular Reservoir. Elastic Extracellular Matrices.

marja
Télécharger la présentation

Structure, function, and temporal expression of Fibrillin in Danio rerio.

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Structure, function, and temporal expression of Fibrillin in Danio rerio. Christian Francom, Jeffrey Caldwell, Barbara Trask, Ph.D.

  2. Extracellular Matrix Functions • Structure—tense or elastic • Cellular Adhesion • Cellular Motility • Molecular Reservoir

  3. Elastic Extracellular Matrices • Component parts • Elastin • Microfibrils • Fibrillins • MAGPs • Fibulins • Other proteins and proteoglycans • Disorders involving the Elastic ECM • Elastin SVAS • MicrofibrilsMarfan Syndrome

  4. Question • How do the microfibrillar proteins contribute to the function of the elastic extracellular matrix? • When and where is each expressed? • What inter-molecular interactions are maintained?

  5. Model Organism • Zebrafish (Daniorerio) • External fertilization and development • Colorless during the first few days of development • Genome sequenced (although not assembled) • **first evolutionary organism to have functional elastic extracellular matrix • Easily maintained http://www.pdn.cam.ac.uk/staff/lewis/zfish3AgesSlide.jpg

  6. Stages of Z-fish Development

  7. Sequences and Primers • Fibrillin atggcagcctgtcctgaattaatcttggaaagagagggaggagaaggagtggatgggggattctttaaggcgaagatgactaaggatttggtggaacgggagattcgcatcacagatgtgcagccggcaaatctgcagcaactgcgtgatgctgtcatctcaaaattggccaaaatctctgaggaatatttccagtaccttgttgaatctttgccacaaagaattgaggcatttgtgaaagcaaaaaggatgtccaacctggagggagccccaggctccaggatattctcagctcaggatgtgaacgagtgcaacgaccccaacacctgcatcagcggtctgtgtctgaacacccctggcagctacatctgcagctgcccccccgactttgagctcaatcccactggagtcggctgcgtcgacactcgctctggaaactgctacctggacgtccagtaccgaggcgatttctctgagagtctgagctgctccaatgagattggagtgggagtgtccaaagcgtcctgctgctgttctaggggtgtggcctggggacagccctgcgaggcctgtccatcactcaactccaagatctgtttcactgcacaacctgaaatctccagtctggtgcagggagagaccagccagtgcttccccaggaggagtatccagtgctgggacacattagcggctcattcccttgggcaagcctggagctcagatggccagctgcacgtttccactcttcatctttgttgtggtgtgaaagaggagaaggagggtttaggaaaacaaaaaagcagtggacagagtgatgttacaaagacaacaggcttcacacagagctctgacagtcgcgtcaaagcgagtgcgctgttattaaaatggtaccgtcccagtgacagctctcacagctttatttttgcagtgtcaggactacggtccaatcatgcaatagtagaggacatccgtctgttcaacagtaagcaatatggagctgagccgctgaaagaactgctcaaactactaccagaagcagatgaggtgaagagattgagagaattcaaaggagacgcaaataagctgacactggttgattccttcatgttccagctcattcaagtaccacgctttgatgtgcgtatcgaggtcatggtacttcaagaagagtttttcccattatgtactgccatgagtcgtgacatcaacattgtgcgccaggcaactgaagctgtgcctgtattagccttctctaagaaaccaatggctcatgaccttttagtcctgacaggtggctatgcaggaaacgctgtaggcttcaaattgtcctctcttctctctctggctgacacaaaggccaacaagcctggcatgaacctgctccattttgtggcattactctcgaggatcttctctgtgttttcacagggtggctatgcaggaaacgctgtaggcttcaaattgtcctctcttctctctctggctgacacaaaggccaacaagcctggcatgaacctgctccattttgtggcattagaagcacagaaaaaggatgaggcacttttaaagttcccagagaagctaacacatattcagagtgcagccagaatttcagtagaaaatatagaaggagagttttcctcactctatgtgaaaacacggtctttggaacagaagattcaagatgacgatgagctgcaaaagcagctagatccatttttgcagagctctgcaaaagccttgcaggaccttaagcaccgcaggctggacttgcgcaaagagggtaatgccctcattgacttcttctgtgaagacaaagacaccttcaaattggatgagtgcttcagaatcttccaagatttctgtctaaagttcaaaaaagctgtgaaggacaactcggacagagaactgaaagaggaagctagacagcgccgcttacgggaactagaggagagacggtgtgcctggggtgcagccaatattgaaacaggtggtaatggaggttttggtcgcagcagcagtgaaaatgatgttgacatcttaaccaaagatggcctcctagacttcctgcttcaatcaagacctcaaagcccccacagtcctttaggacgctcagctagtgctcgccgctaccaccacacatctagtgaccgcagcctccgtggctatatggagctggtctctgggtcttcagcaagtgaatacgccaaattcagcagcctgccacgttcaagcaggaaccatcaaagaaagaccatgccatggcttgtcttgcaggatgataatagagagttgattccacaaagtcaggtttaccaagagaagttagcagtttcacaaagggttgagactgaaccaatcagtcctttggcaagatactcaaatttaggctacaacttgaacagtgatccctatagtaataataataacaattatagttctctttctgaaggtggtcttacacctcgattaaataaaacaaagaatgtctttcaggggacatctggtcaaggcattggacacatgaatgtttctgttgagaaacgcaaactagtgcctgggcttcaaccctttgagataaccagccagatcaataataacagtcatgtttctttggacaatcaacatgaagtgtgtgtgactgatctggaacgagaagaccttcccaaagcttttgttcttgatacaccaccatctagtagaagttctgaggaggggcataagctagaaaaagtatggaatgacaaaatcatctcatcacagagagacgaagcagacacaagtaccttctcttcaacatcttgtgacaccccattgccccttgatccctccagtaaaaaacctgtgttttatgtcatggactgcactgaaacggattgttccattgtgttggactgttctgagattgagagttctcctattataaaagaaggacctggacttatcccaaagttacaggatgtcaactttccaagaatcattcaagacaccaattctctttcatccaattttgagtcttcagactgtcactttgtgtccacatcaagtgatgaaccatttgtaggaagggtcgatcaaacttcagaatcagtcagtccatctgtaacaactgaagaagcagagacagacagctgtgacacagctgaagggcaaagagttgatgagaaaggtgttcagacaggtagccccaaaacgacacttatcaaaaccaagacttcatccaaagggatgactgctcacaccagccgtcctacaagaaccctcacagctactgagactcagaacatgcgaaaagtagtgcccattacaaggcaaaatcgatctagtagcagcatgagaaaggttgataaaccacctgtacatgagagctctgaacccagacgtccatttcgagaccaaagcatgccagccagaagaggcgagagacaaggtagacctacaagacattcaagccttccaccagaggatcctaggacaccaagaggttcgacaccaagcagctctcgttggggaagggactcaattcaacacaggccctccatcaagaaagccaatgccaaacctgttcgtaacattcctaaaccagcaccagaggaaaagatgtgtcgctctacaatgcgtgccttggctcaagctcaggctcaaggaacctcagaaggaggtaccgctgacagccctaacaccaccccaaaaagttcttcacctttgcctagctttgctcgcaacactgtggcatctacttctagatctaaaaaagacatgccatcagcaactgtaaccggaacaccatcaaagagcctatcgagaactagctctcagagattgcctggtggtagaacagagtctgtcagtagcgggcctttaccaaggagtgaggataaactaggaggctcgattaggagggttcaaagtgtaagagcttccagtcgaagcagtgacacaccttcacaatctggtcgggatcactcccttaaaagtagcagtttctcagacagatccacaccaactagagacaccatcatctccagcaggaccagcaaacccacatggaaatag • MAGP atgagactctgttacctgcttctgctgtcactgccgggtctgctggctcaggtgcagaactatgatgaggattatacagacatcttcggtaacagcaacaactacgacaaccccttcttctaccagcagcagatattacagacgcagacgtcagggcctgaacagaacgcgctggatggacagacagagccgactgaacctggaccgctggactgtcgtgaggagcagtatccctgcactcggctctattctgtccatcagccgtgtaaacagtgtctgaacagcatctgcttctacagtctgcgcaggatgtatgtgatcaataaggaggtgtgtgtgcggacggtgtgtgcacatgaggagctcttgagagcggatctgtgtcgtgaccagttcccccgctgtggtgtaatggcgcacattggccagtgtggggcgatggggagcaactgcgccaggagttgtggatcctgctaa

  8. Expression Data Fibrillin MAGP MAGP GAPDH Control

  9. Future Possibilities • More replicates should be done • Analysis with Real-Time PCR • Alternative primers should be made • In situ hybridization should localize expression

  10. Question • How do the microfibrillar proteins contribute to the function of the elastic extracellular matrix? • When and where is each expressed? • What inter-molecular interactions are maintained?

  11. Intermolecular interactions Fibrillin interacts with: MAGP interacts with: • MAGP • CSPG • LTGF-β • Others? • Fibrillins • CSPG • Elastin • Others?

  12. Mammalian Structure of Fibrillin-1

  13. Isolated Bands of LTGF-β Domains 3 5 6 300bp 200bp 100bp

  14. LTGF-β Binding Domains ….MAACPELILEREGGEGVDGGFFKAKMTKDLVEREIRITDVQPANLQQLRDAVISKLAKISEEYFQYLVESLPQRIEAFVKAKRMSNLEGAPGSRIFSAQDVNECNDPNTCISGLCLNTPGSYICSCPPDFELNPTGVGCVDTRSGNCYLDVQYRGDFSESLSCSNEIGVGVSKASCCCSRGVAWGQPCEACPSLNSKICFTAQPEISSLVQGETSQCFPRRSIQCWDTLAAHSLGQAWSSDGQLHVSTLHLCCGVKEEKEGLGKQKSSGQSDVTKTTGFTQSSDSRVKASALLLKWYRPSDSSHSFIFAVSGLRSNHAIVEDIRLFNSKQYGAEPLKELLKLLPEADEVKRLREFKGDANKLTLVDSFMFQLIQVPRFDVRIEVMVLQEEFFPLCTAMSRDINIVRQATEAVPVLAFSKKPMAHDLLVLTGGYAGNAVGFKLSSLLSLADTKANKPGMNLLHFVALLSRIFSVFSQGGYAGNAVGFKLSSLLSLADTKANKPGMNLLHFVALEAQKKDEALLKFPEKLTHIQSAARISVENIEGEFSSLYVKTRSLEQKIQDDDELQKQLDPFLQSSAKALQDLKHRRLDLRKEGNALIDFFCEDKDTFKLDECFRIFQDFCLKFKKAVKDNSDRELKEEARQRRLRELEERRCAWGAANIETGGNGGFGRSSSENDVDILTKDGLLDFLLQSRPQSPHSPLGRSASARRYHHTSSDRSLRGYMELVSGSSASEYAKFSSLPRSSRNHQRKTMPWLVLQDDNRELIPQSQVYQEKLAVSQRVETEPISPLARYSNLGYNLNSDPYSNNNNNYSSLSEGGLTPRLNKTKNVFQGTSGQGIGHMNVSVEKRKLVPGLQPFEITSQINNNSHVSLDNQHEVCVTDLEREDLPKAFVLDTPPSSRSSEEGHKLEKVWNDKIISSQRDEADTSTFSSTSCDTPLPLDPSSKKPVFYVMDCTETDCSIVLDCSEIESSPIIKEGPGLIPKLQDVNFPRIIQDTNSLSSNFESSDCHFVSTSSDEPFVGRVDQTSESVSPSVTTEEAETDSCDTAEGQRVDEKGVQTGSPKTTLIKTKTSSKGMTAHTSRPTRTLTATETQNMRKVVPITRQNRSSSSMRKVDKPPVHESSEPRRPFRDQSMPARRGERQGRPTRHSSLPPEDPRTPRGSTPSSSRWGRDSIQHRPSIKKANAKPVRNIPKPAPEEKMCRSTMRALAQAQAQGTSEGGTADSPNTTPKSSSPLPSFARNTVASTSRSKKDMPSATVTGTPSKSLSRTSSQRLPGGRTESVSSGPLPRSEDKLGGSIRRVQSVRASSRSSDTPSQSGRDHSLKSSSFSDRSTPTRDTIISSRTSKPTWK

  15. LTGF-β Binding Domains ….MAACPELILEREGGEGVDGGFFKAKMTKDLVEREIRITDVQPANLQQLRDAVISKLAKISEEYFQYLVESLPQRIEAFVKAKRMSNLEGAPGSRIFSAQDVNECNDPNTCISGLCLNTPGSYICSCPPDFELNPTGVGCVDTRSGNCYLDVQYRGDFSESLSCSNEIGVGVSKASCCCSRGVAWGQPCEACPSLNSKICFTAQPEISSLVQGETSQCFPRRSIQCWDTLAAHSLGQAWSSDGQLHVSTLHLCCGVKEEKEGLGKQKSSGQSDVTKTTGFTQSSDSRVKASALLLKWYRPSDSSHSFIFAVSGLRSNHAIVEDIRLFNSKQYGAEPLKELLKLLPEADEVKRLREFKGDANKLTLVDSFMFQLIQVPRFDVRIEVMVLQEEFFPLCTAMSRDINIVRQATEAVPVLAFSKKPMAHDLLVLTGGYAGNAVGFKLSSLLSLADTKANKPGMNLLHFVALLSRIFSVFSQGGYAGNAVGFKLSSLLSLADTKANKPGMNLLHFVALEAQKKDEALLKFPEKLTHIQSAARISVENIEGEFSSLYVKTRSLEQKIQDDDELQKQLDPFLQSSAKALQDLKHRRLDLRKEGNALIDFFCEDKDTFKLDECFRIFQDFCLKFKKAVKDNSDRELKEEARQRRLRELEERRCAWGAANIETGGNGGFGRSSSENDVDILTKDGLLDFLLQSRPQSPHSPLGRSASARRYHHTSSDRSLRGYMELVSGSSASEYAKFSSLPRSSRNHQRKTMPWLVLQDDNRELIPQSQVYQEKLAVSQRVETEPISPLARYSNLGYNLNSDPYSNNNNNYSSLSEGGLTPRLNKTKNVFQGTSGQGIGHMNVSVEKRKLVPGLQPFEITSQINNNSHVSLDNQHEVCVTDLEREDLPKAFVLDTPPSSRSSEEGHKLEKVWNDKIISSQRDEADTSTFSSTSCDTPLPLDPSSKKPVFYVMDCTETDCSIVLDCSEIESSPIIKEGPGLIPKLQDVNFPRIIQDTNSLSSNFESSDCHFVSTSSDEPFVGRVDQTSESVSPSVTTEEAETDSCDTAEGQRVDEKGVQTGSPKTTLIKTKTSSKGMTAHTSRPTRTLTATETQNMRKVVPITRQNRSSSSMRKVDKPPVHESSEPRRPFRDQSMPARRGERQGRPTRHSSLPPEDPRTPRGSTPSSSRWGRDSIQHRPSIKKANAKPVRNIPKPAPEEKMCRSTMRALAQAQAQGTSEGGTADSPNTTPKSSSPLPSFARNTVASTSRSKKDMPSATVTGTPSKSLSRTSSQRLPGGRTESVSSGPLPRSEDKLGGSIRRVQSVRASSRSSDTPSQSGRDHSLKSSSFSDRSTPTRDTIISSRTSKPTWK

  16. Future Directions • Clone for expression • Protein binding studies

  17. Acknowledgements • Josh McBride • Aric Elmer • OUR

More Related