Molecular Mechanisms Underlying Cockroach Host Manipulation by a Parasitoid Wasp

Molecular Mechanisms Underlying Cockroach Host Manipulation by a Parasitoid Wasp Thesis submitted in partial fulfillment of the requirements for the degree of “DOCTOR OF PHILOSOPHY” by <ul style="display: flex;"><li style="flex:1">Maayan </li><li style="flex:1">Kaiser Paltin </li></ul>Submitted to the Senate of Ben-Gurion University of the Negev June 2018 Beer-Sheva Molecular Mechanisms Underlying Cockroach Host Manipulation by a Parasitoid Wasp Thesis submitted in partial fulfillment of the requirements for the degree of “DOCTOR OF PHILOSOPHY” by <ul style="display: flex;"><li style="flex:1">Maayan </li><li style="flex:1">Kaiser Paltin </li></ul>Submitted to the Senate of Ben-Gurion University of the Negev 30/12/2018 Approved by the advisor Approved by the Dean of the Kreitman School of Advanced Graduate Studies June 2018 Beer-Sheva This work was carried out under the supervision of Prof. Frederic Libersat In the Department of Life Sciences Faculty of Natural Sciences Research-Student's Affidavit when Submitting the Doctoral Thesis for Judgment I, Maayan Kaiser Paltin, whose signature appears below, hereby declare that: ___ I have written this Thesis by myself, except for the help and guidance offered by my Thesis Advisors. ___ The scientific materials included in this Thesis are products of my own research, culled from the period during which I was a research student. ___ This Thesis incorporates research materials produced in cooperation with others, excluding the technical help commonly received during experimental work. Therefore, I am attaching another affidavit stating the contributions made by myself and the other participants in this research, which has been approved by them and submitted with their approval. 31/12/18 Date: _________________ Maayan Kaiser Paltin Student's name: ________________ Signature:______________ Affidavit stating the contributions to present work: The venom proteomics which is described at chapter 2, was performed in collaboration with Prof. Michael Adams and Dr. Ryan Arvidson from the University of California, Riverside. Mass spectrometry analysis was based on the transcriptomes and differential expression analysis of the venom apparatus, which were done there (described at method section, chapter 2). In chapter 3, in collaboration with Prof. Michael Adams, I used the transcriptome of the cockroach cerebral ganglia as a database for mass spectrometry analysis. Acknowledgments First, I would like to express my utmost gratitude to my advisor and mentor, Prof. Frederic Libersat, for giving me the opportunity to work in his lab, for having the patient when it was needed, for teaching me the basics of being a scientist, and for his wise advices and help in all matters. I would like to thanks some people who led me to the decision of pursuing science. First and most important are my parents, Rene and Leonardo Kaiser, who I have so much to thank for, but especially on always insisting on the importance of expanding own knowledge. My parents inhered me the love for the written word, such an important thing a parent can give to his child. This introduced me to Daglas Adams and Richard Dawkins who inspired me to pursuit biology. I thank those great persons for leading my way to where I needed to be. Thanks to my other and better half, my favored person in the whole universe, Daniel. I could not have done a thing without him, as support and inspiration. Thanks to my brothers Noam and Ariel. Noam, the best person I know, for teaching me so much, and Ariel, for always taking care of me. Thanks to my parents at law, Veronika and Uri Paltin, for being the amazing people they are, for their precious and wise advices, for their kindness and help in anything I could possibly need. I would like to thank Gustavo Glusman, who made the lab more like a home, for his kindness and help. My gratitude goes also to Ram Gal, who mentored me at the beginning of my way and Stav Emanuel, for the companionship and moral support. I would like to thank many more from our life sciences department: Prof. Raz Zarivach and Nitzan Kutnowski for their help with the venom affinity chromatography, Prof. Noam Zilberberg and Prof. Uri Abdu for their wise advices and comments and to all members of Prof. Ofer Yifrach and Prof. Noam Zilberberg labs, for their kind help. Special thanks to Prof. Michael Adams for the collaboration, helpful comments and editing part of this thesis and to Dr. Ryan Arvidson for his collaboration on the venomics of the wasp and for his work on the transcriptomic of the cockroach cerebral ganglia. To Lia Table of Contents <a href="#8_0">List of figures and tables </a><a href="#8_0">..................................................................................................</a><a href="#8_0">.</a><a href="#8_0">1 </a><a href="#12_0">Abstract </a><a href="#12_0">..........................................................................................................................</a><a href="#12_0">.</a><a href="#12_0">4 </a><a href="#14_0">General Introductio</a><a href="#14_0">n</a><a href="#14_0">........................................................................................................</a><a href="#14_0">.</a><a href="#14_0">6 </a><a href="#18_0">1.The role of the cerebral ganglia in the venom-induced behavioral manipulation of </a><a href="#18_0">cockroaches stung by the parasitoid Jewel Wasp</a><a href="#18_0">............................................................... </a><a href="#18_0">10 </a> <a href="#18_1">1.1 Abstrac</a><a href="#18_1">t</a><a href="#18_1">................................................................................................................ </a><a href="#18_1">10 </a><a href="#18_2">1.2 Introduction </a><a href="#18_2">.......................................................................................................... </a><a href="#18_2">10 </a><a href="#0_0">1.3 Material and Method</a><a href="#0_0">s</a><a href="#0_0">............................................................................................ </a><a href="#0_0">13 </a><a href="#0_1">1.4 Results</a><a href="#0_1">.................................................................................................................. </a><a href="#0_1">16 </a><a href="#0_2">1.5 Discussion </a><a href="#0_2">............................................................................................................ </a><a href="#0_2">22 </a><a href="#0_3">2. Parasitoid Jewel Wasp Mounts Multi-Pronged Neurochemical Attack to Hijack a Host Brain </a><a href="#0_3">...................................................................................................................................... </a><a href="#0_3">25 </a> <a href="#0_4">2.1 Abstrac</a><a href="#0_4">t</a><a href="#0_4">................................................................................................................ </a><a href="#0_4">25 </a><a href="#0_5">2.2 Introduction </a><a href="#0_5">.......................................................................................................... </a><a href="#0_5">25 </a><a href="#0_0">2.3 Material and Method</a><a href="#0_0">s</a><a href="#0_0">............................................................................................ </a><a href="#0_0">27 </a><a href="#0_3">2.4 Results</a><a href="#0_3">.................................................................................................................. </a><a href="#0_3">31 </a><a href="#0_6">1.5 Discussion </a><a href="#0_6">............................................................................................................ </a><a href="#0_6">39 </a><a href="#0_3">3. Molecular cross-talk in a unique parasitic manipulation strategy </a><a href="#0_3">.................................... </a><a href="#0_3">46 </a><a href="#0_7">3.1 Abstrac</a><a href="#0_7">t</a><a href="#0_7">................................................................................................................ </a><a href="#0_7">46 </a><a href="#0_8">3.2 Introduction </a><a href="#0_8">.......................................................................................................... </a><a href="#0_8">46 </a><a href="#0_3">3.3 Methods</a><a href="#0_3">................................................................................................................ </a><a href="#0_3">48 </a><a href="#0_9">3.3 Results</a><a href="#0_9">.................................................................................................................. </a><a href="#0_9">51 </a><a href="#0_3">3.4 Discussion </a><a href="#0_3">............................................................................................................ </a><a href="#0_3">71 </a> <a href="#0_3">4. General Discussion and Future Research Direction</a><a href="#0_3">s</a><a href="#0_3">...................................................... </a><a href="#0_3">80 </a><a href="#0_10">References </a><a href="#0_10">..................................................................................................................... </a><a href="#0_10">88 </a><a href="#0_3">Appendix 1 </a><a href="#0_3">.................................................................................................................... </a><a href="#0_3">99 </a><a href="#0_3">Appendix 2 </a><a href="#0_3">.................................................................................................................. </a><a href="#0_3">100 </a><a href="#0_3">Appendix 3 </a><a href="#0_3">.................................................................................................................. </a><a href="#0_3">101 </a><a href="#0_3">Appendix 4 </a><a href="#0_3">.................................................................................................................. </a><a href="#0_3">103 </a>List of figures and tables <a href="#15_0">Figure 1: </a><a href="#15_0">Life cycle of the Jewel Wasp.</a><a href="#15_0">........................................................................</a><a href="#15_0">7 </a><a href="#19_0">Figure 1.1</a><a href="#19_0">: The venom is injected directly into the cockroach cerebral ganglia.</a><a href="#19_0">.....</a><a href="#19_0">11 </a><a href="#0_11">Figure 1.2 </a><a href="#0_11">Brain exposure for the surgical procedures</a><a href="#0_11">.............................................</a><a href="#0_11">16 </a><a href="#0_12">Figure 1.3: </a><a href="#0_12">A procaine injection to the CX decreases spontaneous walking.</a><a href="#0_12">...........</a><a href="#0_12">17 </a><a href="#0_13">Figure 1.4</a><a href="#0_13">: Representative images for postmortem verification of injection site. </a><a href="#0_13">....</a><a href="#0_13">18 </a><a href="#0_14">Figure 1.5: </a><a href="#0_14">A procaine injection to the MBs increases spontaneous walking.</a><a href="#0_14">.........</a><a href="#0_14">19 </a><a href="#0_15">Figure 1.6. </a><a href="#0_15">A venom injection to the CX decreases spontaneous walking.</a><a href="#0_15">...............</a><a href="#0_15">20 </a><a href="#0_16">Figure 1.7: </a><a href="#0_16">Stung crushed CirC cockroaches show decreased spontaneous walking. </a><a href="#0_16">......................................................................................................................................</a><a href="#0_16">21 </a><a href="#0_17">Figure 2.1: </a><a href="#0_17">Proteins/peptides in the venom are necessary for the behavioral </a><a href="#0_17">manipulation. </a><a href="#0_17">..............................................................................................................</a><a href="#0_17">31 </a><a href="#0_18">Figure 2.2: </a><a href="#0_18">Morphological analysis of A. compressa venom apparatus</a><a href="#0_18">.</a><a href="#0_18">..................</a><a href="#0_18">32 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587762" target="_blank">Figure 2.3: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587762" target="_blank">Domains of venom proteins.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587762" target="_blank">....................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587762" target="_blank">33 </a><a href="#0_19">Figure 2.4: </a><a href="#0_19">Proteomic analysis of A. compressa venom.</a><a href="#0_19">...........................................</a><a href="#0_19">34 </a><a href="#0_20">Figure 2.5: </a><a href="#0_20">Partial list of A. compressa venom proteins with expression and </a><a href="#0_20">abundance values</a><a href="#0_20">.</a><a href="#0_20">.......................................................................................................</a><a href="#0_20">37 </a><a href="#0_21">Figure 2.6: </a><a href="#0_21">Bioinformatic analysis of the venom proteome </a><a href="#0_21">......................................</a><a href="#0_21">38 </a><a href="#0_22">Figure 2.7: </a><a href="#0_22">Comparative genomic analysis of A. compressa venom proteins. </a><a href="#0_22">.........</a><a href="#0_22">39 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">Figure.2.8: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">Schematic representation of venom proteins that could be localized at the </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">synspase</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">......................................................................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587767" target="_blank">45 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">Figure 3.1: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">Venom affinity column revealed multiple bands in a wide range of </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">molecular weights</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">. </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">......................................................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587768" target="_blank">52 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587769" target="_blank">Figure 3.2: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587769" target="_blank">The venom targets database is enriched with proteins that are associated </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587769" target="_blank">with synaptic processes. </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587769" target="_blank">..............................................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587769" target="_blank">54 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">Figure 3.3: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">The venom targets database is enriched with proteins that are associated </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">with synaptic processes</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">. </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">..............................................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587770" target="_blank">55 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587771" target="_blank">Figure 3.4</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587771" target="_blank">: Venom targets in brain show enriched terms that that are associated with </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587771" target="_blank">cytoskeleton organization and synapse assembly. </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587771" target="_blank">.....................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587771" target="_blank">56 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587772" target="_blank">Figure 3.5: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587772" target="_blank">Venom targets in SEG show enriched terms that are associated with </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587772" target="_blank">cytoskeleton organization and synapse assembly. </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587772" target="_blank">.....................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587772" target="_blank">57 </a><a href="#0_23">Figure 3.6: </a><a href="#0_23">The percentage of differentially expressed (DE) proteins in the cerebral </a><a href="#0_23">ganglia of stung cockroaches, at the different time points after the wasps sting</a><a href="#0_23">.</a><a href="#0_23">....</a><a href="#0_23">58 </a>1<a href="#0_24">Figure 3.7</a><a href="#0_24">: Heatmap of differentially expressed proteins in CX, 24 hours after the </a><a href="#0_24">sting</a><a href="#0_24">.</a><a href="#0_24">.............................................................................................................................</a><a href="#0_24">61 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587775" target="_blank">Figure 3.8: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587775" target="_blank">Changes that are associated with the long term effect of the venom.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587775" target="_blank">....</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587775" target="_blank">62 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587776" target="_blank">Figure 3.9: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587776" target="_blank">Heatmap of differentially expressed proteins in brain, 24 hours after the </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587776" target="_blank">sting.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587776" target="_blank">.............................................................................................................................</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587776" target="_blank">63 </a><a href="#0_25">Figure 3.10</a><a href="#0_25">: Heatmap of differentially expressed proteins in SEG, 24 hours after the </a><a href="#0_25">sting.</a><a href="#0_25">.............................................................................................................................</a><a href="#0_25">64 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587778" target="_blank">Figure 3.11: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587778" target="_blank">Changes that are associated with the short term effect of the venom.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587778" target="_blank">.</a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587778" target="_blank">64 </a><a href="#0_26">Figure 3.12: </a><a href="#0_26">Changes that are associated with recovery from venom effect. </a><a href="#0_26">...........</a><a href="#0_26">67 </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587780" target="_blank">Figure 3.13: </a><a href="file:///C:/Users/kaiserm/OneDrive/Thesis%20FINAL/Thesis%20Maayan%20final.docx%23_Toc533587780" target="_blank">The highest count of enriched Go terms is found 24 hours after the sting. </a>

Molecular Mechanisms Underlying Cockroach Host Manipulation by a Parasitoid Wasp

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