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Divergent Evolution of Animal Excretory Systems

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Divergent Evolution of Animal Excretory Systems DIVERGENT EVOLUTION OF ANIMAL EXCRETORY SYSTEMS: EVIDENCE FROM MOLECULAR AND FUNCTIONAL STUDIES IN PLANARIANS by Hanh Thi Kim Vu A dissertation submitted to the faculty of The University of Utah in partial fulfillment of the requirements for the degree of Doctor of Philosophy Department of Neurobiology and Anatomy The University of Utah May 2015 Copyright © Hanh Thi Kim Vu 2015 All Rights Reserved The University of Utah Graduate School STATEMENT OF DISSERTATION APPROVAL The dissertation of Hanh Thi Kim Vu has been approved by the following supervisory committee members: Alejandro Sánchez Alvarado , Chair 12-04-2014 Date Approved Charles L. Murtaugh , Member 12-04-2014 Date Approved Tatjana Piotrowski , Member 12-04-2014 Date Approved Yukio Saijoh , Member 12-04-2014 Date Approved Monica L. Vetter , Member 12-04-2014 Date Approved and by Monica L. Vetter , Chair/Dean of the Department/College/School of Neurobiology and Anatomy and by David B. Kieda, Dean of The Graduate School. ABSTRACT Animals have developed extraordinary capacities to maintain homeostasis in the face of severe osmoregulatory challenges from their environment. For instance, with respect to salt and water homeostasis, freshwater animals continuously eliminate excess water while conserving solutes, whereas land-dwelling organisms have to conserve water and solutes as much as possible. Comparative morphological studies suggest that animals have tackled the problems of excretion and osmoregulation by evolving a specialized structure: the excretory organ. Animal excretory organs are extremely diverse. Some are unicellular, such as the excretory cell in nematodes. Others are multicellular and highly specialized, such as the protonephridia/metanephridia in invertebrates or the kidneys in vertebrates. In light of such anatomical and functional diversity, the evolutionary origins of animal excretory systems pose an interesting question in biology. However, the hypotheses proposed thus far remain highly controversial for two main reasons. First, many evolutionary arguments are based solely on morphology in organisms for which no molecular data are available, precluding rigorous genetic comparisons. Second, while invertebrates are critical elements of this evolutionary puzzle, the molecularly tractable ones studied to date display highly derived excretory systems. C. elegans possesses a single excretory cell, while the ultrafiltration of nephrocytes is uncoupled from the absorption/secretion of Malpighian tubules in D. melanogaster. Comparative morphological studies have demonstrated the existence of more complex excretory organs amongst many other invertebrates, including planarians. Planarians have a protonephridial excretory system in which each protonephridial unit consists of a tubule, opening distally via a nephridiopore at the surface of the animal and ending proximally in one or more terminal structures called flame cells. Protonephridia are commonly found amongst many invertebrates. Since protonephridia combine ultrafiltration with filtrate modification, planarians close an “invertebrate gap” in the study of excretory system biology. Taking advantage of a rapidly expanding list of molecular tools in recent years, this dissertation project aims to perform a comprehensive molecular and functional study of planarian protonephridia in order to provide new insights into the longstanding question on the evolutionary relationship between vertebrate and invertebrate excretory systems and gauge planarians’ potential as a novel invertebrate model for human kidney development and disease. iv TABLE OF CONTENTS ABSTRACT ....................................................................................................................... iii LIST OF FIGURES .......................................................................................................... vii ACKNOWLEDGEMENTS .................................................................................................x Chapters 1. OVERVIEW ..................................................................................................................1 A Brief Overview of Kidney Development ...................................................................6 Invertebrate Model Systems for Kidney Development and Diseases ..........................20 The Planarian Protonephridial Excretory System ........................................................21 Research Summary ......................................................................................................27 References ....................................................................................................................28 2. THE MAINTENANCE AND REGENERATION OF THE PLANARIAN EXCRETORY SYSTEM IS REGULATED BY EGFR SIGNALING .......................34 Abstract ........................................................................................................................35 Introduction ..................................................................................................................35 Results ..........................................................................................................................38 Discussion ....................................................................................................................68 Materials and Methods .................................................................................................78 References ....................................................................................................................81 3. STEM CELLS AND FLUID FLOW DRIVE CYST FORMATION IN AN INVERTEBRATE EXCRETORY SYSTEM..............................................................85 Abstract ........................................................................................................................86 Introduction ..................................................................................................................86 Results ..........................................................................................................................89 Discussion ..................................................................................................................160 Materials and Methods ...............................................................................................165 References ..................................................................................................................172 4. CONCLUSIONS AND PERSPECTIVES .................................................................178 Planarian Protonephridia – New Perspectives on the Evolutionary Origin of Vertebrate Nephrons ..................................................................................................179 Planarian Protonephridia – A Novel Invertebrate Model for Better Understanding Kidney Biology ..........................................................................................................184 Planarian Protonephridia – New Opportunities to Study Stem Cell Based Kidney Regeneration ..............................................................................................................189 Conclusion: A Model for All Reasons .......................................................................191 References ..................................................................................................................191 Appendices A. SUMMARY INFORMATION OF THE PLANARIAN HOMOLOGS OF SOLUTE CARRIER GENES ....................................................................................................195 B. EXPRESSION DOMAINS OF SOLUTE CARRIER GENES IN THE PLANARIAN PROTONEPHRIDIA .................................................................................................216 C. EXPRESSION DOMAINS OF SOLUTE CARRIERS IN THE RODENT METANEPHROS ......................................................................................................218 D. SUMMARY INFORMATION OF THE PLANARIAN HOMOLOGS OF HUMAN KIDNEY DISEASE GENES .....................................................................................223 vi LIST OF FIGURES 1.1. Cartoon representing the basic structural organization of excretory systems across animal kingdom .......................................................................................................4 1.2. Basic structural organization of the vertebrate nephron ..........................................7 1.3. Development of the vertebrate kidney ...................................................................10 1.4. Genetic networks controlling early lineage determination of the metanephric kidney, ureteric bud branching, nephron induction, and segmentation .................14 1.5. The planarian protonephridial excretory system ....................................................23 2.1. Distribution of protonephridia ...............................................................................39 2.2. Ultrastructure of protonephridial cell types in high-pressure frozen specimens ...43 2.3. Molecular anatomy of protonephridia ...................................................................46 2.4. Protonephridia regeneration ...................................................................................50 2.5. Proximal tubule defects cause edema formation ...................................................53 2.6. Smed-EGFR-5 is required for protonephridia function and expressed in flame cells .......................................................................................................................55
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