Adaptation and stress defence in intertidal and subtidal Antarctic limpets (Nacella concinna) [Elektronische Ressource] : a study of the plasticity of molecular and biochemical stress response in Antarctic invertebrates = Anpassung und Stressabwehr bei der Antarktischen Napfschnecke Nacella concinna aus dem Sublitoral und Eulitoral / vorgelegt von Ellen Weihe

Ellen Weihe

Sujets

• Biologie

Informations

• Publié par: universitat_bremen
• Publié le: 01 janvier 2009
• Nombre de lectures: 28
• Langue: Deutsch
• Poids de l'ouvrage: 2 Mo

Extrait

Adaptation and stress defence
in intertidal and subtidal Antarctic limpets (Nacella concinna):
A study of the plasticity of molecular and biochemical stress response in Antarctic
invertebrates


Anpassung und Stressabwehr
bei der Antarktischen Napfschnecke Nacella concinna aus dem Sublitoral und Eulitoral:
Eine Studie über die Plastizität molekularer und biochemischer Stressreaktionen in
Antarktischen Wirbellosen




Dissertation
zur Erlangung des akademischen Grades
– Dr. rer. nat. –



dem Fachbereich 2 Biologie / Chemie
der Universität Bremen



vorgelegt von
Ellen Weihe
Diplom-Biologin



Bremen 2009




















Prüfungsausschuß:
1. Gutachter: PD Dr. Doris Abele (Alfred-Wegener Institut für Polar- und Meeresforschung,
Bremerhaven)
2. Gutachter: Prof. Dr. Ralf Dringen (Universität Bremen)
1. Prüfer: Prof. Dr. Hans-Otto Pörtner (Alfred-Wegener Institut für Polar- und
Meeresforschung, Bremerhaven)
2. Prüfer: Dr. Magnus Lucassen (Alfred-Wegener Institut für Polar- und
Contents
Table of Contents
Table of Contents.................................................................................................. v
List of Figures...................................................................................................... vii
List of Tables viii
Terms, Definitions, and Abbreviations............................................................... ix
Zusammenfassung..............................................................................xiii
Summary.............................................................................................xvii
1 Introduction.......................................................................................1
1.1 Living in the Antarctic intertidal................................................................. 1
1.2 Reactive oxygen species and low oxygen concentrations...................... 3
1.3 Cellular oxygen sensing and response ..................................................... 7
1.4 Organismal response to changing oxygen (marine) .............................. 10
1.5 Anaerobic metabolism .............................................................................. 11
1.6 The hypoxia inducible factor.................................................................... 13
1.7 Objectives of the study ............................................................................. 16
2 Materials and Methods...................................................................19
2.1 The Antarctic limpet Nacella concinna.................................................... 19
2.1.1 Animal collection............................................................................................21
2.2 Methods...................................................................................................... 23
2.2.1 Experimental set-ups and tissue sampling..................................................24
2.2.2 Whole animal analyses ..................................................................................26
2.2.3 Biochemistry...................................................................................................28
2.2.4 Molecular biology ...........................................................................................30
3 Publications ....................................................................................37
Differences in the physiological response of inter- and subtidal Antarctic limpets
Nacella concinna to aerial exposure...........................................................................39
v Contents
Differences in heavy metal concentrations and in the response of the antioxidant
system to hypoxia and air exposure in the Antarctic limpet Nacella concinna......55
Characterisation of hypoxia inducible transcription factor HIF-in the Antarctic
limpet Nacella concinna ...............................................................................................67
4 Additional Results ..........................................................................99
4.1 Adenylate concentrations during hypoxia .............................................. 99
4.2 Ascorbyl radical content during hypoxia ...............................................100
5 Discussion.....................................................................................103
5.1 Hypoxic Metabolism in N. concinna .......................................................103
5.2 The transcription factor HIF- in N. concinna........................................107
5.2.1 Regulation of HIF-during stress...............................................................109
6 Conclusion and Perspectives......................................................113
7 References ....................................................................................117
8 Appendix .......................................................................................130
Danksagung........................................................................................................132

vi Contents
List of Figures
Figure 1-1: Limpets during aerial exposure..............................................................................2
Figure 1-2: The univalent reduction of the oxygen molecule....................................................4
Figure 1-3: ROS formation and antioxidant defence. ...............................................................5
Figure 1-4: Electron transport chain in the mitochondrial membrane.......................................6
Figure 1-5: Summary of bioenergetic and biosynthetic O sensors and signalling 2
mechanisms. ............................................................................................................................9
Figure 1-6: Anaerobic metabolic pathways. ...........................................................................12
Figure 1-7: HIF-peptide structure. .......................................................................................14
Figure 1-8: HIF-regulation scheme......................................................................................15
Figure 2-1: Distribution of Nacella concinna...........................................................................20
Figure 2-2: Nacella concinna in the intertidal on King George Island during low tide. ...........21
Figure 2-3: cinna in the sub-littoral in 13m depth at King George Island.............21
Figure 2-4: Map of King George Island. .................................................................................22
Figure 2-5: Map of Potter cove...............................................................................................22
Figure 2-6: Set up for air exposure experiment......................................................................25
Figure 2-7: Hypoxia incubation of Nacella concinna. .............................................................25
Figure 2-8: Shells of sub-littoral and intertidal Nacella concinna............................................26
Figure 2-9: Experimental set-up of PO measurement in the inner shell water......................272
Figure 2-10: Picture of the sub-littoral in Potter Cove showing plumes of sedimentation. .....28
Figure 2-11: Schematic overview of the P. vulgata HIF-cDNA fragment, its functional
domains, and the used primers..............................................................................................31
Figure 2-12:the N. concinna HIF-cDNA and the used primers......32
Figure 4-1: Ascorbyl radical concentration in the digestive gland of intertidal and sub-littoral
N. concinna during hypoxia..................................................................................................101
Figure 5-1: Anaerobic metabolites in foot muscle of sub-littoral Nacella concinna. .............105
Figure 5-2: in foot muscle of intertidal . ................107
Figure 5-3: Reaction scheme of HIF-hydroxylation by PHDs............................................110
Figure 8-1: Primary sequence and deduced protein sequence of N. concinna HIF- cDNA
.............................................................................................................................................131

vii Contents
List of Tables
Table 2-1: Overview of expeditions, conducted experiments, measured parameters and
where the results are published. ............................................................................................23
Table 2-2: Overview of measured parameters and tissues....................................................24
Table 2-3: List of all primers used in this study for sequencing..............................................33
Table 2-4: Primer and reporter used for Real-Time PCR.......................................................34
Table 4-1: Adenylate concentrations in foot muscle of intertidal and sub-littoral Nacella
concinna during hypoxia.......................................................................................................100
Table 5-1: Km and Ki values of parameters influencing prolyl hydroxylase activity. ............112
viii Abbrevia