Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae:
Gespeichert in:
| 1. Verfasser: | |
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
2012
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| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | VI, 131 S. Ill., graph. Darst. |
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| adam_text | IMAGE 1
TABLE OF CONTENT
TABLE O F CONTENT I
SUMMARY. I L L
ZUSAMMENFASSUNG V
1. INTRODUCTION 1
1.1 CYTOKINESIS IN EUKARYOTES 1
1.2 CYTOKINESIS IN SACCHAROMYCES CEREVISIAE 2
1.3 THE CYTOKINETIC MACHINERY 3
1.3.1 SEPTINS 3
1.3.2 MYOSIN AND ACTIN 4
1.3.3 COMPONENTS OF THE AMR-DEPENDENT AND INDEPENDENT PATHWAYS 6
1.3.4 TRILAMINAR SEPTUM 8
1.4. CELL CYCLE CONTROL OF CYTOKINESIS 10
1.4.1 FUNCTION OF THE MITOTIC EXIT NETWORK (MEN) IN CYTOKINESIS 10
1.5 CELL SEPARATION AND THE RAM PATHWAY 12
1.5.1 THE FAMILY OF CONSERVED NDR/LATS KINASES 13
1.5.2 THE NDR/LATS KINASE CBKL AND THE REGULATORY FACTOR MOB2 14
1.5.3 THE TRANSCRIPTION FACTOR ACE2 16
1.5.4 THE ACE2 DOWNSTREAM TARGETS 17
1.6 CELL CYCLE CONTROL O F CELL SEPARATION 17
1.7 THE HOMOLOGUES TO THE RAM PATHWAY IN FUNGI 19
1.7.1 MOR AND SIN IN SCHIZOSACCHAROMYCES POMBE 20
2. AIM O F THE STUDY 21
3. RESULTS. 23
3.1 FUNCTION O F THE ESSENTIAL C2 DOMAIN PROTEIN INNL IN CYTOKINESIS 23
3.1.1 CONSTRUCTION OF INN1 TEMPERATURE SENSITIVE DEGRON ( INNL-TD ) 23
3.1.1.1 CYK3 LOCALIZATION AT THE DIVISION SITE IS SIGNIFICANTLY REDUCED
IN INNL-TD CELLS 25 3.1.1.2 HOFL CONTRACTS ASYMMETRICALLY IN INNL-TD
CELLS 26
3.1.1.3 CHS2 ACCUMULATES STRONGER AT ONE SIDE OF THE NECK UPON INNL
DEGRADATION 27 3.1.1.4 CYK3 DOES NOT INFLUENCE INNL-GFP BUD NECK
LOCALIZATION 28
3.1.2 CONSTRUCTION OF INN1 TEMPERATURE SENSITIVE MUTANT ( INNL-TS ) 29
3.1.2.1 INN1 IS GENETICALLY LINKED TO THE CYTOKINETIC COMPONENTS HOF1,
CYK3 AND MYOL ..31 3.1.2.2 INN1 IS GENETICALLY LINKED TO THE MEN
COMPONENTS 32
3.2 FUNCTION O F LREL IN CELL SEPARATION 35
3.2.1 MULTICOPY SUPPRESSOR SCREENING OF TEML-3 CYK3A DOUBLE MUTANT 35
3.2.2 LRE1 OVEREXPRESSION ANALYSIS IN TEML-3 AND CYK3A MUTANTS 37
3.2.3 LREL DIRECTLY BINDS CBKL AND THE REGULATORY SUBUNIT MOB2 39
3.2.4 LREL COLOCALIZES AT THE BUD NECK WITH CBKL 43
3.2.5 LREL IS A SUBSTRATE OF CBKL 44
3.2.6 CBKL CATALYTICAL ACTIVITY IS TOXIC IN TEML-3 CYK3A CELLS 46
3.2.7 LREL INHIBITS CBKL CATALYICAL ACTIVITY 48
3.2.8 LREL INFLUENCES CBKL DAUGHTER NUCLEAR LOCALIZATION 50
3.2.9 LREL ACTS ON THE CBKL-DEPENDENT ACE2 PATHWAY 52
3.2.10 LRE1 OVEREXPRESSION AND CBK1 DELETION RESTORE THE FORMATION OF
THE SEPTUM IN TEML3 CYK3-AID CONDITIONAL DOUBLE MUTANT 56
I
HTTP://D-NB.INFO/103249770X
IMAGE 2
3.2.11 CDCL4-DEPENDENT DEPHOSPHORYLATION OF LREL PROMOTES THE
INACTIVATION OF CBKL 59
3.3 PRELIMINARY DATA O F CYK3 FUNCTION IN CELL SEPARATION 63
3.3.1 CYK3 IS A MULTI DOMAINS CYTOKINETIC COMPONENT 63
3.3.2 CYK3 BINDS LREL IN VITRO 66
3.3.3 THE INTERACTION BETWEEN LREL AND CBKL IS NOT IMPAIRED IN ABSENCE
OF CYK3 66
4. DISCUSSION 69
4.1 REGULATION OF CYTOKINESIS BY THE ESSENTIAL C2 DOMAIN COMPONENT INNL
AND THE SH3 DOMAIN PROTEIN CYK3 69
4.1.1 INNL PROMOTES CYTOKINESIS VIA THE RECRUITMENT OF CYK3 AT THE BUD
NECK 69 4.1.2 DISSECTING THE FUNCTION OF THE MULTI DOMAIN PROTEIN CYK3
IN CYTOKINESIS 70
4.2 LREL NEGATIVELY REGULATES CELL SEPARATION VIA THE INHIBITION O F THE
NDR/LATS KINASE CBKL 72
4.2.1 LREL DOWN REGULATES THE ACE2-DEPENDENT CELL SEPARATION PATHWAY 72
4.2.2 A MODEL OF THE REGULATION OF CELL SEPARATION BY THE MEN-TRIGGERED
PHOSPHATASE CDCL4, VIA THE ACTIVATION OF LREL 75
4.3 FUTURE PERSPECTIVES 78
5. MATERIAL AND METHODS. 81
5.1 MATERIAL 81
5.1.1 CHEMICALS 81
5.1.2 BUFFERS AND SOLUTIONS 81
5.1.3 ENZYMES 83
5.1.4 ANTIBODIES 83
5.1.5 PROTEASE AND PHOSPHATASE INHIBITORS 84
5.1.6 PRIMERS 85
5.1.7 PLASMIDS 88
5.1.8 STRAINS 89
5.1.9 GROWTH MEDIA 93
5.2 METHODS 97
5.2.1 MOLECULAR BIOLOGY TECHNIQUES 97
5.2.2 SACCHAROMYCES CEREVISIAE SPECIFIC METHODS 103
5.2.3 GENERAL BIOCHEMICAL TECHNIQUES 106
5.2.4 PROTEIN EXPRESSION AND PURIFICATION FROM E. COLI 109
5.2.5 SPECIFIC SACCHAROMICES CEREVISIAE BIOCHEMICAL METHODS 110
5.2.6 SYNCHRONIZATION OF YEAST CELLS 111
5.2.7 MICROSCOPY METHODS 112
6. REFERENCES 117
7. APPENDIX. 125
7.1 SPLIT UBIQUITIN SCREENING 125
7.2 SYNTHETIC GENETIC ARRAY ANALYSIS 127
8. ABBREVIATIONS 129
II
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| dewey-search | 571.84429563 |
| dewey-sort | 3571.84429563 |
| dewey-tens | 570 - Biology |
| discipline | Biologie |
| format | Thesis Book |
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| language | English |
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| spelling | Mancini Lombardi, Ilde Verfasser aut Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae [presented by Ilde Mancini Lombardi] 2012 VI, 131 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Heidelberg, Univ., Diss., 2012 (DE-588)4113937-9 Hochschulschrift gnd-content DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=025684211&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Mancini Lombardi, Ilde Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae |
| subject_GND | (DE-588)4113937-9 |
| title | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae |
| title_auth | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae |
| title_exact_search | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae |
| title_full | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae [presented by Ilde Mancini Lombardi] |
| title_fullStr | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae [presented by Ilde Mancini Lombardi] |
| title_full_unstemmed | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae [presented by Ilde Mancini Lombardi] |
| title_short | Regulation of cytokinesis and cell separation in Saccharomyces cerevisiae |
| title_sort | regulation of cytokinesis and cell separation in saccharomyces cerevisiae |
| topic_facet | Hochschulschrift |
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