Einführung in die Genetik
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1 Einführung in die Genetik Prof. Dr. Kay Schneitz (EBio Pflanzen) #genetiktum FB: Plant Development TUM Prof. Dr. Claus Schwechheimer (PlaSysBiol)
2 Einführung in die Genetik - Inhalte 1 Einführung KS 2 Struktur von Genen und Chromosomen KS 3 Genfunktion KS 4 Transmission der DNA während der Zellteilung KS 5 Vererbung von Einzelgenveränderungen KS 6 Genetische Rekombination (Eukaryonten) KS 7 Genetische Rekombination (Bakterien/Viren) KS 8 Rekombinante DNA-Technologie CS 9 Kartierung/Charakterisierung ganzer Genome CS 10 Genmutationen: Ursache und Reparatur CS 11 Veränderungen der Chromosomen CS 12 Genetische Analyse biologischer Prozesse CS 13 Transposons bei Eukaryonten CS 14 Regulation der Genexpression KS 15 Regulation der Zellzahl - Onkogene CS
3 Transposons Genetics 13
4 Summary Discovery of transposons (Mais) Structure of transposons (simple vs. composite transposons) Replicative vs. conservative transposition Transposons and bacterial antibiotic resistance RNA transposons, retrotransposons (retrovirus, gag, pol, env) Experiment (retrotransposons, RNA intermediate) DNA transposons (P elements, Ac/Ds) Excision (short DNA repeat = footprint) Transposons and genome evolution Human genome (LINE and SINE RNA transposons; DNA transposons) Plant genomes (genome size and transposons) Safe havens (intergenic regions, introns, rrna genes, gene promoters)
5 Regulation of Gene Expression Genetics 14
6 Topics Gene regulation in bacteria Genetic analysis of gene induction Basic logical and molecular principles of genetic switches Gene regulation in eukaryotes Chromatin
7 Overview of transcription
8 E. coli: Initiation
9 E. coli: The promoter
10 Eukaryotes: Initiation
11 Why?
12 Gene regulation in bacteria
13 Genetic Switches An/Ausschalter
14 Positive vs negative regulation
15 Modular functionality: e.g., allosteric effectors
16 Model: the lac system of E. coli Induction of ß-galactosidase by the disaccharide lactose Francois Jacob André Lwoff Jacques Monod
17 The inducer: lactose
18 Simplified lac operon model
19 No lactose present
20 Lactose present
21 Genetic dissection of lac system Genetic components I I -, I s Y Y - P P - Z Z - O O c A A -
22 Generating partial diploids in E. coli
23 O c mutants: Synthesis of ß-galactosidase/permease
24 O c mutants: Synthesis of ß-galactosidase/permease
25 O c mutants: Synthesis of ß-galactosidase/permease
26 O c mutants: Synthesis of ß-galactosidase/permease
27 O c mutants: Synthesis of ß-galactosidase/permease
28 O c mutants: Synthesis of ß-galactosidase/permease O c is a constitutive mutation
29 O c mutants: Synthesis of ß-galactosidase/permease O c is a constitutive mutation Operator (O) is cis-acting
30 Interpretation
31 Interpretation Operators are cis-acting
32 I - mutants: Synthesis of ß- galactosidase/permease
33 I - mutants: Synthesis of ß- galactosidase/permease
34 I - mutants: Synthesis of ß- galactosidase/permease
35 I - mutants: Synthesis of ß- galactosidase/permease
36 I - mutants: Synthesis of ß- galactosidase/permease
37 I - mutants: Synthesis of ß- galactosidase/permease Repressor (I) is trans-acting
38 Interpretation
39 Interpretation Repressors are trans-acting
40 I s alleles: Synthesis of ß- galactosidase/permease
41 I s alleles: Synthesis of ß- galactosidase/permease
42 I s alleles: Synthesis of ß- galactosidase/permease
43 I s alleles: Synthesis of ß- galactosidase/permease
44 I s alleles: Synthesis of ß- galactosidase/permease I s : repressor is hyperactive
45 Interpretation
46 Interpretation Repressor contains a lactose-binding site
47 Lactose present
48 Genetic analysis: I vs O both elements act in repressing lac operon they fundamentally differ in their mode of action (cis vs trans mode) reveals important aspects of the molecular mechanism of lac repression
49 Control of lac system: Lactose vs glucose
50 Catabolite repression of the lac operon
51 Catabolite repression of the lac operon Glucose is a catabolite of lactose camp/cap complex is an activator of transcription
52 Control of the lac operon
53 Molecular anatomy of the genetic switch
54 The operator is a specific DNA sequence
55 The operator is a specific DNA sequence Very specific sequence! One base change enough to eliminate O function
56 Many DNA binding sites are symmetrical
57 Binding of CAP bends DNA
58 Binding of CAP bends DNA Recognition of specific CAP-binding site
59 CAP/RNA pol: Binding sites
60 Helix-turn-helix is a common DNA-binding motif
61 Helix-turn-helix is a common DNA-binding motif Specific contacts with bases in major groove
62 Helix-turn-helix is a common DNA-binding motif Dimers Specific contacts with bases in major groove
63 AA side chains determine specificity of DNA binding Homeodomain
64 Repression vs activation
65 Genetic switches are often part of a cascade mechanism A TF B C
66 Summary Cells respond to intrinsic and extrinsic signals by modulating transcriptional control of certain genes Gene activity is the result of the function of cis- and trans-acting factors Trans-acting proteins react to environmental signals by using built-in sensors that continually monitor cellular conditions Coordinated gene regulation in bacteria gene are often clustered into operons on the chro and transcribed together into multigenic mrnas one cluster of regulatory sites per operon is sufficient to regulate expression of several genes Negative vs positive regulation repressor proteins bind to DNA at operator site thereby blocking transcription (e.g., lac operon) activator proteins activate transcription by binding to DNA at the promoter region (e.g., camp/cap regulation of lac operon) Molecular anatomy of genetic switch regulatory proteins have DNA-binding domains (e.g., HLH) and protein-protein interaction domains (modular specificity of gene regulation depends on specific protein-dna interactions mediated by the chemical interactions between aa side chains and chemical groups of DNA bases
67 Gene regulation in eukaryotes
68 Drosophila: MSL complex and dosage compensation
69 Overview of transcriptional regulation
70 Eukaryotic promoter
71 The yeast GAL system
72 The transcriptional activator Gal4
73 The transcriptional activator Gal4 TF: sequence-specific binding to regions outside the promoters of target genes
74 TFs: Modular Proteins
75 Transcriptional activators and the transcription machinery
76 Enhancer action: Mechanism
77 Enhancer action: Mechanism Cooperativity Synergism
78 Disperse distribution of enhancer elements DPP of Drosophila kb
79 Modular and combinatorial control eve TATA lacz
80 Chromatin
81 Chromatin dynamics
82 Chromatin remodeling e.g., by SWI-SNF complex
83 Histone modifications and the histone code
84 Tup1, a histone deacetylase from yeast, is a corepressor
85 Linking TFs and chromatin dynamics
86 Enhanceosome
87 Enhanceosome Cooperativity Synergism
88 Enhanceosome Cooperativity Synergism
89 Summary Eukaryotic gene regulation resembles bacterial gene regulation trans-acting factors binding to cis-regulatory elements on the DNA this regulatory factors determine the level of transcription by regulating the binding of RNA pol II to the promoter of a gene Enhancers/UAS cis-regulatory elements, possibly located quite far away (>10-50kb) from promoter combinatorial interactions among different transcription factors enhanceosome: complexes of regulatory proteins that interact in cooperative and synergistic fashion --> high levels of transcription through recruitment of RNA pol II Gene regulation and chromatin eukaryotic genes are packed in chromatin activation/repression requires specific modifications to chromatin genes are mostly turned off and kept silent in part by nucleosomes and condensed chromatin histone code: pattern of posttranslational modifications of histone tails (acetylation, methylation, phosphorylation etc). histone code is an epigenetic mark involved in nucleosome positioning and chromatin condensation that can be altered by TFs Tfs recruit for example ATP-dependent chromatin remodelers (e.g., SWI-SNF)
90 THE END
Einführung in die Genetik
Einführung in die Genetik Prof. Dr. Kay Schneitz (EBio Pflanzen) http://plantdev.bio.wzw.tum.de schneitz@wzw.tum.de Prof. Dr. Claus Schwechheimer (PlaSysBiol) http://wzw.tum.de/sysbiol claus.schwechheimer@wzw.tum.de
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