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Uni of Notts, Structure Function & Analysis of Genes, year 2, topic 9
| Term | Definition |
|---|---|
| RNA Pol I RNA Pol II RNA Pol III | Transcribes rRNA (excluding 5S rRNA) Transcribes pre-mRNA Transcribes tRNA, 5S rRNA, & other small RNAs e.g., Okazaki templates |
| Why eukaryotes share common cellular features with prokaryotic ancestors - Brenner | Nature can't build novel biological systems from nothing, only improve on what exists. Many parts that kept prokaryotes alive are conserved due to there being no pressure to change them |
| Modularity of elements in genetic regulation | Regulatory DNA & protein factors are discrete, reusable parts which may function independently & in combination for a variety of different effects |
| Benefits of genetic regulation being modular | Combinatorial coding of generic modules means no need for bespoke regulators for each gene & tissue-specific expression while the expression of these modules is more efficient |
| Why structures of genetic control factors is so similar across eukaryotes | The genetic code is universal, all DNA has common chemical properties, identical structural motifs & fold patterns are conserved due to their predictable effect on DNA |
| Hox protein family | Homeotic ubiquitous gene-selector (TF) proteins involved in morphogenesis & body patterning by binding to many DNA sequences with variable affinity |
| Homeotic meaning | Genes regulating the development of anatomical structures & body patterning in development. Homeosis is body segmentation |
| How lacz staining can test for Hox | Inserting a lacz gene (to code for β-galactosidase) down stream of a Hox promotor then culturing with X-Gal will create a blue stain, showing if the Hox gene is active in that tissue |
| Pax6 | Hox protein involved in eye, forebrain, & islets of Langerhans development. Contains paired & homeodomain helix-turn-helix subdomains with a C-terminal transactivation domain to recruit cellular machinery |
| Pax6 expression in non-eye tissues | Competent embryonic cells have cellular machinery to create any organs, redirecting signalling pathways & imposing spatial differentiation causes sightless eyes to form in any tissue expressing Pax6 |
| Why Pax6 gene taken from a donor organism doesn't change eye development when expressed in the host organism | Pax6 doesn't directly encode any eye characteristics, only transcription factors to activate specific genes encoding eye characteristics already present in the host DNA |
| Generic eukaryotic TF structures | Beads-on-string structure, similar to chromatin, of patterns of modular globular DNA binding & activator domains linked with IDR peptide linkers. The combination of the same domains & linkers are different in different genes |
| Intrinsically Disordered protein Regions (IDRs) & 2 protein folding paradigms | Polypeptides which serve cellular functions while remaining in 1' structures under physiological conditions. Follows disorder-function paradigm as opposed to the structure-function paradigm |
| How pH affects chemistry of TF domain properties | DNA binding sites are basic proteins to form ionic bonds with phosphate backbone in the grooves, activator regions are acidic have internal disorder so they can be flexible & bind to basic co-activators & cellular machinery |
| Alternative splicing in TF generation | Alters exons in mRNA to change function of the same TF across different cell lines by altering DNA binding & regulator domain structure, creates pattern of gene regulation isoforms depending on splicing factors |
| UAS | Upstream Activation Sequence in GAL genes |
| Gal4 Gal80 Gal3 | Dimeric transcriptional activator Gal4 repressor Galactose sensor, derepresses Gal4 |
| 4 UASs of GAL | Each has different affinity for Gal4, in low concentrations only high affinity will bind but remodel chromatin to raise affinity. The more UAS occupied, the greater the GAL expression |
| Combinatorial coding of GAL UAS | Different combinations of occupied binding sites recruit cellular machinery & remodel nucleosomes differently causing graded expression |
| Regulation of GAL1: -glucose, -galactose | Gal80 sequesters Gal4 on the UAS & no expression occurs |
| Regulation of GAL1: -glucose, +galactose | Gal3 causes Gal80 to dissociate from Gal 4 allowing for GAL1 expression relative to galactose concentration |
| Regulation of GAL1: +glucose, +galactose | Recruits the TUP-Mig1 repressor to bind to the Mig1 repression site & prevent expression by compacting chromatin since the cell favours glucose metabolism |
| Why regulation is achieved by repression rather than activation in bacteria | Repression is faster & energetically cheaper when they have no chromatin & their transcription & translation are coupled since genes that aren't immediately useful are a waste or resources |
| Eukaryotic parallels with Lac operon | Chromatin is inherently repressive but certain conditions mean the gene shouldn't be expressed, like Lacl binding to the Lac-operon until lactose binds causing it to dissociate |
| Effects of knockout mutation on TFs | Knockout of either the DNA binder or activator causes complete loss of function unless the activator is bound to another binding domain & the target of that domain replaces the UAS |
| How activators COULD work independently (E.Coli nitrogen repressor/activator example) | It's possible that in truly saturating conditions, just the activators could randomly bump into DNA by mass action. In E. Coli the gene will only be turned on when phosphorylated in low N conditions |
| Why activators must be physically close to transcription | Transcription needs direct protein-protein contact between factors & RNA Pol & intermolecular forces are short-ranged |
| Effects of mutation in Gal4: Deleting acidic activator | Reduces expression when bound but still leads to some expression & doesn't prevent Gal80 from binding |
| Effects of mutation in Gal4: Deleting acidic activator & adding the sequence to Gal80 + what this implies | GAL1 strongly expressed when Gal80 binds, demonstrating this segment is the only part of Gal4 which initiates transcription & the rest of the structure is for support & tethering |
| Difference between GAL & E. Coli Mer activation | GAL simply relies on physical proximity to promotor whereas Mer operon transmits regulatory information through structural distortion of the DNA in order to realign promotor elements |
| Chromatin Immunoprecipitation (ChIP) | Crosslinks to proteins bound to living cell DNA & can be measured to identify location of occupancy on DNA & binding enrichment at different points in time |
| ChIP analysis of GAL -galactose | Gal4 conc. is high at UAS with TATA-binding protein (TBP) low or absent, chromatin is repressed by Gal80 & TUP-Mig1 |
| ChIP analysis of GAL +galactose | Gal3 sequesters Gal80 lowering conc. Gal4 conc. remains constant, TBP, RNA Pol II, & other mediators are recruited increasing enrichment |
| Effect of DNase1 on specialised tissues | Tissue speciality is determined by combination of active genes. Active genes have loose chromatin which DNase1 can penetrate & destroy. Inactive tightly packed genes aren't affected |
| Mediator | Multidomain complex, similar to tau in prokaryotes but is more functional that structural, bridges TFs with RNA Pol II at promotors. The combination of activators & repressors determines gene expression |
| TF chromatin penetration | All TFs can penetrate chromatin to some extent, although most need histone acetylation to do so & can function by mass action solely at saturating concentrations |
Created by:
Denny12
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