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Gene Interaction

Gene Interaction. “Standard” interpretation of complementation test. Hawley & Gilliland (2006) Fig. 1. “Mutation” of a gene might be due to changes elsewhere!. ald is Drosophila mps1 homolog; isolated four mutations (all rescued by ald + transgene)

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Gene Interaction

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  1. Gene Interaction

  2. “Standard” interpretation of complementation test Hawley & Gilliland (2006) Fig. 1

  3. “Mutation” of a gene might be due to changes elsewhere! • ald is Drosophila mps1 homolog; isolated four mutations (all rescued by ald+ transgene) • two ald alleles cause meiotic and mitotic defects (ald sequence changes) • two ald “mutations” cause only meiotic defects (normal ald sequence) • both contain Doc element insertion into neighboring gene • (silences transcription of neighboring genes in germline cells) Hawley & Gilliland (2006) Fig. 2

  4. “False positive” of transgenic rescue • Ku and Dmblm genesboth involved in DNA repair and closely linked on the chromosome • Old mutations of mus309 map to the region genetically • DNA lesions of mus309 lie in Dmblm, but can be rescued with extra copies of Ku (provided on a transgene)

  5. Shared regions between genes

  6. Exceptions to “Non-Complementation = Allelism” • Intragenic complementation (usually allele-specific) • Multi-domain proteins (e.g., rudimentary) • Transvection – pairing-dependent allelic complementation (stay tuned!) • Second-Site Non-Complementation (“SSNC”) • “Poisonous interactions” – products interact to form a toxic product • (usually allele-specific) • “Sequestration interactions” – product of one mutation sequesters the other to a suboptimal concentration in the cell (usually one allele-specific) • Combined haplo-insufficiency (allele non-specific)

  7. Intragenic complementation in multi-domain proteins

  8. Transvection: synapsis-dependent allele complementation E. Lewis (1954) among BX-C mutations in Drosophila Numerous other genes in Drosophila and similar phenomena observed in Neurospora, higher plants, mammals Most due to enhancer elements functioning in trans (allele-specific)

  9. Exceptions to “Non-Complementation = Allelism” • Intragenic complementation (usually allele-specific) • Multi-domain proteins (e.g., rudimentary) • Transvection – pairing-dependent allelic complementation • Second-Site Non-Complementation (“SSNC”) • “Poisonous interactions” – products interact to form a toxic product • (usually allele-specific) • “Sequestration interactions” – product of one mutation sequesters the other to a suboptimal concentration in the cell (usually one allele-specific) • Combined haplo-insufficiency (allele non-specific)

  10. Example of a “Poisonous interaction” SSNC Non-complementation of non-allelic mutations Hawley & Gilliland (2006) Fig. 4 (after Stearns & Botstein (1988) Genetics119: 249–260)

  11. A model for synthetic lethality Figure 6-23

  12. A model for recessive epistasis Figure 6-19

  13. Dominant epistasis due to a white mutation Figure 6-21

  14. Recessive epistasis due to the yellow coat mutation

  15. Suppression: a form of epistasis whereby the expression of one mutation (the “suppressor” mutation) normalizes the phenotype of another mutation (the “suppressed” mutation). The suppressor mutation may display no other phenotype. Intragenic suppression: “pseudo-reversion”; can be same codon or different/interacting region of gene.

  16. Intragenic suppression by compensatory missense mutation: • Example: • p53 tumor suppressor* gene; DNA-binding protein; numerous mutations catalogued • Yeast reporter system (p53 binding site-UAS-URA3) requires p53 binding • Expressed mutant human p53 (does not drive URA3 expression) • Created variety of second-site mutations within p53, using gap-repair-mediated replacement of mutagenic PCR fragments into p53-containing plasmid (site-directed/not random mutagenesis) • Screening for URA3 expression identified array of second-site mutations

  17. identified suppressors for each of three original p53 mutations • (Ura+ and FoaR phenotypes) • most suppressing mutations were obtained multiple times • luciferase-reporter transfection assays in mammalian cell lines gave similar results; • p53-induced apoptosis assays in mammalian cells also gave similar results • created structural models for basis of suppression

  18. Val contributes to hydrophobic core of β sandwich Asn>Asp may shift H-bonds to maintain β sandwich Gly “fits” within DNA contact loop Asn>Tyr may provide new DNA contact (Y with PO4) Ser>Asn may provide new stabilizing H-bond to loop Arg is within H-bond network stabilizing DNA contact loop, eliminates quanidinium His>Arg may create new quanidinium interaction Thr>A/P is known to enhance DNA-binding affinity of wild-type p53

  19. Extragenic suppression: suppressor mutation is in different gene than suppressed mutation Classic example: tRNA anticodon mutations that suppress nonsense/frameshift mutations in other genes Classic example: eye color suppression in Drosophila

  20. A molecular mechanism for suppression Figure 6-22

  21. The Wnt Signaling Pathway

  22. Ommatidium: 8 photoreceptors (R1-8) 12 accessory cells sev mutations remove R7 photoreceptor (cell develops as non-neuronal cone cell) *non-lethal: functions only in precursor of R7 photoreceptor! sev encodes transmembrane protein tyrosine kinase closely related to mammalian c-ros Created sev ts mutation based on known v-src ts mutants, using in vitro mutagenesis

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