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Position Effect Variegation

Position Effect Variegation. 1930- first described. “The mosaic phenotype caused by a chromosomal position effect in which a rearrangement breakpoint displaced the white gene from its normal euchromatic location and placed it in the vicinity of heterochromatin”. Wakimoto Cell 93:321, 1998.

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Position Effect Variegation

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  1. Position Effect Variegation 1930- first described “The mosaic phenotype caused by a chromosomal position effect in which a rearrangement breakpoint displaced the white gene from its normal euchromatic location and placed it in the vicinity of heterochromatin” Wakimoto Cell 93:321, 1998

  2. Position effect variegation- pp448, Alberts Definition- Translocation of a gene from a euchromatic region to a heterochromatic region resulting in inactivation of nearby heterochromatic genes. • Called “heterochromatic spreading”, but is an incomplete definition

  3. Position Effect Variegation The white gene produces red eyes Variegated Wild-type Dorer and Henikoff Cell 77:993, 1994

  4. Single copy of “mini-white” locus inserted near centromere • in null-white fly strain Heterochromatin tandem repeat three repeats four repeats PEV- effect of transgene repeats Inverted repeat Single copy Thus, repeat number and orientation affect PEV

  5. PEV can be suppressed by modifiers Six-copy mini-white gene at 50C in Su(var)295 flys controls

  6. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD Models of PEVA. Cis-spreading Note: Histone acetylation effects PEV 1. Cis-spreading, block factor binding 2. Cis-spreading, form repressor complex with factors Thus, more spreading = more variegation

  7. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD Models of PEVA. Cis-spreading Problems with cis-spreading model • Some hetero-euchromatin rearrangements induce PEV several megabases away • PEV is sensitive to interchromosomal interactions • Thus, trans-interactions are suggested

  8. ModelsB. Nuclear compartment modelA trans-effect model Evidence in support- • Centromeres and most heterochromatin is located at one end of nucleus, telomeres at opposite end • Displaced heterochromatic regions interact with other heterochromatic regions • prevented by modifiers of PEV • However- have not yet correlated measured transcriptional activity and nuclear localization

  9. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD How does PEV Occur? Lets look at Telomere Position Effect RAP1- • Telomere Position Effect- Rap1 in complex with SIR proteins (SIR2/SIR3/SIR4) and histones H4 + H3 • Functions- heterochromatin assembly; recruitment of SIR proteins Folding-back mechanism

  10. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD Models of PEV • Over 120 modifiers (enhancers and suppressors) of PEV identified • Only some are directly involved • HP-1, Su(var)3-7 both co-localize to heterochromatin, interact in yeast two hybrid assay • Neither binds DNA

  11. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD What about genes normally active in heterochromatin? • Flys have over 20 expressed genes located in heterochromatin • >7 of these genes require placement in heterochromatin for normal expression • If place into euchromatic region- PEV results! • 1/2 of mutations that suppress PEV of euchrom. genes also enhance PEV of heterochrom. genes

  12. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD What about genes normally active in heterochromatin? Heterochromatin binding proteins interact with transcription factors to activate transcritpion or mediate long-range enhancer-promoter communication Thus, Rap1p may may have repressor role in euchromatin, activator role in heterochromatin

  13. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD How is PEV maintained? • No current model is satisfactory • Not DNA methylation -(Flys don’t do this) • GAGA protein binds to heterochromatin, remains throughout cell cycle • DNA must be “tagged” to maintain a given level of PEV during subsequent cell divisions • Competition for factors at each cell cycle?

  14. Advanced Molecular and Cellular Biology Bio4751 Spring 2003 Gary A. Bulla, PhD Recent result Mini-white gene Gal4-responsive Green Fluor. Protein • What happens if have two genes (GFP and mini-white) near centromere? Centromere Ahmad and Henikoff, Cell 104:839, 2001.

  15. Ahmad and Henikoff, Cell 104:839, 2001. GFP in near heterochromatin GFP in euchromatin High GAL4 Low GAL4 Euchrom. Heterochrom. • Observe GFP expression is variegated next to heterochromatin • And as increase Gal4, suppress variegation

  16. What happens to a nearby gene (the mini-white gene)? Ahmad and Henikoff, Cell 104:839, 2001. Miniwhite near heterochromatin Miniwhite in euchromatin GAL4? No Yes No Yes Thus, GAL4 binding counteracts silencing at nearby mini-white locus

  17. Ahmad and Henikoff, Cell 104:839, 2001. Miniwhite near heterochromatin Note- GFP on, mini-white off!! Can GFP and mini-white variegation be uncoupled? • Thus, GFP and mini-white silencing can be uncoupled.! • Heterochromatic boundary may be within 2 kb of DNA. • Heterochomatic spreading in not continuous

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