The cis regulatory elements Current Topics in Genomics and Epigenomics – Lecture 2
Jan 11, 2016
The cis regulatory elements
Current Topics in Genomics and Epigenomics – Lecture 2
Transcription Process in Eukaryotes
Transcription in Eukaryotes
Takes place in the nucleus
The lower strand is used as template
RNA is made from 5’ to 3’ direction
Multiple steps Class II genes are
transcribed by RNAP2 and requires post-transcriptional processing
The 5´ Capping
Process: A GTP is attached to the 5´
end via a 5´-5´ bond (Enzyme:Guanyltransferase)
First few nucleotides (including GTP at N#7) are methylated (Enzyme: Guanine- methyltransferase)
Effects: Facilitates splicing Stabilizes the mRNA
transcript Facilitates transport of
mRNA to cytoplasm Recognition by ribosomes in
translation
RNAbranch point
Splicing
Splicing and lariat formationvia successive trans-esterification reactions
Two exons are ligated, intron is released as a branched lariat molecule
Transcriptional initiation
RNAPII loading to promoters requires general transcription factors (GTF)
Transcriptional regulatory elements and their mechanisms of action
cis-regulatory elements in the genome
Insulator(Barrier)
Promoter Enhancer
Insulator (Enhancer blocking)
Silencing element
Insulator(Barrier)
Promoters
Promoters are where transcription initiates Many promoter elements have been found TATA box was discovered in 1979 by comparison
of a number of mRNA start sites
How to Locate promoters in the human genome?
Determine the full length transcripts and align to the genome
Cap Analysis of Gene Expression
How to Locate promoters in the human genome?
Determine the full length transcripts and align to the genome
Locate promoters based on binding sites of the pre-initiation complex
ChIP-based analysis is a general approach for mapping transcription factor binding sites
Locate promoters in the human genome
Locate promoters based on binding sites of the pre-initiation complex
ChIP-chip analysis is a general approach for mapping transcription factor binding sites
ChIP-seq provides a more cost-effective, genome-wide approach
Locate promoters in the human genome
- Kim et al., Nature 2005
Sequence features of promoters
CpG island is associated with nearly 80% of the promoters
TATA box is not a general feature of human promoters
Other sequence elements exists
cis-regulatory elements in the genome
Insulator(Barrier)
Promoter Enhancer
Insulator (Enhancer blocking)
Silencing element
Insulator(Barrier)
Levine M; Small S.
Promoter Enhancers
A cis regulatory element of Shh
Lettice et al., Hum Mol Gen 2003, 12 (14) pp. 1725-35Lettice et al, Proc Natl Acad Sci USA 2002 vol. 99 (11) pp. 7548-53
Enhancer
Preaxial polydactyly
Model of enhancer function
RNAP
RNAP
Coactivators
p300 is a transcriptional co-activator
-- www.web-books.com/ MoBio/Free/Ch4G2.htm
Locating enhancers by mapping the p300 binding sites
Z. Li in collaboration with Visel, Pennachio, and Rubin @ LBNL
Visel et al., Nature 2009
Confirmation of enhancer activities of the p300 binding sites in transgenic mouse embryos
• 53/67 (79%) forebrain predictions, 28/34 (82%) midbrain predictions and 20/25 (82%) limb predictions were active in vivo enhancers at e11.5
Visel et al., Nature 2009
cis-regulatory elements in the genome
Insulator(Barrier)
Promoter Enhancer
Insulator (Enhancer blocking)
Silencing element
Insulator(Barrier)
Gaszner and Felsenfeld Nature Reviews Genetics advance online publication;published online 15 August 2006 | doi:10.1038/nrg1925
Barrier Elements
Gaszner and Felsenfeld Nature Reviews Genetics advance online publication;published online 15 August 2006 | doi:10.1038/nrg1925
Gaszner and Felsenfeld Nature Reviews Genetics advance online publication;published online 15 August 2006 | doi:10.1038/nrg1925
Cohesins and CTCF colocalize on insulators
The ENCODE (ENCyclopedia Of DNA Elements) Project
The ENCODE Project was launched in 2003
Goal: Identify candidate functional elements in the human genome. Produce large scale datasets for public access.
Three phases
Pilot phase (2003-2007) – develop high throughput methods, focusing on 1% of genome
Production phase (2007 – 2012) – ramping up to genome-wide, focusing on a limited number of cell lines; model organisms (worm, fly and mouse) were also included.
Full production phase (2012 – 2016),just launched, aimed for comprehensive analysis.
http: //www.genome.gov/ENCODE
The ENCODE (ENCyclopedia Of DNA Elements) Project
Myers et al., PLoS Biol. 2011 vol. 9 (4) pp. e1001046
Genes & TranscriptsCis regulatory sequences
ChIP-seq Zhang et al. Genome Biol 2008
Annotating cis-elements from ChIP-seq data of Transcription factor binding
DNase-seq
Song & Crawford, Cold Spring Harb Protoc 2010
Chromatin State and accessibility in human DNA measured by ChIP-seq and DNase-Seq
Ecker J, Nature 2012, 489:53
>80% of the genome participates in one or more biochemical event (making RNA, binding TFs, etc) in at least one cell type;
Defined 399,124 regions with enhancer-like features and 70,292 regions with promoter-like features
4.5 million Dnase I hypersensitive sites defined Single nucleotide polymorphisms (SNPs) associated with
disease by GWAS are enriched within non-coding functional elements
Maurano et al… J. A Stamatoyannopoulos, Science 2012