Gene Structure and Identification III

1
Gene Structure and Identification III
BIO520 Bioinformatics Jim Lund
2
For real prediction we need

• Solve the protein folding problem
• Solve the molecular docking/binding problem
• Develop realistic simulations of molecules in
  cells
• Simulate multicellular systems

3
Promoter/Enhancer analysis

• Regulatory Sequences
• Known Consensus Sequences
• Consensus Sequence Generation
• Using functional (experimental) Data
• Real examples

4
Gene Regulatory Sequences

• Functional sites
• Consensus
• Experimental tests
• Inferred sites
• Transcriptome analysis

5
Sequnce Logos

• http//weblogo.berkeley.edu/

6
(No Transcript)
7
Position Weight Matrix

• PO A C G T
• 01 6 4 4 6 N
• 02 4 9 3 4 N
• 03 12 4 3 1 A
• 04 6 1 11 2 R
• 05 3 2 11 4 G
• 06 3 3 4 10 N
• 07 3 10 3 4 N
• 08 11 2 4 3 A
• 09 4 9 3 4 N
• 10 3 6 3 8 N

8
EUKARYOTES

• More complex signals
• Basal/core promoter
• Promoter
• Enhancers
• More genes
• More dispersed signals
• Larger promoters, distant enhancers, regulatory
  sites in introns.
• Combinatoric regulation common

9
Basal Promoter Analysis
Myers and Maniatis, Genes VI, 831

• TATA-box -25 to -30 TBP
• CCAAT-box -212 to -57 CTF/NF1
• GC-box -164 to 1 SP1
• K C W K Y Y Y Y 1 to 5 cap signal

1
10
Finding PolII sites (transcription start site)

• Promoter Scan
• TSSG/TSSW (TSSP for plants)
• Core-Promoter
• FPROM
• BCM Search Launcher

11
Enhancer Elements

• Octamer OCT1, OCT2
• ?B NF ?B
• ATF ATF
• AP1 AP1
• ..

False , False -
12
Consensus Sequence Databases

• TRANSFAC
• TFD (transcription factor database)

13
Consensus Sequence Databases

• Finding sites in promoter regions
• TESS
• http//www.cbil.upenn.edu/cgi-bin/tess/tess
• TFSEARCH
• http//www.cbrc.jp/research/db/TFSEARCH.html
• BCM Search Launcher
• http//searchlauncher.bcm.tmc.edu/seq-search/gene-
  search.html

14
HBB promoter (TESS)
15
Sequence-based algorithms

• Genes from
• Microarray transcription analysis
• ChIPchip experiments
• Orthologous sequences
• Experimental/other
• Programs for finding consensus sites
• MEME analysis of clusters
• AlignAce
• BioProspector/CompareProspector

16
Practical Gene Finding

• Use ALL tools
• Predictive Stitch together a consensus
• ORF finders
• Find patterns (and WWW pattern searches)
• HMM GRAIL, Genscan
• Comparative
• BLASTN, BLASTX
• Compare genomes (humanmouse)
• cDNA, protein, genetic evidence

17
ORFs-aldolase gene
18
Genomic DNA-cDNA alignment
P
DNA sequencing
Align (GAP)
cDNA
19
Comparative Genomics

• Conservation of coding regions
• Identification of transcription signals
• words in common
• Example-yeast comparisons

20
Ensembl prediction pipeline
DNA
RepeatMasker
Genscan
Pmatch all human Proteins and cdnas
Blast genscan peptides v Protein,unigene,est,vert
mrna
MiniGenewise MiniEst2genome
Genes
21
(No Transcript)
22
(No Transcript)
23
Genscan features

• Model both strands at once
• Each state may output a string of symbols
  (according to some probability distribution).
• Explicit intron/exon length modeling
• Advanced splice site modeling
• Complete intron/exon annotation for sequence
• Able to predict multiple genes and partial/whole
  genes
• Parameters learned from annotated genes
• Separate parameter training for different CpG
  content groups (lt 43, 43-51, 51-57,gt57 CG
  content)

24
GENSCAN predictions

• Gn.Ex Type S .Begin ...End .Len Fr Ph I/Ac Do/T
  CodRg P.... Tscr..
• ----- ---- - ------ ------ ---- -- -- ---- ----
  ----- ----- ------
• 7.00 Prom 63096 63135 40
  -2.75
• 7.01 Init 63183 63274 92 2 2 103 77
  142 0.997 14.61
• 7.02 Intr 63403 63625 223 1 1 83 96
  181 0.999 15.61
• 7.03 Term 64524 64652 129 2 0 101 50
  83 0.373 3.00
• 7.04 PlyA 64758 64763 6
  1.05
• 8.00 Prom 70508 70547 40
  -4.75
• 8.01 Init 70595 70686 92 1 2 103 77
  133 0.990 13.71
• 8.02 Intr 70817 71039 223 2 1 100 96
  217 0.999 20.91
• 8.03 Term 71890 72018 129 0 0 116 43
  119 0.827 7.40
• 8.04 PlyA 72126 72131 6
  1.05
• 9.00 Prom 74399 74438 40
  -8.25
• 9.01 Sngl 76602 76847 246 2 0 71 50
  218 0.886 11.13
• 9.02 PlyA 76928 76933 6
  1.05

25
GENSCAN predicted exons
26
Annotated predicted exons
27
HBB gene

• HBB exons 1-3
• 70545..70686
• 70817..71039
• 71890..72150

• GENSCAN
• 70595 70686
• 70817 71039
• 71890 72018