Transcription and gene regulation

RNA in a mammalian cell Amount#genes Protein-coding RNA 5%20000 (mRNA) Non-coding RNA95%<1000 (tRNA, rRNA, miRNA)

mRNA expression levels X axis: organ (skin, heart, brain and so on) Y axis: mRNA expression level

Tnnt2 TROPONIN T, CARDIAC MUSCLE ISOFORM Actc1 ACTIN, ALPHA CARDIAC Srf SERUM RESPONSE FACTOR Transcripts per cell Number of genes Rare and abundant transcripts

Expression patterns

Co-expression

How are transcript levels controlled? 1. Synthesis Changes in the chromatin Transcription 2. Decay Degradation by RNases

Accessible and non-accessible chromatin Quantitative chromatin profiling of the CD2 gene (Dorschner et al, Nature Methods)

How is a gene made (in)accessible for transcription? 1. Chemical modifications of histones Acetylation  open configuration Deacetylation  closed configuration 2. Chemical modification of the DNA Methylation  no transcription

RNA polymerase II The enzyme that makes protein-coding transcripts RNA pol II is non-specific in its pure form: ANY DNA  RNA COPY ~40 basal transcription factors are needed to make RNApol II promoter-specific THE DNA OF A GENE  RNA COPY ~2000 transcription factors are needed to regulate the action of RNApol II THE DNA OF THE RIGHT GENE  RNA COPY

Transcription initiation

Gene regulation We want to define … 1.The cis regulatory elements (CREs) 2.The transcription factors that act on the CREs

Studying gene regulation Traditional experiments Global experiments Computational methods

Example 1: a traditional approach Mack and Owens, Circ. Res. 84:852, 1999 LacZ

Angiotensin II ? Transcriptional Regulation of SMC Genes is Dependent on Complex Combinatorial Interactions of Many Cis Elements and Trans Factors Int CArG CArG B CArG A SRF TATA TBP Pol II Complex TCE KLF5 + KLF4 - + TGF  GATA Factors CRP2 PIAS1 EBoxes E12/E47 Myocardin MHOX/Prx1 + ?

Example 2: a modern experimental approach Which are the targets of transcription factor X?

MyoD and myogenin targets in muscle development (Blais et al, Genes Dev 2005)

Example 3: a typical bioinformatic approach Which genes are targeted by factor X? 1. Identify genomic regions likely to contain regulators 2. Identify potential sites for factor X

MEF2 SRF OTHERS Two views on heart development ! Cripps RM, Olson EM. Control of cardiac development by an evolutionarily conserved transcriptional network. Dev Biol Jun 1;246(1): TGIF/MEIS

Conclusions: RNA levels are tightly regulated by transcription and degradation Transcription is a product of chromatin modifications cis regulatory elements transcription factors There is a number of experimental and computational methods

Master’s projects 1. EGFR pathway – towards combinatorial treatment of solid tumours? 2. From stem cell to B cell 3. Evolution of Serum Response Factor regulation

Previous students Erik Larsson Graduate student, Wallenberg Lab / GU Tanya Lobovkina Graduate student, Physical chemistry / Chalmers Nino Demetrashvili

Reading tips About transcriptional regulation ’Genes and Signals’ by M Ptashne ’Genomic regulatory systems’ by E Davidson Some labs doing nice work Transcript exploration: P Kapranov Chromatin: R Young Nucleus biology: P Silver Genome analysis: D Haussler