Genetic transcription and regulation

Transcription

1 Genetic transcription and regulation

2 Central dogma of biology DNA codes for DNA DNA codes for RNA RNA codes for proteins not surprisingly, many points for regulation of the process

3 DNA codes for DNA replication fork Okazaki fragments nucleotides long in eukaryotes, 10x longer in bacteria! replisome error rate is 1 in bp!

4 DNA codes for RNA Why the need for a messenger? RNA is more flexible additional regulation RNA polymerase Roger Kornberg received the 2006 Nobel Prize (Chemistry) for structures DNA is sequestered in the nucleus in eukaryotes (protected) and... AMPLIFICATION

5 DNA codes for RNA Can get MANY copies simultaneously from the same DNA strand

6 Transcription starts at a promoter promoter is a sequence recognized by the polymerase upstream of the transcription initiation site consensus sequence in bacteria: -10 bps and -35 bps (always just a fraction present though!) RNA polymerase is promiscuous, can bind non-specifically but with lower energy p bound = 1 1+ N NS P eβ ϵ pd PBoC 6.1.2

7 the lac operon operon is just a cluster of genes with one promoter repressor binding site promoter sequences activator binding site PBoC 4.4.1

8 Gene regulation PBoC genes can be regulated by controlling polymerase access to promoter gene repression gene activation about 10% (2600) of our genes code for transcription factors!

9 Gene activation instead of two, now have four distinct states also have three relevant interaction energies (P-DNA, A-DNA, P-A) What is pbound for the promoter? p bound = 1 1+[N NS /P F reg (A)]e β ϵ pd F reg (A) = 1+(A/N NS)e β ϵ ad e β ϵ ap 1+(A/N NS )e β ϵ ad PBoC

10 Transcription factors are also regulated binding of inducers alters factor-dna affinity (lactose induces lac operon expression) lactose One data point is sufficient to fit ΔFloop even the energy of DNA looping affects expression PBoC

11 Genetic switches a genetic switch is a repressor and the gene it controls most interesting are cases where two promoters regulate each other s expression in a feedback loop PBoC

12 Genetic switches p bound (c 1 )= K bc n 1 1+K b c n 1 Hill function - n is cooperativity, i.e., number required for reaction r r(1 p bound (c 1 )) = 1+K b c n 1 r is basal (default) rate in absence of repressor degradation rate basal rate of production dc 1 dt = γc 1 + r 1+K b c n 2 du dt = u + α 1+ν n dc 2 dt = γc 2 + r 1+K b c n 1 due to repression dν dt = ν + α 1+u n PBoC make dimensionless by change of variables

13 Genetic switches du dt = u + dν dt = ν + α 1+ν 2 α 1+u 2 assume n = 2 (clever algebra) steady state (u 2 αu +1)(u 3 + u α) =0 switch-like: concentrations are inversely related (u 2 αu +1) two zeroes for α > 2 ν + 1 u =0 uν =1 (u 3 + u α) PBoC one real zero u = ν not switch-like! (concentrations are always the same)

14 Stability analysis vectors denote (du/dt, dv/dt) α = 1 α = 3 filled - stable unfilled - unstable stable Stable equilibrium for α > 2, observe bistable behavior (two dominant states) unstable stable PBoC

15 Stability analysis u u + δu ν ν + δν d dt ( δu δν ) = A ( δu δν ) A = ( 1 f (ν ) f (u ) 1 - f (x) = nαxn 1 (1 + x n ) 2 ) Solution (sum of exponentials) determined by the eigenvalues of A λ 1,2 = 1 ± f (u )f (ν ) if eigenvalues are negative (positive), solution decays (diverges) over time f (u )f (ν ) < 1 required for stability α = rk 1/n b /γ PBoC α- a combination of basal rate of production, repressor binding constant, and degradation rate - being greater than some critical value determines if switch-like behavior is observed

16 Genetic circuits and clocks Synthetic circuit could be used to, e.g., report on concentrations of multiple chemicals in environment Moon, et al. Genetic programs constructed from layered logic gates in single cells. Nature. 491: (2012). Genetic clock the regulated expression of multiple genes generates ~24h molecular oscillations establishing the circadian rythm Ko, Takahashi. Molecular components of the mammalian circadian clock. Human Mol. Genetics. 15:R (2006).

17 And now for something completely different

18 Radiating genomes of cichlid fish -over 2000 species in just three lakes -500 species in Lake Victoria arose in only 100k years -exhibit a diversity of morphological and ecological traits, e.g., what they eat How did the cichlid fish evolve so quickly? five species have just been sequenced to help answer it Brawand et al. The genomic substrate for adaptive radiation in African cichlid fish. (2014) Nature. 513:

19 How many biologists does it take to sequence a fish?

20 Molecular mechanisms of evolution at work -burst of gene duplication (20% of new genes expressed in a completely new, tissue-specific domain) # of duplications species divergence -accelerated evolution in protein-coding genes (the boring answer), e.g., opsin in the eye and a signaling protein involved in jaw development Brawand et al. The genomic substrate for adaptive radiation in African cichlid fish. (2014) Nature. 513:

21 Molecular mechanisms of evolution at work -high rate of change in gene regulatory elements, which changes how and where certain genes are expressed CNE: conserved non-coding element -high turnover of micrornas (including 40 new genes) for suppressing gene expression to stabilize/refine new expression patterns micrornas: about 22 nucleotides long, interfere with mrna AFTER transcription but BEFORE translation Brawand et al. The genomic substrate for adaptive radiation in African cichlid fish. (2014) Nature. 513:

22 Transposable elements (jumping genes) TE: transposable elements Three waves of TE insertions were detected in each of the cichlid genomes Brawand et al. The genomic substrate for adaptive radiation in African cichlid fish. (2014) Nature. 513:

23 C.D. Jiggins. Evolutionary biology: Radiating genomes. (2014) Nature. 513: Neutral drift or positive selection? The authors attribute the great diversity of changes seen across these genomes to a period of relaxed selection that occurred early in the radiation. During this time, the selective pressures that maintained the stability of the genome were reduced, thereby allowing genetic variation to accumulate and produce subsequent diversification into the lineages we observe today. However, accelerated evolution can result either from neutral evolution due to relaxed selection, or from positive natural selection acting through new selective pressures. Most of the genomic signatures in the paper do not strongly distinguish between these two possibilities. Indeed, it seems most likely that the retention of gene duplicates and rapid genetic divergence were primarily driven by positive natural selection, as species adapted to the great diversity of ecological niches available in the lakes. Subsequent extinction of early lineages could have led to an apparent burst of rapid change on the branch leading to the extant species. There may be no need to invoke a genetic revolution when plain old natural selection can explain the observed patterns.