Transcription: Pausing and Backtracking: Error Correction Mamata Sahoo and Stefan Klumpp Theory and Bio-systems group, Max Planck Institute of Colloids and Interfaces, Potsdam, Germany
Transcription Transcription is the efficient regulatory process in cells,organisms and tissues Control the complex form of gene expression. what happens? The genetic information stored in DNA RNA transcript. How? Transcription RNA polymerase moves along the length of a DNA template by a single base pair per stochastic nucleotide addition creating a complementary RNA.
Transcriptional pausing 1 Heterogeneity in transcription rate Transcription is not continuous interrupted by pausing events. Pauses:RNAP gets halt for times forms inactive configuration. 2 Two general classes of Pauses most frequent. 1 K. Adelman et al., PNAS 99, 13538(2002) 2 I.Artsimovitch et al., PNAS 97, 7090(2000)
Backtracking during transcription: Backtracking pauses 3 Backtracking occurs in three phases. Phase 1: Backtracking Phase 2: Sliding diffusional in nature. Phase 3: Recovery of transcription 3 J.W. Shaevitz, et al., Nature 426, 684(2003)
Questions Addressed? What happens to the transcription pause and backtracking? Pauses have negative effect on transcription High transcription rate requires the pausing events to be suppressed. Backtracking pauses automatically suppressed by the trailing RNAP from behind. However, backtracking is required for the error correction and further recovery of transcription. Making a pause creating an error, Cleaving the transcript Correcting the error. Questions?? What fraction of errors are corrected?? How the efficiency of error correction limited controlled?? How the accuracy can be improved??
Model studied for transcription D D D k c D D 1 k c α p ǫ ǫ ǫ ǫ
Transcription with pausing and backtracking J 0.012 0.01 0.008 0.006 D 1 =0.28, D=0.07 K c =0.07 without Pausing with pause+backtracking P=0.0 P=0.00007 P=0.0007 P=0.007 0.004 0.002 0 0 0.02 0.04 0.06 0.08 0.1 α Both initiation and elongation limited. 4 Low density and maximal current phase. At high transcription intiation rate transcription starts limiting by elongation. Strongly affected by pausing events elongation limited regime. Suppresses with pausing and backtracking. 4 L.B. Shaw, et al., Phys.Rev.E 68, 021910(2003)
D1 Single RNAP transcription: Efficiency of error correction (fec) D D D kc kc p ǫ ǫ ǫ ǫ Efficiency of error correction,fec = For single RNAP transcription,fec = 1 Following the relation,fec = K ca K ca+ǫ 1 (1 a) ; P m=1 KcPm P m=1 KcPm+ǫ 1P m 1 ǫ 1+ P 1 m=1 KcPm a = (1 + Kc 2D ) 2D 1 (4D 2 +Kc 2 +4K c D 4DD 1 ). = (1 + Kc 2D ) Kc 2D (1 + 4D K c ) (ford=d 1 ).
Fec with diffusive rate (D) 1 0.9 0.8 K c =0.07 D 1 =0.28, D=0.007 D 1 =0.28, D=0.07 D 1 =0.28, D=0.2 D 1 =0.28, D=0.4 fec 0.7 0.6 0.5 0.4 0.3 0 0.025 0.05 0.075 0.1 α 0.125 0.15 0.175 0.2 Fec is also both initiation and elongation limited. Increase of D affect strongly in the elongation limited regime. Strong diffusivity suppresses the error correction RNAP spends much time in diffusive manner in any of the backtracked sites.
Fec with backward stepping rate(d 1 ): Single RNAP and Multi-RNAP transcription 1 1 0.9 0.8 0.7 0.6 K c =D/10=0.007 α=0.0 α=1.0 0.9 0.8 0.7 0.6 K c =D=0.07 α=0.0 α=1.0 fec 0.5 fec 0.5 0.4 0.4 0.3 0.3 0.2 0.2 0.1 0.1 0 0 0.1 0.2 0.3 0.4 0.5 0.6 D 1 0 0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4 0.45 0.5 0.55 0.6 0.65 D 1 Fec in multi-rnap transcription is always reduced comparatively with single-rnap transcription Lack of free spaces that restricts diffusion of backtracked RNAP. The difference is strongly affected for higherd 1 regime. Further increase ofk c reduces the difference between both cases Push back effect of the trailing RNAP from behind in the multi-rnap transcription.
Fec with the cleavage rate(k c ):Single-RNAP and Multi-RNAP transcription 1 1 0.8 0.6 D=D 1 =0.07 α=0.0 α=1.0 0.8 0.6 D 1 =0.28, D=0.07 α=0.0 α=1.0 fec fec 0.4 0.4 0.2 0.2 0 0 0.1 0.2 0.3 0.4 0.5 0.6 K c 0 0 0.1 0.2 0.3 0.4 0.5 0.6 K c Fec for single-rnap transcription is always above the fec for multi-rnap transcription Available free spaces for error correction. Fec for multi-rnap transcription is always reduced Dense traffic effect. Error correction in multi-rnap case is improved for higher K c. Further improvement is achieved with increase ind 1.
Fec with both cleavage rate(k c ) and backward stepping rate(d 1 ) D 1 0.1 α=0.0 0.1 α=1.0 0.01 0.01 0.1 0.01 0.01 0.1 K c Fec is strongly controlled both byd 1 andk c. Error correction Strongly improved increasing both by backward stepping rate,d 1 and cleavage rate,k c.
Fec with distance(l) between an active RNAP and a paused RNAP 1 0.9 0.8 0.7 Simulation Analytical 0.6 fec 0.5 0.4 D1=D=K c =0.07 0.3 0.2 0.1 0 0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 L fec(l) =fec single max {1 exp( (L/L 0 ))} Approximation:L 0 = ǫ K c. Gap distribution,p(l) = ( α ǫ )(ǫ α ǫ ) L. fec increases with the distance: More free space available for error correction. Larger gap size Better error correction.
Efficiency of error correction:multi-rnap transcription fec 1 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 D=D 1 =K c =0.07 α c =0.04 α c =0.08 Simulation Analytical 0.1 0 0 0.025 0.05 0.075 0.1 0.125 0.15 0.175 0.2 0.225 0.25 0.275 α Analytical results valid for low value of α Semianalytical. The deviation starts from the crictical value,α c =0.04 where the density starts saturating. Beyond α c, the error correction may depend on other parameters.
Summary Transcription rate suppressed both by pausing and backtracking (reduced saturated density effect). We exactly calculate the efficiency of error correction for a single-rnap and multi-rnap transcription in a semi-analytical way. Error correction can be strongly improved by increasing both the backward stepping rate and the transcript cleavage rate.
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