Luca Giorgetti lab @FMI
@lucagiorgetti
We study transcriptional regulation and chromosome folding using an interdisciplinary approach combining wet- and dry-lab methods. @fmiscience.bsky.social
6/ In this simple model, enhancer position modulates the rate at which the promoter switches from the low to the high burst regime. Interestingly, this occurs only rarely: from only few times per cell cycle when the enhancer is 5k away, to ~0.1 times per cell cycle when the SCR is ~230 kb away!
5/ The most economic model that explains this behavior is that the promoter can operate in two distinct 2-state (ON/OFF) regimes: 1. a low-burst frequency regime in which it bursts only rarely 2. a high-burst frequency regime to which it occasionally (and temporally) switches:
4/ This means that bursts do not occur as single events but rather in clusters of bursts - and this at all distances we studied, as visible in these binarized burst/no burst transcription site intensity traces:
3/ Time periods between consecutive bursts occur on 3 typical time scales: ~3 min, ~25 min and ~160 min, and their durations are correlated
2/ As a consequence, genomic distance from the enhancer also controls the cell-to-cell variability in the number of bursts produced by the promoter in a given interval of time: the closer the enhancer is to the promoter, the more precise the transcriptional output becomes.
1/ Enhancer location controls promoter burst frequency, but not really burst frequency or size, at least in this ultra-simplified mechanistically interpretable setup. See panel d. here: interburst durations become longer as the enhancer is moved to larger genomic distances :
7/ Our studies thus converge on the idea that long-range enhancer-promoter communication depends on long-lived, cohesin-bridged, loop-extrusion-driven encounters mediated by cohesins that land in specific genomic regions depending on whether or not there are CTCF sites, and where they are located:
6/@elphegenoralab.bsky.social & Leonid Mirny's labs' study now brings this concept one level further and shows that the same mechanisms QUANTITATIVELY predicts for the very first time how CTCF site position and orientation determine an enhancer's effect on a distal promoter! This is REALLY exciting.
5/ Just like in @elphegenoralab.bsky.social and Leonid's study, the hypothesis that long-lived cohesin-driven encounters mediate E-P communication explains how enhancer effect scales with genomic distance (exponentially) and is decoupled from total contact probabilities measured in Hi-C (power-law)
4/ In our vert recent study we showed that extrusion-driven encounters last susbtantially longer than random contacts: www.biorxiv.org/content/10.1... @mattiaubertini.bsky.social @nesslfy.bsky.social
3/ Cohesin bridges in @elphegenoralab.bsky.social & Leonid Mirny's new study identify with the loop-extrusion-driven long-lived encounters that we recently discovered using polymer simulations and super-high resolution live imaging:
17/ ... and nicely predicts @elphegenoralab.bsky.social @karissalhansen.bsky.social's data at the Car2 locus www.biorxiv.org/content/10.1..., using changes in cohesin occupancy and extrusion velocity measured with @gfudenberg.bsky.social in www.biorxiv.org/content/10.1...:
15/ This happens because by selecting longer and longer encounters, one selects more and more loop-extrusion-driven events, the probability of which is exponential as a function of genomic distance!
14/ This simple model also predicts that transcription levels should decrease exponentially as a function of genomic distance between an enhancer and promoter, exactly as we verified using data from our previous work!
13/ Strikingly, this simple hypothesis predicts that: 1) average transcription levels should increase nonlinearly as a function of enhancer-promoter contact probabilities, as we and others previously observed: www.nature.com/articles/s41... and also work from e.g. Boettiger, de Laat, Wysocka labs
8/We then asked if we could detect extrusion-driven encounters in living cells. We resorted to a cell line previously established in the lab by @piamach.bsky.social, allowing to measure the distance between two operator arrays separated by 150kb in cells mESC where RAD21 can be inducibly degraded.
6/ We showed that these encounters only arise when cohesin loads approximately midway between two loci, extrudes them into the encounter radius, and pushes them out after some time.
5/Mattia also discovered that loop extrusion creates a new class of encounters which are extremely rare, but last substantially longer than random polymer collisions. The duration of such encounters depends explicitly on the velocity of extrusion, and it thus a signature of active extrusion!
2/ We started by asking: How often, and for how long, do two genomic sequences meet in the cell nucleus within an arbitrary encounter radius? And how does this depend on the loop extrusion activity of cohesin?
(8/n) This model not only fits the observations on single clones but also correctly predicts the effect of changing the position of the enhancer under the hypothesis that the enhancer only modulates the promoter’s ability to switch from the low- to the high-frequency regime.
(7/n) … in which the promoter stochastically switches between a ‘basal’ low burst-frequency regime and a high burst-frequency regime where clusters of bursts become more frequent
(5/n) We also observed that bursts occur in clusters, with inter-burst duration distributed across three timescales and correlated in time
(4/n) When it is located close to the promoter, the enhancer drives more frequent bursts than when it is located far away, resulting in a more uniform transcriptional output. Thus enhancer location within a cis-regulatory landscape is a major determinant of how accurately a gene is transcribed!
(3/n) We show that enhancer position controls the promoter burst frequency, but not burst duration and size
(2/n) We modified our enhancer mobilization assay (www.nature.com/articles/s41...) to enable MS2-mediated visualization of active transcription sites when the SCR enhancer is hopped and reinserted into different random positions around an ectopic copy of the Sox2 promoter inside a ‘neutral’ TAD.
Celebrating 10 years of our lab with a new preprint: www.biorxiv.org/content/10.1... How does enhancer location within a TAD control transcriptional bursts from a cognate promoter? Experiments by Jana Tünnermann and modelling by Gregory Roth
(8/n) This model not only fits the observations on single clones but also correctly predicts the effect of changing the position of the enhancer under the hypothesis that the enhancer only modulates the promoter’s ability to switch from the low- to the high-frequency regime.
(5/n) We also observed that bursts occur in clusters, with inter-burst duration distributed across three timescales and correlated in time
(4/n) When it is located close to the promoter, the enhancer drives more frequent bursts than when it is located far away, resulting in a more uniform transcriptional output. Thus enhancer location within a cis-regulatory landscape is a major determinant of how accurately a gene is transcribed!