Mir Lab

Huge props to our grad student @manyakapoor.bsky.social for developing the simulations and the new MS2 analysis amongst other things! We also thank the transcription community for their invaluable feedback and always welcome further discussion😊

WE ARE HIRING! The lab got the Packard AND an R35! Please spread this PostDoc Ad around! Philly and Upenn are a great place to live and work!

5/n: Please reach out directly to me if you are interested. This is a unique opportunity at a time where increased centralized resources are critical to maintain and accelerate the pace of scientific discovery.

4/n We are seeking a microscope specialist/engineer. The role will include developing and implementing new microscope hardware, control software, customized analysis pipelines as well as collaborating with core users.

3/n We will provide custom built ad hoc optical microscopy solutions, advanced image analysis, and consultation services to researchers across campus and beyond. We will implement technology in response to experimental needs instead of working within the bounds of commercially available technology.

2/n: ACME is co-directed by myself (Mustafa Mir), Melike Lakadamyali @melikel.bsky.social (www.lakadamyali-lab.com), Yihui Shen (www.imaging-systems-metabolism-lab.com), and Andrea Stout @aleestoutphd.bsky.social (www.med.upenn.edu/cdbmicroscopycore)

1/n We have an exciting new initiative at UPenn to provide custom advanced microscopy solutions: Advanced Core for Microscope Engineering (ACME): www.med.upenn.edu/cdbacme/ We are looking for a specialist to help run this new facility, job description is attached please help share widely!

9/ We welcome feedback, questions, and critical discussion from the community. Huge thanks to all our amazing co-authors for their insight, support, and collaboration throughout this work.

8/n: We suggest that enhancers encode the biophysical properties of transcription factor hubs (intensity and stability) through binding affinities and cofactors. Hub properties dictate gene occupancy and transcriptional kinetics, suggesting hubs are active executors of enhancer-encoded regulation.

7/n: Hub intensity and duration correlate with burst amplitude, loading rate, and output suggesting that persistently interacting hubs with more molecules tune gene expression. Consistent with the idea that hubs facilitate transcription factor binding and promote expression.

6/n: Hub presence at a gene is correlated with increased transcription burst amplitude, Pol II loading rate, and total output. Surprisingly, transient interactions between Dorsal hubs and hunchback (lacking Dorsal binding) boosts transcription in ventral nuclei where Dorsal concentration is highest

5/n: We found:
• snail has persistent hub interactions with high-intensity hubs
• sog: has slightly less persistent hub interactions with lower-intensity hubs
• hunchback (negative control): transient interactions without stable hub formation

4/n: Next we examined Dorsal hub interactions at two target genes with distinct Dorsal occupancy and enhancer grammars, snai and sog, and as a negative control examined hunchback which has no dorsal sites.

3/n: First, we find that hubs form rapidly after mitosis and last through interphase. Nuclear hub density doesn't depend on nuclear concentration but hub intensity does, suggesting that hub formation may be independent of concentration, but hubs accumulate more protein with increased concentrations.

2/n: Transcription factor hubs (dense local accumulations) regulate gene expression by promoting the frequency of binding. They are often discussed as having uniform properties based on protein sequence and concentration. We wondered if their biophysical properties vary in a gene dependent manner.

Transcription factor hubs exhibit gene-specific properties that tune expression The spatial and temporal control of gene expression relies on transcription factors binding to and occupying their target sites. Transcription factor hubs - localized, high-concentration microenvironm...

1/n: Excited to share our new preprint. We find that Dorsal transcription factor hubs exhibit gene-specific biophysical properties that tune transcriptional output. Read here: www.biorxiv.org/content/10.1.... Project was led by amazing graduate student Samantha Fallacaro (not on bsky)

Martino Ugolini and I wrote a review on how useful embryos are to understand the role of transcription bodies in gene expression. I am sure
@mirlab.bsky.social, @harrisonflylab.bsky.social, @lennarthilbert.bsky.social and others would agree. Enjoy the read! portlandpress.com/biochemsoctr....

In light of policies to defund biomedical research, and with judicial hearings happening today, we hope to highlight the many ways biomedical research improves people's lives, and to convey how damaging the proposed cuts would be... not just for us researchers, but for everyone.

And all the MS2 interaction analysis was done by an amazing bioengineering graduate student @manyakapoor.bsky.social who just joined Bsky!

Major oversight in my earlier postdoc: This work was led by our wonderful postdoc @mukherja.bsky.social who also built the microscope that enabled these experiments

We can't follow the same cluster but see a shift in cluster kinetics across nuclei. We can't say anything about initiation factors since we are just looking at pol2 and we are testing effects of recruitment vs elongation with the drugs. Will check our language to make sure this is clear. Thanks.

7/n Overall we show that Pol II clusters are functionally labile and suggest that they likely represent collections of molecules engaged at a single gene locus. Please take a read and send us your critical feedback!

6/n Using simultaneous imaging of nascent transcription and Pol II clusters we found that a single cluster remains associated with an active gene during a transcription burst and the intensity of the cluster is highly correlated in real time with the amount of nascent transcription.

5/n Using inhibitors of initiation and elongation we found that cluster formation is dependent on initiation and that transcription elongation leads to a reduction in cluster lifetime.

4/n We found that Pol II clusters undergo a transformation as the zygotic genome is activated. From initially being mostly composed of molecules engaged in transcription initiation to becoming hotspots of elongating molecules.

3/n We used single-molecule tracking and lattice light-sheet microscopy in live Drosophila embryos to ask how clusters change during zygotic genome activation (when there is a ~3x increase in transcription) and how clusters influence transcription levels.

2/n Since their discovery in the 1990's the functional roles and mechanisms of formation of RNAPII clusters have been heavily investigated. Models have ranged from transcription factories containing many genes to condensates that transiently "kiss” active genes.

1/n Excited to share our new preprint! Using live imaging in Drosophila embryos, we show that RNA Pol II clusters switch from sites of initiation to elongation during zygotic genome activation and that they are stably associated with an active gene: www.biorxiv.org/content/10.1...

Big congratulations to our MD/PHD student @driaadigun.bsky.social on receiving a F31 award from NIGMS to "Illuminate the Molecular Mechanisms of Replication and Transcription Coordination" during zygotic genome activation and support her training.

Glad you appreciate this article! Thanks for sharing with others.