iiinova — where your digital self keeps evolving

Dave Shechner @shechnerlab.bsky.social · 20/09/2025

AWESOME CONFERENCE ALERT!! I had an absolute blast at the Fusion “Genome Regulation Through RNA” meeting back in 2024, and psyched to see its upcoming return! Let’s geek out about all things Chromatin and RNA together, in Cancun! Talk abstract deadline: 10/17 Details👇

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Dave Shechner @shechnerlab.bsky.social · 30/06/2025

Hello! Ever wonder what's "talking to" your favorite transcript, but were too scared to ask? In our review in @cp-cellreports.bsky.social, @mardakheh.bsky.social and I highlight new RNA-focused tools for discovering RNA interactions across organizational scales. Checkit! tinyurl.com/ydn6e3ac

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Dave Shechner @shechnerlab.bsky.social · 17/12/2024

Aw, yeah! It’s en-dash time, people!

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Dave Shechner @shechnerlab.bsky.social · 30/11/2024

Just updated my phone OS, and apparently the calendar doubles as a karyotype now?

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

And that's a wrap (for now)! I hope you've enjoyed this thread and give our preprint a read. Let us know what you think! (30/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Taken together, our data allow us to propose the first-ever model for the molecular architecture of a single-locus subnuclear compartment. This dramatically expands our understanding of the factory's functional scope, and its architectural disruption in DCM (24/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

This explains some of our observations in the factory transcriptome, since many pre-TTN-proximal transcripts are putative targets of pre-TTN-proximal RBPs. Thus, these RNAs may be recruited via their interactions with factory-resident proteins (23/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

In fact, nearly the entire pool of SAFB (an RBP and "nuclear matrix" factor) and of QKI (a splicing factor that's also mutated in DCM) are localized within TTN RNA factories! (22/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

The factory proteome reveals a remarkable compartment that integrates RNA biogenesis with chromatin compartmentalization. Tons of exciting chromatin regulators (cohesins; SAFBs), RBPs (QKI; SRSF's; ELAVL1) and, surprisingly, DNA repair factors (FEN1; APEX1; MCM's) (21/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Factory localization also correlates with splicing efficiency, as TTN-proximal transcripts are more differentially spliced than other RNAs. Many alternative splicing events are defects (e.g. Retained Introns) specific to KO cells. Almost none are direct RBM20-targets (19/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

We next used O-MAP-Seq to map the TTN RNA factory transcriptome, revealing hundreds of RNAs that localize near nascent TTN in WT and ∆RBM20 cardiomyocytes. Few of these RNAs are encoded in TTN-proximal DNA loci, suggesting that they're localized post-transcriptionally (18/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Surprisingly, in WT cells TID loci bear repressive chromatin marks, and TID-enclosed genes are lowly expressed. Upon RBM20 loss we see a modest, though significant uptick in expression, with downstream effects on the broader transcriptome (17/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Strikingly, upon loss of the RMB20 splicing factor, all of these chromatin interactions disappear. This implies either that RBM20 is required to build the TTN RNA compartment, or alternatively, that RBM20 is needed to keep nascent TTN localized within that compartment. (16/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

With this in hand, we set out to systematically map the chromatin, RNAs, and proteins in the TTN RNA factory. We began by probing the factory's genomic interactions, using O-MAP-ChIP. This revealed scores of Mb-scale domains in both cis and trans. (14/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

To our delight, this yielded exceptionally precise biotinylation in both cell types: just two biotin foci—corresponding to the two alleles of nascent TTN. So, in the context of 2 meters of folded DNA, O-MAP can home in and affinity-tag a single-locus nuclear compartment! (13/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

As a test case, we chose a cardiomyocyte-specific "RNA factory" recently discovered by @berteroale and Chuck Murry. This factory forms around nascent pre-mRNAs encoding the giant protein Titin (TTN), and compartmentalizes the splicing factor RBM20, a key TTN regulator (10/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

We reasoned that we could adapt O-MAP to probe individual nuclear compartments by targeting the nascent transcripts contained therein. But how can we selectively probe these nascent transcripts (and avoid mature RNAs)? By targeting O-MAP to introns! (9/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

O-MAP is a nearly universal tool for RNA-targeted microenvironment-mapping. It uses FISH-like DNA oligos to localize the proximity-biotinylating enzyme HRP to a target RNA. HRP then biotinylates nearby molecules, enabling their discovery (8/30).

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Notably, few tools are available for the targeted biochemical characterization of single loci. If you want to dissect the "neighborhood" around an individual locus—unbiasedly discovering the nearby chromatin domains, RNAs, and proteins—you're in for a challenging time. (5/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

Over the past few decades, our field has developed many powerful tools for probing this compartmental architecture. But there are still core aspects of nuclear organization that remain almost impossible to interrogate by existing technologies. (4/30)

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

This project was driven by the unstoppable Evan Kania (now *Dr.* Kania) and aided by an incredible team of collaborators—including @dschweppe.bsky.social, and @oligopain.bsky.social, @shaoenong, @berteroale, and Chuck Murry. Find the preprint below. Now let's jump it! (2/30) tinyurl.com/yurzfyjp

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Dave Shechner @shechnerlab.bsky.social · 21/11/2024

It's my great pleasure to present the next big preprint from SheqLab! An exciting application of our O-MAP platform that I hope will transform the study of nuclear architecture. If you've ever wanted to dissect the subnuclear "neighborhood" around an individual locus, read on! (1/30)

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Dave Shechner @shechnerlab.bsky.social · 15/11/2024

This is an outrage, I tell you! AN OUTRAGE!!

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Dave Shechner @shechnerlab.bsky.social · 13/11/2024

Meanwhile, basically my whole feed on the muskier platform:

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