Matthew Taliaferro

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

20/ We tested this by measuring Arp2/3 buildup at the midbody across our cell lines. Arp2/3 accumulation was lost in Net1 knockouts, restored by the full-length UTR rescue, and NOT restored by the ΔLE rescue, exactly what our model predicted.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

18/ Abscission requires actin to build up at the cut site, and that build-up depends on a protein complex called Arp2/3. One GTPase, Rac1, is known to switch on Arp2/3.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

16/ So maybe a local pileup of Net1 mRNA (and therefore protein) creates a local pool of active GTPase. To test this, we rescued our knockout with a Net1 transgene that still had the LE in its UTR but carried a point mutation that stopped the protein from binding GTPases.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

14/ But what's the mechanism? Does having Net1 mRNA at the midbody actually build up more Net1 protein there? Using our knockout/rescue system: yes! Only Net1 protein made from RNA containing the LE showed up in meaningful amounts near the midbody.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

12/ So where exactly does the cell cycle stall when Net1 RNA can't get to the midbody? Net1 knockout cells linger too long in telophase, the last stage of mitosis, pointing to a problem with abscission itself. The full-length UTR transgene fixed this. The ΔLE version didn't.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

10/ Now we finally knew what to target! We used antisense oligos (ASOs), short synthetic sequences that bind to and block a specific stretch of RNA, against the LE. They blocked Net1 RNA from reaching the midbody. And, unexpectedly, cell division slowed way down! ⏳

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

8/ Okay, but which sequences *within* that 3' UTR are doing the work? To find out, we used a massively parallel reporter assay (MPRA). We tiled ~500 short DNA pieces across the Net1 3' UTR, put each into a reporter RNA, and measured how well each one reached the midbody.

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

6/ Luckily, we can isolate the RNA contents of midbodies and compare them to whole cells. We and others had already found that specific RNAs are shuttled to the midbody. We started with one of the most enriched RNAs, Net1, and asked: how does it get there?

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

4/ Some background: the midbody recruits a series of proteins to carry out abscission. But are RNA molecules recruited too? And if so, does it actually matter? Does having RNA there help the midbody do its job?

Excited to share our newest work! We asked whether RNA molecules need to be in a specific place in the cell to do their job. We found that a single RNA must reach the midbody, the structure that pinches two dividing cells apart, for cell division to finish efficiently. 🧵⬇️

This work was led by an incredible graduate student, Katie Vaeth, and was a truly wonderful collaboration between our lab and cell division expert @prekerislab.bsky.social @cuanschutz.bsky.social www.biorxiv.org/content/10.6...

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

19/ There are a few possible reasons. Perhaps our in vitro structures may differ from those in cells, and/or perhaps important features (like G-quadruplexes) may be invisible to our methods.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

17/ To test the functionality of those structures, we built another pool. We mutated every stem, plus made structure-preserving "compensatory" mutations that should restore the fold.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

15/ Again, shuffling the critical element killed activity. But the support elements tolerated even wide shuffling windows. So they work through bulk nucleotide content, with exact nucleotide order mattering much less.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

13/ Same story. Any mutations in the critical element killed activity. The support element tolerated small mutation windows but not larger ones.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

11/ These critical elements aren't just a quirk of our workhorse neuronal cell line. With Michael Kiebler's lab, single-molecule imaging showed they're also essential for shipping RNA to axons in primary hippocampal neurons.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

9/ We call these "critical elements." Necessary, but not sufficient on their own, which fits the picture that the full, active element is just really big.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

7/ Surprisingly, only pieces keeping 200–250 nt of the element still worked. These localization elements are therefore HUGE, way bigger than the elements that control splicing or translation.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

5/ But there's a puzzle. If you line these elements up across genes, they look nothing alike. So what features within these 260 nt elements are important for their function? Total mystery.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

3/ Some background: Our lab often profiles where RNAs go by physically separating parts of a cell and sequencing the RNA in each fraction. In neurons, we split the cell body (soma) from the projections (neurites) to see which RNAs get trafficked to neurites.

New paper! How do RNAs "know" where to go inside a cell? We dug into the sequence elements that route RNAs to the right place. It turns out that, in mammals, they're surprisingly massive (>200 nt), multipartite, and wonderfully complicated. 🧵

In this time of introspection On the eve of study section I say to my reflection God, please spare me more rejection My reviewers criticize me And my colleagues all despise me Try to put it all behind me But that funding cliff is nipping at my heels