The Lewis Lab

We are excited to share our new preprint demonstrating that nucleic acid interactions with SUZ12 constrain PRC2 activity, establishing a kinetic buffer essential for targeted gene silencing and revealing vulnerabilities in diffuse midline gliomas. tinyurl.com/4yrwftrn

This paper is wild. After 3 rounds of directed evolution, they converted a DNA polymerase into an enzyme that can do: - RNA synthesis - Reverse transcription - Synthesis of "unnatural" nucleotides - Synthesis of DNA-RNA chimeras One of the best papers I’ve read recently. For context: In nature, it is DNA polymerase that takes a DNA sequence as a template and then copies it. These enzymes are crucial in replicating the genome for cell division, and they are EXTREMELY specific for DNA over RNA. This is key because RNA nucleotides are present in the cell at concentrations ~100x higher than DNA nucleotides, so the enzyme has evolved clever strategies to select one over the other. RNA polymerases, for comparison, are the enzymes that take a DNA sequence as template and then convert it into RNA. They are involved in gene expression, for example. To convert a DNA polymerase into an RNA polymerase (and all the other functions I mentioned earlier), the authors did a fairly straightforward directed evolution experiment. First, they took four DNA polymerase enzymes belonging to various archaea. These DNA polymerases don’t check for DNA vs. RNA as stringently as other types of cells, so they’re a good starting point to evolve RNA polymerases. The authors inserted some targeted mutations into these enzymes, based on known mutations in the literature. For example, they swapped the amino acid at position 409 for a smaller amino acid, thus removing a “gate” that keeps RNA building blocks from entering the enzyme. Next, they took the four genes encoding these DNA polymerases and cut them up into 12 segments each. They randomly stitched these 12 segments together — from the four different genes — to build millions of unique variants. Each shuffled gene was inserted into an E. coli cell. Then, they grew up these cells (each carrying a unique polymerase) and put them into microfluidic droplets. A device isolates each droplet, lyses the cell open, and releases the polymerase. The droplet also contains RNA building blocks and a DNA template, encoding a fluorescent reporter. If the polymerase begins synthesizing RNA, it will produce a detectable signal. They screened about 100 million droplets in 10 hours of work, searching for those with a signal. For each well that yields a fluorescent signal, the researchers isolated the DNA and sequenced it to figure out which polymerase it was. They repeated this 3x times, finally isolating a really excellent RNA polymerase variant which they called "C28." C28 has 39 mutations compared to the wildtype enzymes. It incorporates about 3.3 nucleotides of RNA per second, with 99.8% fidelity. The crazy thing is that this enzyme can also copy DNA or RNA templates back into DNA (reverse transcription), or use chimeric DNA-RNA molecules as a template and amplify them. It is just a super versatile polymerase that can act on DNA, RNA, or modified nucleotides, to build just about anything.

Stress controls epigenetic inheritance! A histone ubiquitylation-based regulatory hub links stress/environmental signaling to heterochromatin self-propagation and epigenetic inheritance-reshaping how we think about development, drug resistance and cancer👉nature.com/articles/s4158…

New online! Cell-cycle-dependent repression of histone gene transcription by histone H4 bit.ly/49lpAYX

Sam Krabbenhoft, @thePeterLewis & @harrisonflylab use fruit flies to study pediatric diffuse midline glioma. A new #GENETICS study identifies genetic pathways that could guide targeted therapies—showing the power of model organisms in cancer research: buff.ly/FwSPt8X

遺伝子スケールのクロマチン物理を研究するため、ヒストン修飾を制御した96-mer (20 kb) の再構成系を確立しました。1分子観察・原子間力顕微鏡・in vitro Hi-Cにより特定の修飾パターンが構造や動態をどう変えるか調べています。論文のビジュ結構いいのでぜひ見てください！ science.org/doi/10.1126/sc…

Mechanisms of DNMT3A–3L-mediated de novo DNA methylation on chromatin nature.com/articles/s4159…

1/ 🚀 AEBP2 isn’t what we thought. You were told that AEBP2 promotes PRC2 activity on chromatin. We found the opposite: the most prevalent AEBP2 isoform inhibits PRC2 activity. Excited to share our new paper surl.li/cgwqcq A thread 🧵

Happy to share our latest work on Polycomb repression reprogramming in early development, spearheaded by two super talented Ph.D students Yitian Zeng and Feng Kong. cell.com/cell-stem-cell…

Our work "Aquarius helicase facilitates HIV-1 integration into R-loop enriched genomic regions," is out in @NatureMicrobiol nature.com/articles/s4156…

Really excited that our manuscript on Synovial Sarcoma is now online. Link to Open Access PDF: tinyurl.com/55bnssws A summary🧵 :

@wendywenderski Thanks, Wendy! We really appreciate your note and are happy you enjoyed the study.

Solid study from the Lewis lab dissecting PRC2 protein domain functions and particularly the role of SUZ12 NBD in constraining polycomb domains to their appropriate sites within the genome.

⁦@BeatrizGerFalc⁩ summarizing some of her latest work with the lab. #EZH2 controls cell plasticity in #ProstateCancer through coordination with activation of #translation and the RNA degrading protein #tristetraprolin ⁦⁦@CPDR_Labs⁩

Today in @Nature we share our back-to-back stories with @nzhenguw revealing chemical-genetic convergence between a molecular glue & E3 ligase cancer mutations. 1/6

The summer has been busy! Along with the move and a PhD defense, we have updated two preprints: biorxiv.org/content/10.110… and biorxiv.org/content/10.110…

Hypertranscription state in early embryos that both shapes and is fostered by the three-dimensional genome organization, revealing an intimate interplay between chromatin structure and transcription @Nature @xielablife nature.com/articles/s4158…

Online Now: Distinct specificity and functions of PRC2 subcomplexes in human stem cells and cardiac differentiation dlvr.it/TMRwWj

VEFS: the NBD is gone. RNA binding is rare (small red bubble). Most PRC2 is RNA-free (large green bubble), catalytically active, and binds both CpG and non-CpG sites.
 Result: diffuse H3K27me3 that pulls PRC1 away from its normal targets, disrupting gene silencing.

PRC2 silences genes by adding H3K27me3 at the right spots in the genome. Our new work shows how the nucleic acid-binding domain (NBD) of SUZ12 keeps this activity under control, and what happens when you remove it.