8/ We propose that H3K27me3 mimicry has repeatedly emerged through evolution to restrict the spread of Polycomb domains by breaking the PRC2-H3K27me3 positive feedback loop.

7/ We find that the allosteric regulatory activities of JARID2 and PALI1 synergise to antagonise PRC2 during mouse development and restrict the spread of Polycomb domains in mECS.

6/ Unexpectedly, the H3K27me3 mimicry-defective JARID2 K116R exhibited a homeotic transformation characteristics of Polycomb gain-of-function!

This seemingly goes against the prevalent paradigm where JARID2 K116me3 is believed to positively regulate PRC2.

5/ To determine what is the function of H3K27me3 mimicry, we used CRISPR/Cas to generate 3 knock-in mouse lines: each included a single K-to-R mutation that specifically prevents JARID2, PALI1 or PALI2 from mimicking H3K27me3.

3/ H3K27me3 binds to PRC2. Some proteins mimic H3K27me3 to directly interact with the H3K27me3-binding site in PRC2.

- The Margueron lab showed it for JARID2: pmc.ncbi.nlm.nih.gov/articles/PMC...
- We showed it for PALI1/2: www.nature.com/articles/s41...

1/ H3K27me3 mimicry has repeatedly emerged through evolution, but what's the physiological relevance?

We show that JARID2 and PALI1 mimic H3K27me3 to antagonise PRC2 in vivo and restrict the spread of Polycomb domains.
🧵
www.biorxiv.org/content/10.1...

1/ Baculovirus vector building without PCR!

We updated our MoClo Baculo preprint: www.biorxiv.org/content/10.1...
🧪
Plasmids are available on @addgene.bsky.social, we are working on making them a kit.

Southern eagle ray glides in Black Rock today #Australia🦑

Beaumaris today🦑

Black Rock today

13/ Fun fact #4: Are you using the biGBac system but wish to swap tags more easily? Or want to use MoClo Baculo to build vectors expressing up to 30 proteins? We added an optional compatibility with the biGBac system, which provides all the benefits of MoClo for biGBac users.

12/ Fun fact #3: Intermediate plasmids (Level 1) can be used for protein expression, making MoClo Baculo useful even if you wish to quickly build vectors that express only a single protein.
(Though, in such case, you'd like to sequence these plasmids).

8/ As a bonus, we show that the same CDS parts can also be used to build yeast expression vectors of the same complex (human PRC2 here). This is done using the MoClo Yeast toolkit, with the aid of some new parts that we added for compatibility.

7/ Does it work?
Yes! As a demo, we constructed multi-gene baculovirus expression vectors of the 4- and 5-subunit PRC2 complexes (plasmids size: ~20 kbp).

6/ We then added BV backbones, promoters and terminator parts. We also added some useful affinity tags for protein purification. Users can easily add more tags.

5/ We first modified the part design of the MoClo Yeast toolkit to allow adding/removing N-terminal tags using the same coding sequence part.