#nucleosome — Public Fediverse posts

RE: https://fediscience.org/@Guillawme/115798148618761432

This article on an ALC1-nucleosome complex with a new conformation was accepted at @ActaCrystD in April and went online today! Check it out!

https://doi.org/10.1107/S2059798326004158

#CryoEM #StructuralBiology #nucleosome #chromatin

RE: https://fediscience.org/@Guillawme/115798148618761432

This article on an ALC1-nucleosome complex with a new conformation was accepted at @ActaCrystD in April and went online today! Check it out!

https://doi.org/10.1107/S2059798326004158

#CryoEM #StructuralBiology #nucleosome #chromatin

One thing I had forgotten to post about this whole story: the fine people running our local cryoEM facility in Uppsala made a 3D print of this ALC1-#nucleosome structure and gifted it to me during the facility's end-of-year social event in December. What a sweet attention!

One thing I had forgotten to post about this whole story: the fine people running our local cryoEM facility in Uppsala made a 3D print of this ALC1-#nucleosome structure and gifted it to me during the facility's end-of-year social event in December. What a sweet attention!

Finished reading this preprint on the ALC1-#nucleosome complex? ⬆️

The methods section details what worked, but not everything that was tried: it would be way too long for the academic article format (this methods section is already quite long).

Well, if you are curious about the thought process and decision making during data analysis, rejoice! For @HRBridges has written it all up in Part 2 of the case study in the CryoSPARC Guide!

Find it here: https://guide.cryosparc.com/processing-data/tutorials-and-case-studies/case-study-processing-of-a-novel-motor-bound-nucleosome-state-empiar-10739-part-2

Discuss it on the forum here: https://discuss.cryosparc.com/t/case-study-exploratory-processing-of-a-motor-bound-nucleosome-empiar-10739-part-2/24438
Or directly here on the Fediverse.

Finished reading this preprint on the ALC1-#nucleosome complex? ⬆️

The methods section details what worked, but not everything that was tried: it would be way too long for the academic article format (this methods section is already quite long).

Well, if you are curious about the thought process and decision making during data analysis, rejoice! For @HRBridges has written it all up in Part 2 of the case study in the CryoSPARC Guide!

Find it here: https://guide.cryosparc.com/processing-data/tutorials-and-case-studies/case-study-processing-of-a-novel-motor-bound-nucleosome-state-empiar-10739-part-2

Discuss it on the forum here: https://discuss.cryosparc.com/t/case-study-exploratory-processing-of-a-motor-bound-nucleosome-empiar-10739-part-2/24438
Or directly here on the Fediverse.

So what does this new structure look like?

ALC1 is loosely bound to the #nucleosome, in a conformation that likely doesn't allow it to slide the nucleosome along the DNA, since the two ATPase lobes are not clamping the DNA. But we finally see the macro domain, for the first time in a nucleosome-bound structure! It interacts with the N-ATPase lobe, preventing the ATPase domain from fully clamping the DNA. So this looks a lot like an intermediate state in the transition from auto-inhibited to active ALC1.

Another striking observation from this structure is a long alpha-helix just upstream of the macro domain, that lines the nucleosome's minor super-groove.

4/5

So what does this new structure look like?

ALC1 is loosely bound to the #nucleosome, in a conformation that likely doesn't allow it to slide the nucleosome along the DNA, since the two ATPase lobes are not clamping the DNA. But we finally see the macro domain, for the first time in a nucleosome-bound structure! It interacts with the N-ATPase lobe, preventing the ATPase domain from fully clamping the DNA. So this looks a lot like an intermediate state in the transition from auto-inhibited to active ALC1.

Another striking observation from this structure is a long alpha-helix just upstream of the macro domain, that lines the nucleosome's minor super-groove.

4/5

As promised, here is a summary of this new preprint on the ALC1-nucleosome complex. 🧵

https://doi.org/10.1101/2025.11.10.687450

First, a reminder of some structural features of the #nucleosome.

Histones H2A and H2B form a negatively charged cleft exposed on the surface of the histone octamer, called the "acidic patch". It is recognized by many chromatin-binding factors. If you have read on these proteins, you have most likely already read about the acidic patch too.

One feature much less commonly mentioned is the "super-groove". The DNA wraps around the histone octamer in such a way that the major and minor grooves perfectly align across the two gyres. In 2004, Dervan, Luger and colleagues synthesized short polyamides able to bind to specific sequences across the super-groove (https://doi.org/10.1073/pnas.0401743101). They hypothesized that some chromatin-binding factors may recognize the super-groove, among other nucleosomal epitopes. Until 2025, there was no direct evidence for this.

1/5

As promised, here is a summary of this new preprint on the ALC1-nucleosome complex. 🧵

https://doi.org/10.1101/2025.11.10.687450

First, a reminder of some structural features of the #nucleosome.

Histones H2A and H2B form a negatively charged cleft exposed on the surface of the histone octamer, called the "acidic patch". It is recognized by many chromatin-binding factors. If you have read on these proteins, you have most likely already read about the acidic patch too.

One feature much less commonly mentioned is the "super-groove". The DNA wraps around the histone octamer in such a way that the major and minor grooves perfectly align across the two gyres. In 2004, Dervan, Luger and colleagues synthesized short polyamides able to bind to specific sequences across the super-groove (https://doi.org/10.1073/pnas.0401743101). They hypothesized that some chromatin-binding factors may recognize the super-groove, among other nucleosomal epitopes. Until 2025, there was no direct evidence for this.

1/5

RE: https://fediscience.org/@Guillawme/111534984107819771

Something really cool happened to me this year!

@HRBridges re-processed a #cryoEM dataset from some previous work of mine and colleagues (publicly available as EMPIAR-10739; see quoted post below for a summary of this work). She significantly improved the results we had struggled to obtain at the time of the initial analysis, and wrote it up as a case study for the CryoSPARC Guide: https://guide.cryosparc.com/processing-data/tutorials-and-case-studies/case-study-end-to-end-and-exploratory-processing-of-a-motor-bound-nucleosome-empiar-10739

This is already super cool! Even cooler: she found a structure we had not found in this dataset. And it is a significant piece of result both to complete our understanding of ALC1 (the protein under study) and more broadly to understand one more way in which chromatin-binding factors can recognize the #nucleosome
We wrote an article about it, the preprint went online in November (now trying to get it peer-reviewed, but it might take some time; I can only work on this in my free time): https://doi.org/10.1101/2025.11.10.687450

I wrote some more about the back story here: https://www.gaullier.org/en/blog/2025/12/28/new-preprint-on-alc1
Long story short: this preprint would not have materialized, had we not both attended the CCP-EM Spring Symposium this year and discussed at the poster session.

I will write a summary thread about this new preprint in the near future (hopefully before the spring semester hits me; it's going to be busy...).

RE: https://fediscience.org/@Guillawme/111534984107819771

Something really cool happened to me this year!

@HRBridges re-processed a #cryoEM dataset from some previous work of mine and colleagues (publicly available as EMPIAR-10739; see quoted post below for a summary of this work). She significantly improved the results we had struggled to obtain at the time of the initial analysis, and wrote it up as a case study for the CryoSPARC Guide: https://guide.cryosparc.com/processing-data/tutorials-and-case-studies/case-study-end-to-end-and-exploratory-processing-of-a-motor-bound-nucleosome-empiar-10739

This is already super cool! Even cooler: she found a structure we had not found in this dataset. And it is a significant piece of result both to complete our understanding of ALC1 (the protein under study) and more broadly to understand one more way in which chromatin-binding factors can recognize the #nucleosome
We wrote an article about it, the preprint went online in November (now trying to get it peer-reviewed, but it might take some time; I can only work on this in my free time): https://doi.org/10.1101/2025.11.10.687450

I wrote some more about the back story here: https://www.gaullier.org/en/blog/2025/12/28/new-preprint-on-alc1
Long story short: this preprint would not have materialized, had we not both attended the CCP-EM Spring Symposium this year and discussed at the poster session.

I will write a summary thread about this new preprint in the near future (hopefully before the spring semester hits me; it's going to be busy...).

#Chromatin fiber's #genomic 'memory' governs the building blocks of life, study reveals.

#transcription #nucleosome

https://phys.org/news/2025-02-chromatin-fiber-genomic-memory-blocks.html

#Chromatin fiber's #genomic 'memory' governs the building blocks of life, study reveals.

#transcription #nucleosome

https://phys.org/news/2025-02-chromatin-fiber-genomic-memory-blocks.html

#Chromatin fiber's #genomic 'memory' governs the building blocks of life, study reveals.

#transcription #nucleosome

https://phys.org/news/2025-02-chromatin-fiber-genomic-memory-blocks.html

Repressed but ready to go: How #bivalency holds developmental #genes in a poised state for #expression.

#histones #H3K27me3 #H3K4me3 #Nucleosome #KAT6B

https://phys.org/news/2025-02-repressed-ready-bivalency-developmental-genes.html

Repressed but ready to go: How #bivalency holds developmental #genes in a poised state for #expression.

#histones #H3K27me3 #H3K4me3 #Nucleosome #KAT6B

https://phys.org/news/2025-02-repressed-ready-bivalency-developmental-genes.html

6/n In addition to the above shown examples for #MolecularMachines, here a few more #molecules we presented in #AugmentedReality at #IdeenExpo: #Tubulin-stathmin complex, #antibody, #nucleosome particle, #SARSCoV2 #polymerase.
#DIYbio #MINT #STEM #chem #AR #education #TiltFive

6/n In addition to the above shown examples for #MolecularMachines, here a few more #molecules we presented in #AugmentedReality at #IdeenExpo: #Tubulin-stathmin complex, #antibody, #nucleosome particle, #SARSCoV2 #polymerase.
#DIYbio #MINT #STEM #chem #AR #education #TiltFive

A #cryoem model of the RSF–NCP complex reveals distinct binding characteristics compared with the SNF2h–NCP complex and biochemical analyses indicate that RSF possesses unique chromatin-remodeling properties #ChromatinRemodeling #RSF #Nucleosome https://t.co/ZP6ZglquKq

Researchers decipher how an #enzyme modifies the #genetic material in the #cell #nucleus.

#chromatin #ISWI #nucleosome

https://phys.org/news/2024-04-decipher-enzyme-genetic-material-cell.html

Our new article has been published in the Biophysical Journal @BiophysJ: "Nucleosome spacing controls chromatin spatial structure and accessibility". We placed a long #nucleosome chain in a box mimicking natural density conditions. We did not see melting of #chromatin fibers at high densities. Accessibility of #chromatin to diffusive processes increases with higher regularity of nucleosome spacing. https://authors.elsevier.com/a/1ijrB1SPT7sCr. #hochschulestralsund #lmumünchen

In the bigger picture of the DNA damage response, this makes a lot of sense: the PARP1- or PARP2-HPF1 complex binds to a DNA break and ADP-ribosylates the histone tails that are most accessible from this location. This modification then activates ALC1 which slides the #nucleosome away from the break, making space for the rest of the double-strand break repair proteins to come in and do their things.

In the bigger picture of the DNA damage response, this makes a lot of sense: the PARP1- or PARP2-HPF1 complex binds to a DNA break and ADP-ribosylates the histone tails that are most accessible from this location. This modification then activates ALC1 which slides the #nucleosome away from the break, making space for the rest of the double-strand break repair proteins to come in and do their things.

In summary, we showed that histone tails closest to a DNA break get ADP-ribosylated more easily, resulting in an asymmetric pattern of ADP-ribosylation on the nucleosome. We then showed that this asymmetry causes ALC1 to slide the #nucleosome in a preferred direction: away from the DNA end that is proximal to the ADP-ribosylated histone.

In summary, we showed that histone tails closest to a DNA break get ADP-ribosylated more easily, resulting in an asymmetric pattern of ADP-ribosylation on the nucleosome. We then showed that this asymmetry causes ALC1 to slide the #nucleosome in a preferred direction: away from the DNA end that is proximal to the ADP-ribosylated histone.

And as the title of the article says, ALC1 is indeed sensitive to this asymmetry! ADP-ribosylation on only one side of the #nucleosome establishes a preferential direction in which ACL1 slides the nucleosome, while symmetric ADP-ribosylation on both sides causes no such bias in sliding direction.

And as the title of the article says, ALC1 is indeed sensitive to this asymmetry! ADP-ribosylation on only one side of the #nucleosome establishes a preferential direction in which ACL1 slides the nucleosome, while symmetric ADP-ribosylation on both sides causes no such bias in sliding direction.

1) We got a much better #cryoEM map than when we had used an enzymatically PARylated #nucleosome resulting in heterogeneous PARylation, for our study published in 2021.

In particular, we now have sufficient resolution to distinguish base pairs and therefore to orient the asymmetric DNA sequence. This provides a definite proof of where ALC1 binds, relative to the side of the nucleosome that is ADP-ribosylated.

1) We got a much better #cryoEM map than when we had used an enzymatically PARylated #nucleosome resulting in heterogeneous PARylation, for our study published in 2021.

In particular, we now have sufficient resolution to distinguish base pairs and therefore to orient the asymmetric DNA sequence. This provides a definite proof of where ALC1 binds, relative to the side of the nucleosome that is ADP-ribosylated.

We devised an experiment to test this hypothesis. We assembled a #nucleosome on an asymmetric DNA with 3 bp on one side of the nucleosome positioning sequence (NPS) and 78 bp on the other side. We also labeled the two H2A-H2B dimers with a different fluorophore. We know which one is on which side because the NPS is asymmetric and always binds an H2A-H2B dimer on one side first.

After enzymatic PARylation, an SDS-PAGE tells us whether one H2A-H2B dimer was more easily PARylated, and which one.

We devised an experiment to test this hypothesis. We assembled a #nucleosome on an asymmetric DNA with 3 bp on one side of the nucleosome positioning sequence (NPS) and 78 bp on the other side. We also labeled the two H2A-H2B dimers with a different fluorophore. We know which one is on which side because the NPS is asymmetric and always binds an H2A-H2B dimer on one side first.

After enzymatic PARylation, an SDS-PAGE tells us whether one H2A-H2B dimer was more easily PARylated, and which one.

From our previous paper on ALC1 in 2021, we were left with the hypothesis that poly-ADP-ribose could be deposited asymmetrically on nucleosomes in response to DNA damage, because not all KS (Lys-Ser) motifs in histone tails would be equally accessible for the PARP1- or PARP2-HPF1 complex bound to the DNA break. This asymmetry would in turn affect in which direction ALC1 slides the #nucleosome and would explain the biphasic saturation curves we got (see: https://fediscience.org/@Guillawme/111535162109035086 ).

From our previous paper on ALC1 in 2021, we were left with the hypothesis that poly-ADP-ribose could be deposited asymmetrically on nucleosomes in response to DNA damage, because not all KS (Lys-Ser) motifs in histone tails would be equally accessible for the PARP1- or PARP2-HPF1 complex bound to the DNA break. This asymmetry would in turn affect in which direction ALC1 slides the #nucleosome and would explain the biphasic saturation curves we got (see: https://fediscience.org/@Guillawme/111535162109035086 ).

To sum up: for maximal activity, ALC1 needs to not only interact with the #nucleosome acidic patch and the H4 tail, like other remodelers do, but also with PAR chains on histones. It also seems like PAR chains are deposited asymmetrically on the nucleosome, which could bias the directionality of nucleosome sliding. This is the hypothesis we addressed in an upcoming new paper! Stay tuned for another summary when it will be published.
Fin.

To sum up: for maximal activity, ALC1 needs to not only interact with the #nucleosome acidic patch and the H4 tail, like other remodelers do, but also with PAR chains on histones. It also seems like PAR chains are deposited asymmetrically on the nucleosome, which could bias the directionality of nucleosome sliding. This is the hypothesis we addressed in an upcoming new paper! Stay tuned for another summary when it will be published.
Fin.

Now, what happens when we perturb the interactions with the H4 tail and the #nucleosome acidic patch? Remodeling is slower at all concentrations of ALC1 tested. By the way, if you think these saturation curves look strange, you're right!

Now, what happens when we perturb the interactions with the H4 tail and the #nucleosome acidic patch? Remodeling is slower at all concentrations of ALC1 tested. By the way, if you think these saturation curves look strange, you're right!

And we see an interaction with the #nucleosome acidic patch. Another thing that Laura https://twitter.com/LauraLe70087690 had already shown, in a study from 2020 I was also involved in. See: https://doi.org/10.1016/j.celrep.2020.108529

And we see an interaction with the #nucleosome acidic patch. Another thing that Laura https://twitter.com/LauraLe70087690 had already shown, in a study from 2020 I was also involved in. See: https://doi.org/10.1016/j.celrep.2020.108529

So far, so good. Now, it would be great to determine a 3D structure of the active complex between ALC1 and a #nucleosome ! This is where it gets hairy. As the name implies, poly-ADP-ribose is a polymer.

The problem is that, when generating this PTM in vitro using PARP1 and HPF1, we have essentially no control over 1) the length, 2) the degree of branching and 3) the site of attachment on histones of the PAR chains (there are multiple known sites of PARylation in histones).

So far, so good. Now, it would be great to determine a 3D structure of the active complex between ALC1 and a #nucleosome ! This is where it gets hairy. As the name implies, poly-ADP-ribose is a polymer.

The problem is that, when generating this PTM in vitro using PARP1 and HPF1, we have essentially no control over 1) the length, 2) the degree of branching and 3) the site of attachment on histones of the PAR chains (there are multiple known sites of PARylation in histones).

We wanted to further understand how ALC1 works. For this, we needed its natural substrate: a PARylated #nucleosome
Turns out one can easily make this in vitro by mixing a nucleosome with PARP1, HPF1 and NAD+.

Then, using a FRET assay, we can measure how fast ALC1 slides the histone octamer in an end-positioned nucleosome away from the short DNA linker.

We wanted to further understand how ALC1 works. For this, we needed its natural substrate: a PARylated #nucleosome
Turns out one can easily make this in vitro by mixing a nucleosome with PARP1, HPF1 and NAD+.

Then, using a FRET assay, we can measure how fast ALC1 slides the histone octamer in an end-positioned nucleosome away from the short DNA linker.

In 2021, we published this paper: Structure and dynamics of the #chromatin remodeler ALC1 bound to a PARylated #nucleosome
https://doi.org/10.7554/eLife.71420

It ended with a hypothesis we addressed in a follow-up paper, which will be out soon, hooray! 🥳
I will post a summary thread of this new paper, but in the meantime, I will repost below the summary of the first paper that I posted on Twitter back then (since I no longer have an account on Twitter).

#cryoEM

In 2021, we published this paper: Structure and dynamics of the #chromatin remodeler ALC1 bound to a PARylated #nucleosome
https://doi.org/10.7554/eLife.71420

It ended with a hypothesis we addressed in a follow-up paper, which will be out soon, hooray! 🥳
I will post a summary thread of this new paper, but in the meantime, I will repost below the summary of the first paper that I posted on Twitter back then (since I no longer have an account on Twitter).

#cryoEM

Extensive multivalent interactions between intrinsically disordered regions in the #BRCA1/#BARD1 #E3 #ligase and #nucleosome substrates facilitate #chromatin recruitment and contribute to histone H2A #ubiquitylation and #DNArepair
Sam Witus, Rachel Klevit, Weixing Zhao et al
https://www.embopress.org/doi/10.15252/embj.2023113565

Extensive multivalent interactions between intrinsically disordered regions in the #BRCA1/#BARD1 #E3 #ligase and #nucleosome substrates facilitate #chromatin recruitment and contribute to histone H2A #ubiquitylation and #DNArepair
Sam Witus, Rachel Klevit, Weixing Zhao et al
https://www.embopress.org/doi/10.15252/embj.2023113565

#3Dprint of #Nucleosome in Complex COMPASS 6UH5 (the Complex of proteins Associated with Set1). Making these protein models feels like being a toy maker than it does a scientist.
@caseysteffen
https://buff.ly/3Xo6Dx3

A new article recommended by #PCIGenomics now published in Peer Community Journal, #genomics section! #Nucleosome patterns in four #plantpathogenicfungi with contrasted genome structures https://peercommunityjournal.org/articles/10.24072/pcjournal.227/ https://doi.org/10.24072/pcjournal.227

#GeneRegulation digest week 2:

🧬DNA methylation atlas of human cells
https://www.nature.com/articles/s41586-022-05580-6

🧬Evolution of #3Dchromatin organization
https://www.sciencedirect.com/science/article/pii/S0959437X22001344

🧬#SMARCA5 (#SNF2H) maintains #nucleosome spacing near CTCF https://www.sciencedirect.com/science/article/pii/S1097276522011753 (in line with our Clarkson et al'2019 https://academic.oup.com/nar/article/47/21/11181/5609524)

🧬Loss of #epigenetic information as a cause of mammalian #aging https://www.sciencedirect.com/science/article/pii/S0092867422015707

🧬#Chromatin assembly of exogenous bacterial #chromosomes in yeast https://www.biorxiv.org/content/10.1101/2022.12.21.520625v2

#GeneRegulation digest week 2:

🧬DNA methylation atlas of human cells
https://www.nature.com/articles/s41586-022-05580-6

🧬Evolution of #3Dchromatin organization
https://www.sciencedirect.com/science/article/pii/S0959437X22001344

🧬#SMARCA5 (#SNF2H) maintains #nucleosome spacing near CTCF https://www.sciencedirect.com/science/article/pii/S1097276522011753 (in line with our Clarkson et al'2019 https://academic.oup.com/nar/article/47/21/11181/5609524)

🧬Loss of #epigenetic information as a cause of mammalian #aging https://www.sciencedirect.com/science/article/pii/S0092867422015707

🧬#Chromatin assembly of exogenous bacterial #chromosomes in yeast https://www.biorxiv.org/content/10.1101/2022.12.21.520625v2

A nucleosome is a structural unit of a eukaryotic chromosome, consisting of a length of #DNA coiled around a core of histones. Explore the x-ray crystallography data and #3Dprint the #Nucleosome COMPASS Complex. #genetics #biotech #CRISP https://buff.ly/3Xo6Dx3

Our new preprint! Nucleosome repositioning in chronic lymphocytic leukaemia (CLL) https://www.biorxiv.org/content/10.1101/2022.12.20.518743v1
▶️Nucleosome changes at cancer-sensitive regions allow patient stratification
▶️Nucleosome repeat length (NRL) decreases with #cancer aggressiveness
▶️Interplay with #methylation, #TFbinding and more!

#CLL, #nucleosome, #leukaemia, #chromatin

Our new preprint! Nucleosome repositioning in chronic lymphocytic leukaemia (CLL) https://www.biorxiv.org/content/10.1101/2022.12.20.518743v1
▶️Nucleosome changes at cancer-sensitive regions allow patient stratification
▶️Nucleosome repeat length (NRL) decreases with #cancer aggressiveness
▶️Interplay with #methylation, #TFbinding and more!

#CLL, #nucleosome, #leukaemia, #chromatin

#SingleCell multi-omics allows functional characterization of #structuralVariants https://www.nature.com/articles/s41587-022-01552-3 "We developed a computational method based on Strand-seq that combines #nucleosome occupancy and structural variation analysis in single cells to identify the functional effects of somatic structural variants in leukemia."

Another talk about long reads: Duncan Sproul on Genome-wide analysis of single-molecule DNA methylation

#Epigenomics22 #epigenetics #nanopore #nucleosome #DNMT

#Introductions
I just moved my account from qoto.org to mastodon.nl. Nothing against the fine folks at Qoto, just a more recognizable address.

My interests are in:
- Epigenetic regulation of stem cells and lineage commitment
- Single cell omics and lineage trajectories of embryonic and heart development
- Gene-regulatory networks of development and differentiation

#devbio #embryo #chromatin #genome #nucleosome #epigenetics

http://www.veenstralab.nl/index.html