Journal: eLife

Article Title: Structure and flexibility of the yeast NuA4 histone acetyltransferase complex

doi: 10.7554/eLife.81400

Figure Lengend Snippet: ( A ) Domain map of NuA4 subunits. Modeled regions are marked with a black bar; numbers indicate starting and ending residues. ( B ) Cryo-electron microscopy (cryo-EM) map in red showing the best-defined parts of NuA4. A transparent lower-threshold cryo-EM map is overlaid to show the flexible density likely corresponding to the histone acetyltransferase (HAT)/Trimer Independent of NuA4 involved in Transcription Interactions with Nucleosomes (TINTIN) modules. ( C ) Cartoon representation of NuA4 modules. ( D ) Venn diagram showing NuA4 subunit organization across different complex modules. Subunits in the core attach to Tra1 and act to tether the TINTIN and HAT modules to the complex. ( E ) Cryo-EM map of the NuA4 hub with individual subunits colored. ( F ) Model of the NuA4 hub with individual subunits colored and labeled.

Article Snippet: Discovery screens consisting of 77 nucleosome substrates ( EpiCypher #16-9001 ) were used to test the HAT (50 nM) and TINTIN (50 nM) modules ( ).

Techniques: Electron Microscopy, Cryo-EM Sample Prep, Labeling

Journal: eLife

Article Title: Structure and flexibility of the yeast NuA4 histone acetyltransferase complex

doi: 10.7554/eLife.81400

Figure Lengend Snippet: ( A ) Model for NuA4 complex organization (NuA4 central hub model from the present cryo-electron microscopy (cryo-EM) structure, with the rest of the models from AlphaFold2 prediction) ( ; ). Spatial constraints are imposed on the position of flexible modules by the length of the linker to the connecting amino acids resolved in the structure. Additional low-resolution density adjacent to the NuA4 hub suggests the approximate location of the flexible modules. ( B ) Domain map showcasing the subunits that link the Trimer Independent of NuA4 involved in Transcription Interactions with Nucleosomes (TINTIN), histone acetyltransferase (HAT), and YAF9 modules to the HUB. ( C ) Sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) (Bio-Rad 4–20%) of purified NuA4 TINTIN and HAT modules, stained with InstantBlue (Expedeon). ( D ) dCypher assay results of nucleosome discovery screen for the purified HAT module. Error bars calculate from duplicate experiments, ( E ) dCypher assay result of nucleosome discovery screen for the purified TINTIN module. Error bars calculate from duplicate experiments. Figure 3—source data 1. Uncropped – sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) (Bio-Rad 4–20%) of purified NuA4 Trimer Independent of NuA4 involved in Transcription Interactions with Nucleosomes (TINTIN) and histone acetyltransferase (HAT) modules, stained with InstantBlue (Expedeon). For panel C.

Article Snippet: Discovery screens consisting of 77 nucleosome substrates ( EpiCypher #16-9001 ) were used to test the HAT (50 nM) and TINTIN (50 nM) modules ( ).

Techniques: Electron Microscopy, Cryo-EM Sample Prep, Polyacrylamide Gel Electrophoresis, SDS Page, Purification, Staining

Journal: eLife

Article Title: Structure and flexibility of the yeast NuA4 histone acetyltransferase complex

doi: 10.7554/eLife.81400

Figure Lengend Snippet: AlphaFold2 prediction for the module structures of Trimer Independent of NuA4 involved in Transcription Interactions with Nucleosomes (TINTIN) (left), histone acetyltransferase (HAT) (middle), and Yaf8/Swc4 (right) ( ; ). ( A ) MSA coverage (MSA search and alignment using Jackhammer). ( B ) Predicted alignment error (blue high, red low). Colored bars on top of the graph indicate separate regions of interaction within the modules. ( C ) AlphaFold2 models (ordered regions) with crosslinks mapped (blue <30 Å, red >30 Å) ( ; ). Models colored based on the AlphaFold2 predicted interacting regions (as seen in panel B). ( D ) NuA4 crosslinking position with the given module indicated by cyan spheres .

Article Snippet: Discovery screens consisting of 77 nucleosome substrates ( EpiCypher #16-9001 ) were used to test the HAT (50 nM) and TINTIN (50 nM) modules ( ).

Techniques:

Journal: eLife

Article Title: Structure and flexibility of the yeast NuA4 histone acetyltransferase complex

doi: 10.7554/eLife.81400

Figure Lengend Snippet: Model of NuA4 chromatin localization and histone acetylation. NuA4 is recruited to genomic loci through interaction of Tra1 with site-specific transcription factors. Once recruited, the flexible reader domains interrogate nearby nucleosomes. Nucleosomes bearing the proper chemical marks are preferentially acetylated.

Article Snippet: Discovery screens consisting of 77 nucleosome substrates ( EpiCypher #16-9001 ) were used to test the HAT (50 nM) and TINTIN (50 nM) modules ( ).

Techniques:

Journal: eLife

Article Title: Structure and flexibility of the yeast NuA4 histone acetyltransferase complex

doi: 10.7554/eLife.81400

Figure Lengend Snippet: ( A ) dCypher nucleosome discovery screen-binding profile for purified histone acetyltransferase (HAT) module. Abbreviations for histone modifications are as follows: me, methyl; ac, acetyl; bu, butyryl; cr, crotonyl; ph, phosphoryl; ub, ubiquitin; cit, citrulline. Error bars calculate from duplicate experiments, ( B ) dCypher nucleosome discovery screen-binding profile for purified Trimer Independent of NuA4 involved in Transcription Interactions with Nucleosomes (TINTIN) module. Same abbreviations as panel A. Error bars calculate from duplicate experiments.

Article Snippet: Discovery screens consisting of 77 nucleosome substrates ( EpiCypher #16-9001 ) were used to test the HAT (50 nM) and TINTIN (50 nM) modules ( ).

Techniques: Binding Assay, Purification

Journal: ACS biomaterials science & engineering

Article Title: Chondroitin Sulfate Glycosaminoglycan Matrices Promote Neural Stem Cell Maintenance and Neuroprotection Post-Traumatic Brain Injury

doi: 10.1021/acsbiomaterials.6b00805

Figure Lengend Snippet: List of Immunohistochemical Markers

Article Snippet: 21 , 22 table ft1 table-wrap mode="anchored" t5 caption a7 target antibody vendor dilutions ref neurons NeuN millipore 1:200 21 , 23 neurons NF200 millipore 1:1000 24 astrocytes GFAP dako 1:1000 19 , 22 macrophages CD68 AbD Serotec 1:500 22 NSCs Sox1 1:200 1:200 25 NSCs Nestin Millipore 1:500 19 , 20 proliferating cells Ki67 BD Biosciences 1:800 26 oligodendrocytes Olig2 Millipore 1:250 27 fibroblast growth factor FGF2 R&D Systems 1:200 23 N-acetylgalactosamine (GalNAc) residues linked to CS-GAGs FITC -WFA Vector Laboratories 50 μ g/mL 28 Open in a separate window List of Immunohistochemical Markers Brain sections collected on glass slides, were rinsed in PBS and subsequently incubated in PBS containing 4% paraformaldehyde and 0.4 M sucrose for 30 min.

Techniques: Immunohistochemical staining, Plasmid Preparation

Journal: ACS biomaterials science & engineering

Article Title: Chondroitin Sulfate Glycosaminoglycan Matrices Promote Neural Stem Cell Maintenance and Neuroprotection Post-Traumatic Brain Injury

doi: 10.1021/acsbiomaterials.6b00805

Figure Lengend Snippet: FGF2 presence in brain tissue. (A) Representative images of the region corresponding to the red dotted box in coronal brain sections obtained from sham animals, and surrounding the lesion area in coronal brain sections obtained from TBI only, NSC only, CS-GAG only, and CS-GAG-NSC treated animals. Cellular nuclei are represented by DAPI (blue); CS-GAG and GalNAc presence is indicated by WFA labeling (green); and FGF2 labeling is indicated in magenta. Merged overlays are presented in the topmost panel. A significantly greater FGF2+ area was visualized in brain sections obtained from CS-GAG and CS-GAG-NSC treated animals when compared to sham and TBI only controls, and NSC only treated animals. Statistical significance is represented by ‘*’, which indicates p < 0.05. Scale = 100 μm.

Article Snippet: 21 , 22 table ft1 table-wrap mode="anchored" t5 caption a7 target antibody vendor dilutions ref neurons NeuN millipore 1:200 21 , 23 neurons NF200 millipore 1:1000 24 astrocytes GFAP dako 1:1000 19 , 22 macrophages CD68 AbD Serotec 1:500 22 NSCs Sox1 1:200 1:200 25 NSCs Nestin Millipore 1:500 19 , 20 proliferating cells Ki67 BD Biosciences 1:800 26 oligodendrocytes Olig2 Millipore 1:250 27 fibroblast growth factor FGF2 R&D Systems 1:200 23 N-acetylgalactosamine (GalNAc) residues linked to CS-GAGs FITC -WFA Vector Laboratories 50 μ g/mL 28 Open in a separate window List of Immunohistochemical Markers Brain sections collected on glass slides, were rinsed in PBS and subsequently incubated in PBS containing 4% paraformaldehyde and 0.4 M sucrose for 30 min.

Techniques: Labeling