Journal: bioRxiv

Article Title: Hydra domain drives SNF2L multimerization and marks ISWI diversification in parasites

doi: 10.1101/2025.09.03.673926

Figure Lengend Snippet: a . Neighbor-joining radial phylogenetic analysis of full-length ISWI proteins from apicomplexan parasites ( T. gondii, N. caninum, E. tenella, P. falciparum, P. vivax, B. microti, C. parvum ), chromerids ( C. velia, V. brassicaformis ), and model eukaryotes ( A. thaliana, H. sapiens, S. cerevisiae ) reveals two distinct apicomplexan-specific clades corresponding to Tg SNF2h- and Tg SNF2L-like proteins. b . Schematic domain organization of human and T. gondii ISWI proteins highlights the presence of a unique insertion in Tg SNF2L, termed Hydra. c . Secondary structures and surface of the domains of Tg SNF2L predicted by AlphaFold v2; C-ter; C-terminal; N-ter, N-terminal. d . Multiple sequence alignment of the Hydra domain from coccidian SNF2L-like ISWI proteins showing conserved residues and predicted secondary structures.

Article Snippet: Recombinant full-length TgSNF2L and TgSNF2LΔHydra were used alongside human SNF2h (HsSNF2h/SMARCA5, EpiCypher, SKU: 15-1024) as a positive control.

Techniques: Sequencing

Journal: bioRxiv

Article Title: Hydra domain drives SNF2L multimerization and marks ISWI diversification in parasites

doi: 10.1101/2025.09.03.673926

Figure Lengend Snippet: a . Recombinant Tg SNF2L and its hydra domain deletion variant (Δhydra) were purified and analyzed by 4-12% NuPAGE, followed by Coomassie blue staining and anti-His tag Western blotting. b . Nucleosome remodeling assay using restriction enzyme accessibility confirms that both full-length and Δhydra recombinant Tg SNF2L retain catalytic activity. Commercial Hs SNF2h (top), recombinant full-length Tg SNF2L (middle), and truncated Tg SNF2L lacking the Hydra domain (bottom) were incubated with EpiDyne nucleosome remodeling substrates. In this assay, remodeling exposes previously occluded GATC sites, enabling cleavage by the restriction enzyme DpnII. The upper band corresponds to intact nucleosomes; the appearance of the lower band indicates successful remodeling. The first lane serves as a -DpnII control, subsequent lanes represent increasing reaction times and the final lane is - ATP control. c . Size-exclusion chromatography coupled with multi-angle light scattering (SEC-MALLS) shows that removing the hydra domain decreases the higher oligomeric forms of Tg SNF2L in the micromolar range. With the loss of the hydra domain, two new forms are detected, corresponding to a Tg SNF2L and Tg SNF2LΔhydra SEC-MALLS (Superose 6 Increase) chromatograms shown as the refractive index curves in blue and orange, respectively. Point measurements of the molecular weight in kDa are displayed as black curves with average masses within the peak regions. d . Mass photometry demonstrates a decrease in tetramer and higher oligomeric forms in the nanomolar range upon hydra domain deletion. The data, shown as normalized counts per molecular weight bin (one representative experiment), compares Tg SNF2L and Tg SNF2LΔhydra in blue and orange, respectively. Monomer, dimer and tetramer peaks are fitted using Gaussian distribution model while higher oligomeric forms are delimited by a dotted line. The relative quantifications of these peaks or windows are shown on the right with the mean and standard deviations shown from duplicate measurements. e . Proposed model: The hydra domain acts as a multimerization module, facilitating Tg SNF2L storage in a functionally primed state. In this model, Tg SNF2L’s multi-oligomeric forms may rapidly release Tg SNF2L and its associated proteins in response to DNA damage or replication fork progression.

Article Snippet: Recombinant full-length TgSNF2L and TgSNF2LΔHydra were used alongside human SNF2h (HsSNF2h/SMARCA5, EpiCypher, SKU: 15-1024) as a positive control.

Techniques: Recombinant, Variant Assay, Purification, Staining, Western Blot, Activity Assay, Incubation, Control, Size-exclusion Chromatography, Multi-Angle Light Scattering, Refractive Index, Molecular Weight

Journal: PLoS Genetics

Article Title: miR-100 Induces Epithelial-Mesenchymal Transition but Suppresses Tumorigenesis, Migration and Invasion

doi: 10.1371/journal.pgen.1004177

Figure Lengend Snippet: (A) Immunoblotting of SMARCA5, HOXA1 and HSP90 in HMLE and MCF7 cells transduced with miR-100. (B) Luciferase activity of the wild-type or mutant HOXA1 3′ UTR reporter gene in 293T cells with ectopic expression of miR-100. (C) Immunoblotting of SMARCA5, E-cadherin and cyclophilin B (CypB) in HMLE cells infected with the SMARCA5 shRNA (shSMARCA5) or the pLKO.1-puro lentiviral vector with a scrambled sequence (Scr) that does not target any mRNA. (D) qPCR of CDH1 in HMLE cells transduced with the control vector (mock), miR-100 alone or in combination with SMARCA5. (E) Immunoblotting of SMARCA5, E-cadherin and HSP90 in HMLE cells transduced with the control vector (mock), miR-100 alone or in combination with SMARCA5. (F) Bisulfite sequencing assay (left panel) and the percentage of CpG methylation (right panel) of the CDH1 promoter in HMLE cells transduced with the control vector (mock), miR-100 alone or in combination with SMARCA5. Open circles: unmethylated CpG sites; solid black circles: methylated CpG sites. Data in (B), (D) and (F) are mean ± SEM, and statistical significance was determined by two-tailed, unpaired Student's t test.

Article Snippet: The human SMARCA5 expression vector was from GeneCopoeia (EX-E2767-Lv105).

Techniques: Western Blot, Transduction, Luciferase, Activity Assay, Mutagenesis, Expressing, Infection, shRNA, Plasmid Preparation, Sequencing, Control, Methylation Sequencing, CpG Methylation Assay, Methylation, Two Tailed Test

Journal: EMBO Reports

Article Title: Expansion of the ISWI chromatin remodeler family with new active complexes

doi: 10.15252/embr.201744011

Figure Lengend Snippet: The expanded family of ISWI chromatin remodelers

Article Snippet: Clonal wells were expanded and tested by Western blotting with antibodies against SMARCA1 (#12483, Cell Signaling Technology), SMARCA5 (NB100‐55310, Novus Biologicals), or tubulin (T9028, Sigma).

Techniques:

Journal: Nucleic Acids Research

Article Title: AURKB-driven dissolution of CIZ1–RNA assemblies from the inactive X chromosome in mitosis

doi: 10.1093/nar/gkag018

Figure Lengend Snippet: CIZ1 C-terminal interaction partners. ( A ) Overview of nuclear protein interaction studies using mammalian cell nuclear extracts from HeLa cells and recombinant GST-tagged hCIZ1 C-terminal fragment C179. Domains coloured as in Fig. . Below, volcano plots showing protein interaction partners identified with high confidence in three independent studies and their relative retrieval by GST-hC179 compared to GST control. Significance (−log 10 FDR q -value) is plotted against log 2 fold change (FC), derived from n = 4 replicates in each case. Significant interaction partners are ≥2-fold more abundant in CIZ1–retrieved samples compared to GST samples, at q -value ≤0.05. Data are given in . ( B ) Venn diagram showing common interaction partners between three independent studies. The 56 core interaction partners are listed in . ( C ) CIZ1 interaction partners in common with 80 Xist interactors identified by CHIRP-MS or iDRIP . Venn diagram indicates those that interact with CIZ1 in all three of our studies. Proteins that were common to both Xist studies and also identified in any of our CIZ1 interaction studies are listed in . ( D ) Simplified STRING diagram showing 56 core CIZ1 interaction partners, clustered using MCL clustering. Three unclustered proteins (dark pink) and one chromatin protein (green) are nuclear matrix-associated proteins . ( E ) Individual abundance (mean of four replicates) across three studies for four nuclear matrix proteins in the core 56 interaction list (SAFB2, SMARCA5, NUMA1, SLTM) and two that appear in at least one of the studies (SAFA, SAFB1). Histograms show fraction of high-confidence peptides in bait and control for each study. ( F ) Example immunofluorescence images of SAFB2 (green) in cycling WT and CIZ1 null PEFs, co-stained for CIZ1-N (red), and DAPI (blue). Box and whisker plots show intensity measures derived from two independent primary cell populations (N) for each genotype. n = number of nuclei measured. Comparison is by t-test, where *** denotes P < .001 and indicates a significant reduction of bound SAFB2 epitope in CIZ1 null cells. ( G ) As in panel (F) but for SAFA, which is not significantly changed in CIZ1 null cells.

Article Snippet: SMARCA5 , Biorbyt orb340809.

Techniques: Recombinant, Control, Derivative Assay, Immunofluorescence, Staining, Whisker Assay, Comparison

Journal: Nucleic Acids Research

Article Title: AURKB-driven dissolution of CIZ1–RNA assemblies from the inactive X chromosome in mitosis

doi: 10.1093/nar/gkag018

Figure Lengend Snippet: Effect of mutation on CIZ1 C-terminal interaction partners. ( A ) Volcano plots displaying protein interaction partners, comparing WT human C179 with C179 DDD or C179 Δtail in independent studies. The majority of proteins were unaffected. Those significantly increased or decreased are highlighted (log 2 fold change ±1, q ≤ 0.05). See also . To the right, STRING diagrams showing functional protein clusters for differentially retrieved proteins. Cluster identities are given in . ( B ) Summary table describing effects of mutations on study-specific interaction partners and core 56 interaction partners (in parentheses). ( C ) Heatmaps showing peptide abundance of the core 56 interaction partners retrieved in control (GST), WT (C179), and mutant reactions, as indicated ( n = 4 replicates). Upper, 10 proteins that are significantly reduced in C179 DDD compared to WT are indicated. Lower, of the core 56 only 5 are significantly changed [see panel (D)]. ( D ) Plot displays log 2 FC of core 56 proteins for C179 DDD (blue) and C179 Δtail (orange), with identities. Ten proteins that are significantly reduced upon phosphomimetic mutation are labelled in blue. Nuclear matrix proteins SAFB2, SLTM, and SMARCA5 are not affected; however, NUMA1 was increased in C179 Δtail (green).

Article Snippet: SMARCA5 , Biorbyt orb340809.

Techniques: Mutagenesis, Functional Assay, Control

Journal: Advanced Science

Article Title: Deciphering the Functional Long Non‐Coding RNAs Derived from MicroRNA Loci

doi: 10.1002/advs.202203987

Figure Lengend Snippet: Molnc‐301b interacts with SMARCA5. A) Fractions of RNAs located in the chromatin, nucleoplasm, and cytoplasm of HSPCs. GAPDH , U1 , and XIST RNAs were used as positive controls for cytoplasm, nucleoplasm, and chromatin location, respectively ( n = 3 replicates). B) Experimental design of the molnc‐301b mechanism study. C) Scatterplots showing DEGs of 301b_wt‐, 301b_m1‐, 301b_m2‐, 301b_del‐overexpressing HSPCs compared with the control. Significantly up‐ or downregulated genes were determined by p <0.05 ( n = 2 replicates). D) GO functional enrichment analysis of coordinately activated and coordinately repressed genes. E) GO functional network showing chromatin‐related terms of molnc‐301b pull‐down nuclear proteins in MS analysis. F) Western blot validation of proteins associated with molnc‐301b in K562 cells. NC refers to proteins pulled down by magnetic beads. G) SMARCA5 RIP assay of K562 cells. Western blot showing SMARCA5 immunoprecipitation (upper panel). The relative fold enrichment of molnc‐301b using SMARCA5 compared with IgG was determined by qPCR analysis (lower panel). SLC25A21‐AS1 and pri‐124 transcripts were used as negative controls. Data represent the mean ± SD ( n = 3 replicates). p ‐Valuess were calculated by unpaired t‐ test. **** p <0.0001, ns, not significant.

Article Snippet: 2 unique shRNA constructs in lentiviral GFP vector of SMARCA5 (TL309248, ORIGENE) were used to package lentivirus and infect K562 cells.

Techniques: Functional Assay, Western Blot, Magnetic Beads, Immunoprecipitation

Journal: Advanced Science

Article Title: Deciphering the Functional Long Non‐Coding RNAs Derived from MicroRNA Loci

doi: 10.1002/advs.202203987

Figure Lengend Snippet: Molnc‐301b acts as an “RNA decoy” by promoting the dissociation of SMARCA5 from chromatin. A) Heatmap showing DEGs of molnc‐301b overexpression (301b_wt, 301b_m1, 301b_m2 and 301b_del) and SMARCA5 knockdown. Significantly up‐ or downregulated genes were determined by p<0.05 ( n = 2 replicates). B) Overlapping DEGs of molnc‐301b overexpression (301b_wt, 301b_m1, 301b_m2 and 301b_del) and SMARCA5 knockdown. The colored boxes and line types denote gene expression events upregulated (red box) or downregulated (blue box) by molnc‐301b (solid line) or SMARCA5 (dashed line). C) Schematic illustration of molnc‐301b promoting the dissociation of SMARCA5 from chromatin and regulating transcription. D) qPCR validation of molnc‐301b in the immunoprecipitation products of ChIRP assay. GAPDH was used as a negative control. E) Scatterplot showing the intersection (red dots, n = 149) between molnc‐301b & SMARCA5 antagonistic (red and blue dots, n = 289) and molnc‐301b‐bound targets revealed by ChIRP‐seq. The overlapping DEGs with log2(fold‐change) of 301_del versus Ctrl and knockdown SMARCA5 shA versus shCtrl are illustrated. F) GO functional enrichment analysis of the molnc‐301b and SMARCA5 co‐targeting genes. G) ChIRP‐qPCR validation of chromatin interaction of molnc‐301b with four erythroid‐associated and another four translation‐associated gene loci. GAPDH locus was used as a negative control. H) SMARCA5 ChIP‐qPCR validation of four erythroid‐associated and four translation‐associated gene loci. I) qPCR validation of four erythroid‐associated and four translation‐associated genes in HSPCs. J) Western blot validation of four erythroid‐associated and four translation‐associated genes. For Figure 5G‐I, data represent the mean ± SD (n = 3 replicates). p‐Valuess were calculated by unpaired t‐test. *p < 0.05, **p <0.01, ***p <0.001, ****p <0.0001, ns, not significant.

Article Snippet: 2 unique shRNA constructs in lentiviral GFP vector of SMARCA5 (TL309248, ORIGENE) were used to package lentivirus and infect K562 cells.

Techniques: Over Expression, Expressing, Immunoprecipitation, Negative Control, Functional Assay, Western Blot

Journal: Advanced Science

Article Title: Deciphering the Functional Long Non‐Coding RNAs Derived from MicroRNA Loci

doi: 10.1002/advs.202203987

Figure Lengend Snippet: Functional molncRNA screening via CRISPR‐based scRNA‐seq. A) Schematic illustration of an integrated approach for CRISPR‐pooled screening of functional molncRNAs with single‐cell transcriptomics. B) Flow cytometric analysis of infected gRNA‐EGFP + /Cas9‐mCherry + K562 cells. MolncRNA (‐) containing more than two pairs of gRNAs for each of the 361 miRNA loci targeted by D1 and D2 gRNAs recognizing the 1–3 kb or 0.5–1 kb downstream region of the miRNA precursor to specifically perturb molncRNA expression. MolncRNA (‐/‐) containing gRNAs for the same 361 target genes, but recognizing the 1–2 kb upstream or 1–3 kb downstream region of the miRNA precursors (U1 and D1 gRNAs) to knock out both molncRNAs and the cognate miRNAs. IC, internal control gRNA. C) Pairwise similarity of transcriptome changes between molncRNA and IC+/IC‐ genes. The similarity of transcriptome changes was calculated as the formula on the lower panel (the method referred to Tian's work [ ⁷⁷ ] ) D) Classification of functional molncRNA gRNAs by comparing with library 1 and library 2. E) Plot displaying a differential expression of erythroid‐associated genes in cells with the indicated functional molncRNA (upper panel, negative molncRNA regulators; lower panel, positive molncRNA regulators) compared to cells with NTC cells. Circle size represents the proportion of cells expressing indicated erythroid differentiation genes. F) Volcano plot showing the DEGs caused by molncRNA or miRNA knockdown. DEGs in red ( p ‐Value < 0.05), or other colors for genes belong to specific GO functional terms labeled in the diagram. G) i – iii . Models of ex‐molncRNA function. i . Linc‐MD1 binds miR‐133 and miR‐135 to act as a competing endogenous RNA (ceRNA) that abolishes miRNA repressing activity on MAML1 and MEF2C and controls muscle differentiation. ii . H19 binds to SAHH and inhibits its activity, thus mediating the demethylation of hematopoietic transcription factors (Runx1 and Spi1) in mouse embryonic hematopoiesis. iii . Molnc‐301b acts as an “RNA decoy” molecule by binding to SMARCA5 to perturb its interaction with chromatin. iv – vi ) Models of in‐molncRNA function. iv ) MIR100HG facilitates the interaction between HuR and its target mRNAs to regulate the cell cycle. v ) LEADeR binds to promoters with an Alu element in the proximity of the interferon regulatory factor (IRF) binding site and negatively regulates prostate basal luminal differentiation. vi ) LncHIFCAR/MIR31HG enhances the recruitment of HIF‐1α and p300 to HIF‐1 target promoters, and activates the HIF‐1 transcriptional network, with the ultimate effect of regulating cancer development.

Article Snippet: 2 unique shRNA constructs in lentiviral GFP vector of SMARCA5 (TL309248, ORIGENE) were used to package lentivirus and infect K562 cells.

Techniques: Functional Assay, CRISPR, Single-cell Transcriptomics, Infection, Expressing, Knock-Out, Labeling, Activity Assay, Binding Assay

Journal: Scientific Reports

Article Title: Altered transcriptional regulatory proteins in glioblastoma and YBX1 as a potential regulator of tumor invasion

doi: 10.1038/s41598-019-47360-9

Figure Lengend Snippet: Immunohistochemistry on tissue microarrays of different grades of astrocytomas ( A ) and MS/MS spectra of representative peptides ( B ) for six selected proteins - NUCKS1, SMARCA5, PARP1, PTBP1, HMGB2 and NFIB. IHC analysis of the proteins tested using commercially available tissue microarrays (US BioMax), confirmed overexpression of these proteins in multiple tumor specimens. The details of tissue microarrays used and IHC procedure are described in the Methods and the staining scoring details are shown in Supplementary Table . MS/MS spectra acquisition is described under Methods.

Article Snippet: Sections were then incubated for 1.5 h at room temperature with primary antibodies - HMGB2 (Abcam, ab11973, 1:200 dilution), PARP1 (Santacruz, sc-8007, 1:100 dilution), NUCKS1 (Abcam, ab84710, 1:100 dilution), SMARCA5 (Atlas Antibodies, HPA008751, 1:200 dilution), NF1B (Abcam, ab186738, 1:150 dilution), PTBP1 (Abcam, ab5642, 1:100 dilution), YBX1 (Abcam, ab12148, 1:500 dilution), EGFR (Atlas antibodies, HPA018530, 1:100 dilution), MAPK1 (Abcam, ab32081, 1:100 dilution), CD44 (BioGenex, AM310-5M, Ready to use), SOX2 (Invitrogen, PA-1-16968, 1:150 dilution), TNC (Cloud-clone corp, MAB975Hu22, 1:100 dilution) and MMP13 (Abcam, ab39012, 1:100 dilution) followed by peroxidase-labeled polymer conjugate to anti-goat, anti-mouse or anti-rabbit immunoglobulins compatible with the primary antibody for 1 h. After washing, sections were incubated for 5–20 min. with the 3, 3 - diaminobenzidine chromogen (DAB) system to develop the stain.

Techniques: Immunohistochemistry, Tandem Mass Spectroscopy, Over Expression, Staining

Journal: Frontiers in Molecular Biosciences

Article Title: Long non-coding RNA lncMGC mediates the expression of TGF-β-induced genes in renal cells via nucleosome remodelers

doi: 10.3389/fmolb.2023.1204124

Figure Lengend Snippet: lncMGC-interacting proteins identified by RNA pull down followed by mass spectrometry. (A) In vitro transcribed RNAs (hlncMGC sense and antisense) were incubated with human HK2 kidney cell lysates and bound proteins isolated and analyzed by mass spectrometry (run in duplicate). 135 proteins were found to interact with sense lncMGC. (B) STRING DB ( https://string-db.org/ ) groups the proteins into RNA processing factors (such as PRPF3 and PTBP1), ribosomal proteins (such as RPL26 and RPL21), and nucleosome remodeling factors. (C) Heatmap of top 10 candidate proteins based on the number of peptides detected by MS (AS1 and AS2 refer to duplicates from antisense; Supplementary S1 and S2 refer to duplicates from sense). (D) RNA immunoprecipitation was used to validate lncMGC binding to the top 10 proteins in HK-2 cells. lncRNA NRON for IQGAP and lncTCF7 for SMARCA5 were used as positive controls ( ; ). Heatmap of the mean of three independent qPCRs is shown. Normalized affinity was calculated as the ratio of lncRNAs/positive control or as the ratio of lncRNAs/lncRNA with the highest affinity to the target protein. (E,F) The structures of SMARCA5 along with histone/DNA are shown in e (from the front) and f (from the side). Protein Data Bank, https://www.ebi.ac.uk/pdbe/entry/pdb/6ne3/analysis#assembly_1 .

Article Snippet: The cross-linked chromatin was sheared and immunoprecipitated with antibodies against H3K27Ac (Abcam, ab4729) and SMARCA5 (Boster Biol.

Techniques: Mass Spectrometry, In Vitro, Incubation, Isolation, RNA Immunoprecipitation, Binding Assay, Positive Control

Journal: Frontiers in Molecular Biosciences

Article Title: Long non-coding RNA lncMGC mediates the expression of TGF-β-induced genes in renal cells via nucleosome remodelers

doi: 10.3389/fmolb.2023.1204124

Figure Lengend Snippet: Differentially expressed candidate genes related to DKD from RNA-seq in WT and lncMGC KO5 MMC. (A) Heatmap (the mean of Log2RPKM from three independent samples) demonstrated genes associated with DKD among the DEGs. The changes in expression of Col4a3, Col4a4, Ucp2, Nox4, Nkd2 , and Pai1 were confirmed by RT-qPCR (B–G) . Effects of siSMARCA5 in WT MMC on the expressions of SMARCA5 (H) , Col4a3 (I) , Col4a4 (J) , Nkd2 (K) , Nox4 (L) , Ucp2 (M) , and Pai1 (N) . Data are shown as the mean of three independent experiments calculated from triplicate qPCRs. One-way ANOVA with post hoc Tukey’s test for multiple comparisons; ±SEM; *, p < 0.05; **, p < 0.01; ***, p < 0.001; and **** p < 0.0001.

Article Snippet: The cross-linked chromatin was sheared and immunoprecipitated with antibodies against H3K27Ac (Abcam, ab4729) and SMARCA5 (Boster Biol.

Techniques: RNA Sequencing, Expressing, Quantitative RT-PCR

Journal: Frontiers in Molecular Biosciences

Article Title: Long non-coding RNA lncMGC mediates the expression of TGF-β-induced genes in renal cells via nucleosome remodelers

doi: 10.3389/fmolb.2023.1204124

Figure Lengend Snippet: Autoregulation of lncMGC expression. (A) H3K27ac at TSS of lncMGC shows clear increase in WT MMC treated with TGF-β but no increase in KO5 MMC under basal or TGF-β-treated conditions. ATAC-seq tracks show a decrease of ATAC-peaks at TSS of lncMGC in KO5 MMC compared to WT MMC in control and TGF-β-treated conditions. (B) Schematic of the genomic region of the promoter of mouse lncMGC/miR-379 cluster depicting Smad and CHOP binding sites and INR (TSS). Arrows indicate positions of PCR primers for Smad and CHOP sites. The arrow (INR) shows the position of INR. (C–H) H3K27ac and SMARCA5 enrichment (ChIP assays) increased by TGF-β in WT MMC at the lncMGC promoter Smad binding site and initiator (INR) site also known as the transcription start site. However, such an increase was not detected in lncMGC-KO5 MMC, suggesting lncMGC regulation of its own promoter. (I–K) Effects of siSMARCA5 on the expression of lncMGC (I) , miR-379 (J) , and H3K27ac at the Smad site of the lncMGC promoter (K) . Data are shown as the mean of three ChIP experiments calculated by triplicate qPCRs each. One-way ANOVA with post hoc Tukey’s test for multiple comparisons; ±SEM; *, p < 0.05; **, p < 0.01; ***, p < 0.001; and **** p < 0.0001.

Article Snippet: The cross-linked chromatin was sheared and immunoprecipitated with antibodies against H3K27Ac (Abcam, ab4729) and SMARCA5 (Boster Biol.

Techniques: Expressing, Control, Binding Assay

Journal: Frontiers in Molecular Biosciences

Article Title: Long non-coding RNA lncMGC mediates the expression of TGF-β-induced genes in renal cells via nucleosome remodelers

doi: 10.3389/fmolb.2023.1204124

Figure Lengend Snippet: Effects of lncMGC deletion and SMARCA5 siRNA (siSMARCA5) on lncMGC neighboring genes. (A) Genomic structure of the Dlk1-lncMGC region and a proposed model of regulation of neighboring genes by lncMGC RNA through SMARCA5. (B–E) Expression levels (RT-qPCR) of lncMGC neighboring genes ( Dlk1, Meg3, Rian, and Mirg ) in KO5 MMC compared to WT MMC. (F) ATAC-seq and H3K27 ChIP-seq tracks in WT and lncMGC KO5 MMC showed changes at the TSS of lncMGC in KO5 MMC compared to WT MMC in control and TGF-β-treated conditions. ATAC-peaks at the TSS of lncMGC adjacent transcripts ( Dlk1, Meg2, and Rian ) are shown in KO5 MMC compared to WT MMC in control conditions and TGF-β-treated conditions. The positions of ATAC-peaks (green) are the same as those of H3K27ac (blue), suggesting those sites are critical for the expression of these genes. (G–J) Effects of siSMARCA5 in WT MMC on Dlk1 (G) , Meg3 (H) , Rian (I) , and Mirg (J) . Data are shown as the mean of three independent experiments calculated from triplicate qPCRs. One-way ANOVA with post hoc Tukey’s test for multiple comparisons; ±SEM; *, p < 0.05; **, p < 0.01; ***, p < 0.001; and **** p < 0.0001.

Article Snippet: The cross-linked chromatin was sheared and immunoprecipitated with antibodies against H3K27Ac (Abcam, ab4729) and SMARCA5 (Boster Biol.

Techniques: Expressing, Quantitative RT-PCR, ChIP-sequencing, Control

Journal: Frontiers in Molecular Biosciences

Article Title: Long non-coding RNA lncMGC mediates the expression of TGF-β-induced genes in renal cells via nucleosome remodelers

doi: 10.3389/fmolb.2023.1204124

Figure Lengend Snippet: Motif analyses and proposed mechanism by which lncMGC and SMARCA5 regulate DKD-related genes by TGF-β. (A–C) Motif analysis (based on differential ATAC-peaks) of WT-TGFb vs. WT-SD Hyper ( p < 0.0001 and enrichment >2) (A) , and KO-TGFb vs. WT-TGFb Hypo ( p < 0.0001 and enrichment >2) (B) and KO-SD vs. WT-SD hypo ( p < 0.0001 and enrichment >2) (C) . (D) Proposed mechanisms of gene regulation by lncMGC mediated by SMARCA5. The promoters of TGF-β-regulated genes are open through Smad and ZF ARID sites even before TGF-β treatment. TGF-β-regulated genes are upregulated by the recruitment of Smad at open chromatin regions. (E) In the absence of lncMGC, the promoters of TGF-β-regulated genes are closed even after TGF-β treatment, and their expression is not increased even with promoter acetylation or TGF-β treatment. Interaction of lncMGC RNA with nucleosome remodelers can enhance the gene expression through the opening of chromatin at the promoter regions of TGF-β-regulated genes.

Article Snippet: The cross-linked chromatin was sheared and immunoprecipitated with antibodies against H3K27Ac (Abcam, ab4729) and SMARCA5 (Boster Biol.

Techniques: Expressing, Gene Expression