EMBO/sd-character-level-ner huggingface dataset:
Figure 3Schematic diagrams of RIα regions subjected to in vitro binding assays. Results from the RIα-SPATC1L interaction assay (Fig. 3B) are indicated as + or - at right. D/D, dimerization/docking domain (binding site for AKAPs); Domain A, catalytic subunit binding domain and cAMP binding domain; Domain B, cAMP binding domain; IS, inhibitor sequence (binding site for the active site of the catalytic subunit). Mapping of the SPATC1L binding site on RIα by in vitro binding assay using purified GST-SPATC1L and His-RIα proteins consisting of regions 1-6. Purified GST protein only was used as a control. Binding was confirmed by immunoprecipitation using an anti-SPATC1L antibody followed by immunoblot analyses of RIα, detected using an anti-His antibody. In, input; Sup, supernatant; IP, immunoprecipitated protein. PKA activity after overexpression of SPATC1L or SPATC1LΔC. Graphs show the average PKA activity of Spatc1l-transfected cells (white bar) and Spatc1lΔC-transfected cells (black bar) in the presence or absence of cAMP, expressed relative to control cells (mock) as a percentage. Representative signals observed in the assay are shown above. The overexpression of SPATC1L was confirmed in every experiment by immunoblotting (data not shown). The amount of Cα associated with RIα in the presence or absence of SPATC1LΔC. After IP of RIα in cells overexpressing SPATC1LΔC or control, immunoblot analysis was performed using anti-Cα antibody. Graph shows densitometric analysis of Cα in relation to immunoprecipitated RIα in immunoblot analysis . In, input; IgG, immunoglobulin G from normal rabbit serum.
a–c) Heart-to-body-weight ratios (a), cardiomyocyte diameters (b) and cardiac fibrosis (c) in Sham-operated mice and mice treated with intravenous injection of either scrambled control (Scr) or miR-132 inhibitors (Ant-132) after TAC.f7(a-c) Heart-to-body-weight ratios (a), cardiomyocyte diameters (b) and cardiac fibrosis (c) in Sham-operated mice and mice treated with intravenous injection of either scrambled control (Scr) or miR-132 inhibitors (Ant-132) after TAC. These mice were analysed 3 weeks after TAC (n=4-11). PSR, picrosirius red staining. (d-f) Echocardiography analysis of cardiac dimensions and function in Sham-operated mice and mice treated with intravenous injection of either control (Scr) or miR-132 inhibitors (Ant-132) after TAC. These mice were analysed 3 weeks after TAC for (d) fractional shortening, (e) end-diastolic area and (f) end-systolic area (n=4-9). (g-i) Cardiac FoxO3 protein levels in mice treated with intravenous injection of either control (Scr) or miR-132 inhibitors (Ant-132) 3 weeks after TAC and treatment (n=4-8). M: size marker. All values represent mean±s.e.m. *P0.05; **P0.01; ***P0.005; #P0.05 against TAC-control; ##P0.01 against TAC-control; ###P0.005 against TAC-control. Scale bars, 50 μm. DAPI, 4′,6-diamidino-2-phenylindole; FC, fold change; WGA, wheat germ agglutinin. (g-i) calcineurin activity (h) and Mcip1.4 mRNA levels (i) in mice treated with intravenous injection of either control (Scr) or miR-132 inhibitors (Ant-132) 3 weeks after TAC and treatment (n=4-8). M: size marker. All values represent mean±s.e.m. *P0.05; **P0.01; ***P0.005; #P0.05 against TAC-control; ##P0.01 against TAC-control; ###P0.005 against TAC-control. Scale bars, 50 μm. DAPI, 4′,6-diamidino-2-phenylindole; FC, fold change; WGA, wheat germ agglutinin.
Figure 5A, Co-IP assay of PIF3-TAP with TOC1-GFP vs. 5X-GFP in N. benthamiana. Protein extracts were immunoprecipitated by IgG resins followed by HRV-3C protease digestion. PIF3 and TOC1 were detected by anti-MYC and anti-GFP, respectively. B, Quantification of protein interaction in A. Results from 3 independent trials were averaged. Error bars indicate SEM, asterisks indicate significant difference (*p<0.05, student's t test). C, ChIP-qPCR assay of TOC1 binding to promoter of pre-dawn-phased PIF3 target genes at ZT14 in Col-0, native promoter TOC1 and 5X lines. D, ChIP-qPCR assay of the chromatin residence of native promoter TOC1-GFP at target promoters in toc1 vs. pif3/4/5 toc1 (pif345t1) mutant backgrounds. Data information: 3-d-old SD-grown seedlings were harvested at ZT14, TOC1-GFP was immunoprecipitated by α-GFP and magnetic protein G beads. Data from 3 independent trials were averaged, error bars indicate SEM. Asterisks indicates significant differences (** p<0.001, *** p<0.0001, Student's t test), n.s=not significant.
Fig.Figure 8Under conditions of magnesium starvation, MgtE is upregulated and open, poised to transport Mg2+ inside the cell. If other divalent cations, such as Mn2+ or Zn2+ are present in the environment at relatively high concentrations, MgtE can transport them or be inhibited by them, which causes toxicity. Under these conditions, the N-terminal cytosolic domain of YqgP binds Mn2+ or Zn2+, and activates YqgP for cleavage of MgtE between its transmembrane helices 1 and 2. YqgP also interacts with FtsH, and presents MgtE or its cleavage product(s) for processive proteolysis by FtsH. The cleavage of MgtE by YqgP is required for its full dislocation and degradation by FtsH. The presentation/adaptor function of YqgP is not dependent on its catalytic activity, but it requires its active site unoccupied. YqgP thus behaves also as a pseudoprotease which are common in the rhomboid superfamily. The connection of a rhomboid-like protease/pseudoprotease (YqgP) with a processive degradative membrane-bound protease/dislocase (FtsH) and its role in membrane protein quality control in B. subtilis that we identify here represent a striking analogy to the eukaryotic role of rhomboid-like pseudoproteases Derlins in ER-associated degradation.
EK1-Lipopeptides showed potent inhibitory activity against SARS-CoV-2 infection.Fig. 4.Amino acid sequences of the designed peptides EK1 and EK1C. The dotted lines represent E-K salt-bridge with i to i + 3, or i + 4 arrangement. b. SARS-CoV-2 S protein-mediated cell-cell fusion in the presence of EK1-scramble (I), EK1 (II), EK1C (III), and EK1P (IV) at 2.5 μM (scale bar: 400 μm). c. Inhibitory activity of EK1-scramble, EK1, EK1C and EK1P against SARS-CoV-2 S-mediated cell-cell fusion. d. Design diagram of EK1-lipopeptides with cholesterol modification, including EK1C1-EK1C7. e. Inhibitory activity of EK1-lipopeptides on SARS-CoV-2 S-mediated cell-cell fusion. f. Inhibitory activity of EK1-lipopeptides on SARS-CoV-2 PsV infection. Experiments were repeated twice, and the data are expressed as means ± SD (error bar).
Adipocyte EV transfer proteins involved in FA metabolism to melanoma cells A) Workflow of the SILAC approach.Figure 1Workflow of the SILAC approach. 3T3-F442A cells were seeded and differentiated in the presence of heavy amino acids. After 14 days of differentiation, the EV secreted by the mature labeled adipocytes were isolated and analyzed by mass spectrometry to evaluate the presence of heavy amino acid-containing proteins. These EV were also added to SKMEL28 cells for 12h, then LC-MS/MS analysis was performed to identify heavy amino-acid containing proteins that had been transferred from adipocytes to melanoma cells via EV. Three independent samples (Exp 1 to 3) of EV secreted by labeled 3T3-F442A cells were analyzed by mass spectrometry (in duplicate injections, Inj1/2). The percentage of proteins bearing at least one peptide containing a heavy amino acid is indicated. Proteins involved in FAO and oxidative phosphorylation (OXPHOS) that are transferred from adipocytes to melanoma cells via EV are shown in red.
Identification of human S/T kinases modulating the AAR2-PRP8 protein interactionA.Figure 5A. Schematic of phospho-dependent interaction of the spliceosomal AAR2 and PRP8. AAR2, a spliceosomal assembly protein, and PRP8 from a stable complex, that dissociates upon phosphorylation of AAR2. PPR8 is then free to bind the spliceosomal interaction partner protein BRR2 (Weber et al, 2013; Galej et al, 2013). This regulatory interaction switch is thought to be conserved between yeast and human. A protein kinase that can modulate the dissociation of AAR2 from PRP8 is unknown. B. Superposition of the yeast Aar2 (orange, PDB entry 3SBT; (Weber et al, 2011)) 3D structure and a homology model of human AAR2 derived from HHpred (marine), based on the yeast Aar2p structure (Santos et al, 2015). The inset on the right shows a close-up view of the C-terminal domain, where serines 253 (yeast) and 284 (human) are displayed as sticks. Residues 154-170 and 318-356 of yeast Aar2 as well as residues 1-18, 170-201 and 359-384 of human AAR2 are not shown for clarity. C. Y2H interaction experiment with human AAR2 (FL) as bait and PRP8 (aa1755-2335) as prey. Growth on selective media is abolished when AAR2 carries a S284E phospho-mimicry amino acid substitution. Mutation of second candidate phospho-site, AAR2(T356), does not influence the AAR2-PRP8 protein interaction. D. Interaction of AAR2 and PRP8 in human cells. HA‑tagged PRP8 (aa1755-2335) was co-expressed in HEK293 cells with either wildtype AAR2, AAR2(S284A) or AAR2(S284E) mutant versions. Immunoprecipitation of PRP8 led to a co-precipitation of wild typeAAR2 and phospho-null AAR2(S284A). In contrast, the phospho-mimicry AAR2(S284E) version did not bind PRP8. E. Results of the phospho-Y2H screen to identify kinases modulating the interaction of AAR2 and PRP8. Growth on selective media (SD5), of strains carrying AAR2 bait and PRP8 prey plasmids is reduced through co-expression of either SGK2 or CK2α1(CSNK2A1). Growth reduction is visible in a 1:10 dilution series on selective agar (SD5) in comparison to the control media (SD3). Ctrl: non-related kinases. F. Interaction of AAR2 and PRP8 in human cells in the presences of SGK and CK2α1 kinases. Experiment as in D. Co-expression of wild type SGK2 and CK2α1 reduces the amount of AAR2 co-immunoprecipitated with RPR8.
Figure 8(A) Representative images of histological analysis for GLUT1 protein expression in BT549-derived xenograft tumors. Scale bar, 100 μm.(C) Representative images of histological analyses for GLUT1 and MIG-6 expressions in TNBC patients in low-grade (stage I) and high-grade (stage III) tumors Scale bar, 200 µm. (D) Scatterplot analysis for the correlation between GLUT1 and MIG-6 expression levels (n=85). The r value was calculated by Spearman's rank correlation coefficient analysis. (E) Representative images of the immunofluorescence multiplex assay for colocalization of MIG-6 and HIF1α protein expression in TNBC tumor specimens. The triangle indicates nuclear colocalization, and the star indicates cytoplasmic colocalization. (F) A working model depicts the mechanism by which MIG-6 exerts its tumor-promoting function in TNBC. MIG-6 overexpression promotes HIF1α stabilization and the subsequent upregulation of GLUT1 gene transcription, metabolic reprogramming, and tumorigenesis.
Taxanes induce micronucleation in the human hair follicle matrixA-B) The presence of micronucleated cells in the hair matrix in paclitaxel and docetaxel treated (100 nM, 24h) hair follicles (HF) compared to vehicle is significant (p = <0.Figure 2The presence of micronucleated cells in the hair matrix in paclitaxel (100 nM, 24h) hair follicles (HF) compared to vehicle is significant (p = <0.0001 [****]). Mann Whitney U test performed using N of 12-13 HFs (paclitaxel) from 3 patients. The presence of micronucleated cells in the hair matrix in docetaxel treated (100 nM, 24h) hair follicles (HF) compared to vehicle is significant (p = <0.0001 [****]). Mann Whitney U test performed using N of 8 HFs (docetaxel) from 3 patients. C) Hoechst 33342 staining of healthy cell nuclei comprising the hair matrix (lined) and dermal papilla in untreated (vehicle) human HFs. 20 µm scale. D) Paclitaxel treatment (100 nM, 24h) induces the formation of micronucleated bodies, as visualised by Hoechst 33342 staining (arrows), localising to the proliferative region of the hair matrix. i - 20 µm scale; ii - 10 µm scale. E) 100 nM docetaxel treatment was also seen to promote the formation of micronucleated bodies (arrows). 10 µm scale. Error bars are standard error.
Fig.Figure 1(A) QRT-PCR assay of PAX6, SOX1, NKX2-1, NKX2-2, TBR1 and TBR2 during dorsal (up) and ventral (down) neural differentiation. (B) QRT-PCR assay of SOX1-OT V1 during dorsal (up) and ventral (down) neural differentiation Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (C) QRT-PCR assay of SOX1-OT V1 in control and V1-PAKI cells on day 16 of dorsal neural differentiation, p=0.0059. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (D) Immunostaining assay of PAX6 (green) in control and V1-PAKI cells on day 10 of neural differentiation. Scale bar, 100 μm. (E) QRT-PCR assay of PAX6 and SOX1 in control and V1-PAKI cells on day 16 of dorsal neural differentiation, p=0.0146. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (F and G) Immunostaining assay of PAX6 (green) and SOX1 (red) in control and V1-PAKI cells on day 16 of dorsal neural differentiation (F), and quantiﬁcation of SOX1+ cells (G), p=0.0001. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (H and I) Immunostaining assay of MAP2 (green) in control and V1-PAKI cells on day 35 of dorsal neural differentiation (H), and quantiﬁcation of MAP2+ cells (I), p=0.0001. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (J and K) Immunostaining assay of PAX6 (green) and SOX1 (red) in control and V1-PAKI cells on day 16 of ventral neural differentiation (J), and quantiﬁcation of SOX1+ cells (K), p=0.0003. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test). (L and M) Immunostaining assay of GABA (green) in control and V1-PAKI cells on day 35 of ventral neural differentiation (L), and quantiﬁcation of GABA+ cells (M), p=0.0001. Scale bar, 100 μm. Data information: Data are presented as mean ±SEM from three independent experiments (n=3). *p<0.05, **p<0.01, ***p<0.001 (unpaired two-tailed Student's t-test).
Figure 2Sqr mRNA levels (A, B, C), in cerebrum (A, D), kidneys (B, E, G) and muscle (C, F, H) of Coq9+/+, Coq9R239X and Coq9Q95X mice. SQR protein levels (D, E, F) in cerebrum (A, D), kidneys (B, E, G) and muscle (C, F, H) of Coq9+/+, Coq9R239X and Coq9Q95X mice. Note that SQRwestern-blots were performed in isolated cerebralmitochondria due to the low levels of this protein in cerebrum. In kidneys and muscle the western-blots were performed in tissue homogenates. SQR activity (G, H) in kidneys (B, E, G) and muscle (C, F, H) of Coq9+/+, Coq9R239X and Coq9Q95X mice.
Figure 9(A) Spinal cord sections from 5-month-old WT, Tmem106b-/-, Grn-/- and Tmem106b-/-Grn-/- mice were stained with anti-neurofascin, anti-LAMP1 and anti-CathD antibodies. Representative images from three independent mice were shown. Scale bar = 10 µm. (B) Quantification of LAMP1 positive vacuoles (> 2 µm) for experiments in (A). 4-6 sections/spinal cord from 3 mice were analyzed for each genotype. Data presented as mean ± SEM. Unpaired Student's t test: ****, p<0.0001.
Figure 3(A) Immunoblot of patient fibroblast cell lines (P1 and P2) and three independent controls (C1-3). Immunoblots were probed with the indicated primary antibodies. Quantification of BET1 protein levels normalized to GAPDH. Data represented are mean ± SD of three independent biological experiments. Significant differences of P1 BET1-levels were checked compared to each control (***=p<0.001, one-way-ANOVA followed by Tukey`s post-hoc test). (B) Subcellular localization of BET1 in control and patient fibroblasts by indirect immunofluorescent confocal microscopy. Representative immunofluorescence images of BET1 (red) and cis-Golgi marker GM130 (green). Scale bar: 20µm.Colocalization of BET1 with GM130 was determined using Pearson's correlation coefficient (PCC) measured per cell (right panel; C1: n=63, C2: n=50, C3: n=49 and P2: n=33) (***=p≤0.001, one-way ANOVA was followed by Tukey's post-hoc test). Means are shown ± SEM. (C) Representative immunofluorescence images of BET1 (red) and ER marker PDI (green). Scale bar: 20 µm. Right panel shows PCCs for BET1 versus PDI (C1: n=61, C2: n=42, C3: n=59 and P2: n=32 of two biological replicates). (***=p≤0.001, one-way ANOVA was followed by Tukey's post-hoc test). Means are shown ± SEM. (D) HeLa-cells stably expressing the pC4-reporter were transfected with control siRNA (ctrl) or two independent siRNAs (#1 and #2) directed against BET1 (siRNA #1 is shown in figure EV 2B). 2d post-transfection the pC4-secretion assay was performed for 0 and 10 min post induction of secretion. Representative images are shown (upper panel ctrl and lower panel #2 siRNA, respectively. Scale bar: 20 µm. Right panels represent magnifications of dashed areas. (E) Quantification of colocalization using the Pearson`s correlation coefficient (PCC) of the pC4-construct (GFP) and the cis-Golgi marker GM130 (0 min: ctrl n=63, #1 n=56, #2 n=64; 10 min: ctrl n=104, #1 n=126, #2 n=110). Shown are means ± SEM. Colocalization is significantly reduced by BET1 knockdown (siRNA #1 and #2) compared to control transfected cells. Significant differences of PCC (pC4:GM130) after 10 min induction with solubilizer were analyzed with a one-way-ANOVA followed by Dunnett`s post-hoc test. ***=p≤0.001. 0 min: ctrl 0.265±0.016, #1 0.268±0.017, #2 0.264±0.018; 10 min: ctrl 0.798±0.009, #1 0.726±0.011, #2 0.715±0.011. (F) Quantification of knockdown efficiency of BET1 in HeLa-pC4 cells by immunoblots. Data represented are mean ± SD of three biological replicates. ***=p<0.001 compared to ctrl cells, one-way-ANOVA followed by Dunnett`s post-hoc test).
BAK is ubiquitinated in response to various mitophagy stimuli and by endogenous Parkin, but is compromised in Parkinson's Disease-associated Parkin mutants.Figure 3(A) HeLaParkin cells were treated with CCCP (10 μM), antimycin A and oligomycin (AO) for 2h or GTPP (10 μM) for 6h. Immunoblots are representative of three independent experiments. Graph shows densitometric analysis of non-ubiquitinated proteins relative to untreated control from three independent experiments. Error bars represent mean +/- (B) SH-SY5Y neuroblastoma cells expressing Cas9 together with a sgRNA targeting PRKN (sgPRKN) or a non-targeting control (sgCon) were assessed for Parkin expression by immunoblotting of cytosolic fractions. SH-SY5Y cells generated in (B) were treated with antimycin A and oligomycin (AO) for 3 h prior to cell lysis and immunoblotting. Mono and di-ubiquitinated BAK in Parkin-expressing cells following AO are indicated (asterisk). Data is representative of three independent experiments. (D) Domain architecture of human Parkin with positions of selected PD-associated mutations. Ubl, ubiquitin-like; IBR, in-between-RING HeLa cells expressing wild-type Parkin or the indicated PD-associated Parkin mutants were treated with antimycin A and oligomycin (AO) for 2h. Immunoblots are representative of three independent experiments. Graph shows densitometric analysis of non-ubiquitinated proteins in AO-treated samples relative to untreated control from three independent experiments. Error bars represent mean +/- SD. All Parkin constructs were N-terminally FLAG-tagged except R104W.
Figure 5Jarid2 is present as a full-length protein in multi-potent and undifferentiated cells. Full-length JARID2 with the help of N-terminal PRC2 interacting domain can recruit PRC2 and suppress differentiation genes. Upon differentiation N-terminal of JARID2 is cleaved (ΔN-JARID2) which leads to reduction in PRC2 recruitment and possibly H3K27me3 levels at differentiation genes. However, PRC2 removal is not sufficient for activation of differentiation genes and, in addition, ΔN-JARID2 is required for their up-regulation likely through, directly or indirectly, facilitating transcription factors such as AP-1.
A schematic presentation of various signalling pathways leading to AMPK activation and AMPK‐dependent autophagy.Figure 7A schematic presentation of various signalling pathways leading to AMPK activation and AMPK‐dependent autophagy.
Figure 5A. Domains AAA5s to AAA2L of wild-type and mutant 5 are shown for the ADP-vi state (color) and the AMPPNP state (black). Box: Visualization of inter alpha carbon distances between AMPPNP and ADP-vi state of mutant 5 and wild-type dynein. Black arrows indicate direction of movement when transitioning from the AMPPNP to the ADP-vi state while the size of the arrow indicates the magnitude of movement. All structures are alignment on AAA1L. For wild-type, the structures of human cytoplasmic dynein 2 (ADP-vi state - PDB: 4RH7 and the yeast cytoplasmic dynein (AMPPNP state - PDB: 4W8F were used. For mutant 5 AMPPNP, we used the class 1 structure. B. Same as in (A) but for domains AAA2s to AAA5L. C. Visualization of inter alpha carbon distances between the AAA domains in the AMPPNP and the ADP-vi state of mutant 5 and wild-type dynein for alignments on AAA1L (left) and AAA2L (right). Black arrows indicate direction of movement when transitioning from the AMPPNP to the ADP-vi state while the size of the arrow indicates the magnitude of movement. We removed the linker for clarity. For wild-type, the structures of human cytoplasmic dynein 2 (ADP-vi state - PDB: 4RH7 and the yeast cytoplasmic dynein (AMPPNP state - PDB: 4W8F were used. For mutant 5 AMPPNP, we used the class 1 structure. D. Model for domain movements in the AAA ring of dynein during ATP hydrolysis at AAA1. The AAA ring can be divided into two halves that are connected by two springs. Upon ATP binding / hydrolysis, the gap between AAA1/ AAA2 closes and moves AAA6 / AAA5 which in turn pulls on AAA4 so that the gap between AAA4 / AAA5 closes as well. In addition, this conformational change will pull the buttress and therewith change the stalk registry. E. In the stalk mutant, the spring between AAA4 / AAA5 does not close upon ATP binding / hydrolysis presumably due to a disruption of the stalk / buttress interface. Moreover, the gap at the AAA4 / AAA5 interface is larger for the stalk mutant in both states, AMPPNP and ADP-vi, than for wild-type in the AMPPNP state. This "loose spring" at AAA4 / AAA5 uncouples these domains from the closure of the AAA1 / AAA2 interface, and this accounts for microtubule independent hydrolysis.
a) Overexpression of miRNA precursors from a library identified miRNAs enhancing cardiomyocyte growth and brain natriuretic peptide (BNP) secretion.f1(a) Overexpression of miRNA precursors from a library identified miRNAs enhancing cardiomyocyte growth and brain natriuretic peptide (BNP) secretion. The miRNA family, miR-212/132, is highlighted by red circles. (b,c) Effects of miR-212 and miR-132 precursors and inhibitors (anti) on cardiomyocyte cell size as compared with the effects of scrambled (Scr) controls (n=5-13). Representative images used for quantification of cardiomyocyte cell size are shown in c. (d) Effects of various pro-hypertrophic stimuli on miR-212 and miR-132 expression in neonatal cardiomyocytes (n=6-10). (e) miR-212 and miR-132 expression levels during pressure-induced left ventricular hypertrophy 3 and 21 days after transaortic constriction (TAC) surgery of mice (n=4). (f) Cardiomyocyte diameters after Sham operation or 3, 14 and 35 days after TAC (n=4 per group). (g,h) miR-212 and miR-132 expression (normalized to sno-202 levels) in fractionated cardiomyocytes and cardiac fibroblasts derived from adult mice after 3 and 35 days of TAC. All values represent mean±s.e.m. *P0.05; **P0.01; ***P0.005. Scale bar in c represents 50 μm. ACTN2, α-cardiac actinin; Ang II, angiotensin-2; a.u., arbitrary unit; d, day; DAPI, 4′,6-diamidino-2-phenylindole; FC, fold change; NS, no significant difference; P/I, phenylephrine/isoprenaline.