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1 DOI: /ncb2535 Figure S1 SOX10 is expressed in human giant congenital nevi and its expression in human melanoma samples suggests that SOX10 functions in a MITF-independent manner. a, b, Representative pictures of immunostaining for SOX10 in 3 subgroups of human congenital nevi (group I, II and III). 17 samples of giant congenital nevi from different human patients were divided into three subgroups based on the size of the congenital nevi: group I (1.5 to 10 cm), group II (11 to 20 cm) and group III (21 to 40 cm). Note the expression of SOX10 in differentiated melanocytes residing in the epidermis, while epithelial cells of the epidermis do not express SOX10 (epidermal-dermal border is demarcated by dashed lines). c, d, Expression analysis of SOX10 and MITF in human melanomas. TMA were scored based on the percentage of SOX10- positive and MITF-positive nuclear staining in melanoma cells and samples were divided into four groups with expression in less than 10% of tumor cells (group I), from 10 to 50 % (group II), from 50 to 90% (group III), and in more than 90% (group IV) of all tumor cells. Insets show high magnification views. c, Analysis of SOX10 and MITF expression in primary human melanomas. Two representative tumors immunostained for SOX10 and MITF from TMA of primary melanoma samples demonstrating that in 11% of cases SOX10 is expressed at grade IV, while MITF expression is evaluated as grade I (n=39 tumors). d, Analysis of SOX10 and MITF expression in metastatic TMA showing that SOX10 expression does not correlate to MITF expression in 16% of all cases analyzed (n=119 tumors). These data demonstrate that SOX10 might have a MITF-independent function in a subset of human melanomas. M, melanocytes, TMA, tissue microarray, BF, bright field; Scale bars, 50 mm. 1
2 Figure S2 Sox10 haploinsufficiency prevents skin hyperpigmentation, expansion of neoplastic cells, and melanoma formation in Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ mice. a, Pictures of Tyr::Nras Q61K INK4a -/- and Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ littermate mice at different time points. At postnatal day 12 (a, left panels) and at 5 months of age (a, right panels), Sox10 haploinsufficiency counteracts hyperpigmentation of paws and snout in Tyr::Nras Q61K INK4a -/- mice. Moreover, also in Tyr::Nras Q61K INK4a - /- mice, Sox10 haploinsufficiency resulted in formation of a belly spot (a, lower left panel). Note that Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ mice have pigmented hairs indicating normal melanocyte stem cell function. Scale bars, 5 mm. b, Pictures of Tyr::Nras Q61K INK4a -/- and Tyr::Nras Q61K INK4a - /- Sox10 LacZ/+ mice showing two examples of Tyr::Nras Q61K INK4a -/- mice with melanoma on the back skin (melanoma is demarcated by dashed lines) and their Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ littermates displaying no signs of melanoma. Please note that there is no hair greying in mice of both genotypes. Scale bars, 5 mm. c, S100 immunohistochemistry of skin sections from Tyr::Nras Q61K INK4a -/- and Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ mice. To decrease hyperpigmentation in the dermis, sections were bleached. Note rare example of few S100-positive cells associated with HFs in Tyr::Nras Q61K INK4a -/- Sox10 LacZ/+ mice and the presence of S100-positive glial cells in close proximity to the SG in both genotypes. Two examples for each genotype are shown. Scale bars, 50 mm. d, Immunostaining for Dct (green) in skin sections from Tyr::Nras Q61K and Tyr::Nras Q61K Sox10 LacZ/+ mice at postnatal day 9. Note the presence of multiple ectopic Dctexpressing cells in Tyr::Nras Q61K mice (indicated by arrows). Strikingly, Sox10 haploinsufficiency completely abolishes the presence of these Dct-positive cells in the dermis. Scale bars, 20 mm. HF, hair follicle, SG, sebaceous gland. 2
3 Figure S3 Characterization of SOX10 expression upon SOX10 knockdown in vitro in human melanoma cell lines. a, Western blot analysis of SOX10 expression in M and A375 human melanoma cell lines upon SOX10 knockdown after 48 and 96 hours showing that SOX10 levels are efficiently reduced as compared to the control samples. For SOX10 knockdown, shrna expression plasmids based on psuper-neo-gfp vectors (SOX10 shrna; scrambled (scr) shrna used as a control) were electroporated into human melanoma cell lines and samples were collected after 48 and 96 hours. b, c, Immunostaining for SOX10 (red) combined with GFP signal showing that in cells electroporated with scr shrna all GFP-positive cells express SOX10, while GFP-positive cells electroporated with SOX10 shrna show no SOX10 expression. Representative examples from A375 melanoma cell line (b) and from M cell line (c) are shown. Scale bars, 20 mm. Uncropped images of blots are shown in Supplementary Fig. S6. 3
4 Figure S4 SOX10 knockdown in human melanoma cell lines leads to decreased expression of melanocytic genes, increase of mesectodermal differentiation, increased apoptosis and deregulation of cell cycle. a, Venn diagram depicting the number of differentially upregulated or downregulated genes in human melanoma M cell line upon SOX10 knockdown after 48 and 96 hours. b, mrna expression of genes of melanocytic lineage after 96 hours of SOX10 knockdown. n=3 independent experiments, each carried out in triplicate. c, mrna expression of mesectodermal lineage genes after 96 hours of SOX10 knockdown. n=2 independent experiments, each carried out in triplicate. d-f, mrna expression of genes involved in the regulation of cell cycle in M cell line after 48 and 96 hours of SOX10 knockdown. n=3 independent experiments, each carried out in triplicate. Data represent the fold change as mean ± s.d. and are normalized to the control. 4
5 Figure S5 Knockdown of SOX10 induces cell cycle arrest and leads to increased apoptosis in human melanoma cells. a-b, FACS analysis of apoptosis in A375 cell line upon SOX10 knockdown. For apoptosis analysis, GFP-positive cells were gated (a) and Annexin V-positive cells were analyzed after 48 hours (b, left panel) and 96 hours (b, right panel) of SOX10 knockdown. c, d, Immunostaining for activated caspase-3 and quantification of activated caspase-3-positive cells in M cell line upon SOX10 knockdown after 96 hours. Percentage of caspase-3-positive cells per DAPI-positive nuclei was counted (n=400 DAPI-positive nuclei in each independent experiment). n=3 independent experiments. Data are presented as mean ± s.d. These data show that SOX10 knockdown influences survival of melanoma cells. e-g, FACS analysis of cell cycle in the A375 cell line upon SOX10 knockdown. GFP-positive cells were gated (e) and subjected further to PI selection gating. Resulting FACS profiles of A375 control and A375 SOX10 shrna cells at 48 hrs (f) and at 96 hrs (g) are presented. h, Quantification of cell cycle phase distribution. Percentage of cells in G1/G0 and S+G2 phases in M and A375 cell lines upon SOX10 knockdown after 48 and 96 hours are presented. n=3 independent experiments. Note that in control A375 cells, more cells are in S+G2 phase than in G1/ G0 phase. Data are presented as mean ± s.d. The data show that SOX10 knockdown results in cell cycle arrest in melanoma cells. hrs, hours, Scale bars, 50 mm. 5
6 Figure S6 Full length blots from Supplementary Fig. 3a. 6
7 Table S1 Clinical data of human patients diagnosed with giant congenital melanocytic nevi. 17 samples of giant congenital nevi from different human patients were divided into three subgroups based on the size of the congenital nevi: group I (1.5 to 10 cm), group II (11 to 20 cm) and group III (21 to 40 cm) and analyzed for SOX10 expression. CMN, congenital melanocytic nevus, f, female, m, male. 7
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DOI:.38/ncb3399 a b c d FSP DAPI 5mm mm 5mm 5mm e Correspond to melanoma in-situ Figure a DCT FSP- f MITF mm mm MlanaA melanoma in-situ DCT 5mm FSP- mm mm mm mm mm g melanoma in-situ MITF MlanaA mm mm
Supplementary Information
Supplementary Information Figure S1: Follicular melanocytes in the wound peripheral area migrate to the epidermis in response to wounding stimuli. Dorsal skin of Trp2-LacZ mice stained with X-gal and analyzed
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DOI: 10.1038/ncb2607 Figure S1 Elf5 loss promotes EMT in mammary epithelium while Elf5 overexpression inhibits TGFβ induced EMT. (a, c) Different confocal slices through the Z stack image. (b, d) 3D rendering
Supplementary Figure 1
Supplementary Figure 1 Kif1a RNAi effect on basal progenitor differentiation Related to Figure 2. Representative confocal images of the VZ and SVZ of rat cortices transfected at E16 with scrambled or Kif1a
supplementary information
DOI: 10.1038/ncb2153 Figure S1 Ectopic expression of HAUSP up-regulates REST protein. (a) Immunoblotting showed that ectopic expression of HAUSP increased REST protein levels in ENStemA NPCs. (b) Immunofluorescent
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SUPPLEMENTARY INFORMATION Supplementary Figure 1. The expression of ephrin-b2 H2BGFP persists in the post-hearingonset organ of Corti and is specifically restricted to supporting cells. Sox2 immunolabeling
EPIGENETIC RE-EXPRESSION OF HIF-2α SUPPRESSES SOFT TISSUE SARCOMA GROWTH
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SUPPLEMENTARY FIGURES 1 Supplementary Figure 1, Adult hippocampal QNPs and TAPs uniformly express REST a-b) Confocal images of adult hippocampal mouse sections showing GFAP (green), Sox2 (red), and REST
Supplementary Figure 1. SA-β-Gal positive senescent cells in various cancer tissues. Representative frozen sections of breast, thyroid, colon and
Supplementary Figure 1. SA-β-Gal positive senescent cells in various cancer tissues. Representative frozen sections of breast, thyroid, colon and stomach cancer were stained with SA-β-Gal and nuclear fast
TEB. Id4 p63 DAPI Merge. Id4 CK8 DAPI Merge
a Duct TEB b Id4 p63 DAPI Merge Id4 CK8 DAPI Merge c d e Supplementary Figure 1. Identification of Id4-positive MECs and characterization of the Comma-D model. (a) IHC analysis of ID4 expression in the
Supplementary Figure 1. A. Bar graph representing the expression levels of the 19 indicated genes in the microarrays analyses comparing human lung
Supplementary Figure 1. A. Bar graph representing the expression levels of the 19 indicated genes in the microarrays analyses comparing human lung immortalized broncho-epithelial cells (AALE cells) expressing
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doi:10.1038/nature12652 Supplementary Figure 1. PRDM16 interacts with endogenous EHMT1 in brown adipocytes. Immunoprecipitation of PRDM16 complex by flag antibody (M2) followed by Western blot analysis
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DOI: 10.1038/ncb2566 Figure S1 CDKL5 protein expression pattern and localization in mouse brain. (a) Multiple-tissue western blot from a postnatal day (P) 21 mouse probed with an antibody against CDKL5.
Supplementary Figures
Supplementary Figures Supplementary Figure 1. Confirmation of Dnmt1 conditional knockout out mice. a, Representative images of sorted stem (Lin - CD49f high CD24 + ), luminal (Lin - CD49f low CD24 + )
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DOI: 10.1038/ncb3021 Supplementary figure 1 Characterisation of TIMPless fibroblasts. a) Relative gene expression of TIMPs1-4 by real time quantitative PCR (RT-qPCR) in WT or ΔTimp fibroblasts (mean ±
Supplementary Figure 1. Validation of astrocytes. Primary astrocytes were
Supplementary Figure 1. Validation of astrocytes. Primary astrocytes were separated from the glial cultures using a mild trypsinization protocol. Anti-glial fibrillary acidic protein (GFAP) immunofluorescent
Supplementary Table 1. List of primers used in this study
Supplementary Table 1. List of primers used in this study Gene Forward primer Reverse primer Rat Met 5 -aggtcgcttcatgcaggt-3 5 -tccggagacacaggatgg-3 Rat Runx1 5 -cctccttgaaccactccact-3 5 -ctggatctgcctggcatc-3
T H E J O U R N A L O F C E L L B I O L O G Y
T H E J O U R N A L O F C E L L B I O L O G Y Supplemental material Amelio et al., http://www.jcb.org/cgi/content/full/jcb.201203134/dc1 Figure S1. mir-24 regulates proliferation and by itself induces
Supplementary Fig. 1 eif6 +/- mice show a reduction in white adipose tissue, blood lipids and normal glycogen synthesis. The cohort of the original
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SUPPLEMENTARY LEGENDS...
TABLE OF CONTENTS SUPPLEMENTARY LEGENDS... 2 11 MOVIE S1... 2 FIGURE S1 LEGEND... 3 FIGURE S2 LEGEND... 4 FIGURE S3 LEGEND... 5 FIGURE S4 LEGEND... 6 FIGURE S5 LEGEND... 7 FIGURE S6 LEGEND... 8 FIGURE
stability and tumor suppression
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DOI: 1.138/ncb3355 a S1A8 + cells/ total.1.8.6.4.2 b S1A8/?-Actin c % T-cell proliferation 3 25 2 15 1 5 T cells Supplementary Figure 1 Inter-tumoral heterogeneity of MDSC accumulation in mammary tumor
Supplemental Figure 1. Intracranial transduction of a modified ptomo lentiviral vector in the mouse
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Table 1. Clinicopathological features of melanoma samples (shown in Fig. 2) exhibiting higher XYZ mrna levels.
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F-actin VWF Vinculin. F-actin. Vinculin VWF
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SUPPLEMENTARY FIGURES AND TABLE
SUPPLEMENTARY FIGURES AND TABLE Supplementary Figure S1: Characterization of IRE1α mutants. A. U87-LUC cells were transduced with the lentiviral vector containing the GFP sequence (U87-LUC Tet-ON GFP).
Type of file: PDF Size of file: 0 KB Title of file for HTML: Supplementary Information Description: Supplementary Figures
Type of file: PDF Size of file: 0 KB Title of file for HTML: Supplementary Information Description: Supplementary Figures Supplementary Figure 1 mir-128-3p is highly expressed in chemoresistant, metastatic
Supplementary Figure 1 IMQ-Induced Mouse Model of Psoriasis. IMQ cream was
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fl/+ KRas;Atg5 fl/+ KRas;Atg5 fl/fl KRas;Atg5 fl/fl KRas;Atg5 Supplementary Figure 1. Gene set enrichment analyses. (a) (b)
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Supplementary Figure 1. Characterization of NMuMG-ErbB2 and NIC breast cancer cells expressing shrnas targeting LPP. NMuMG-ErbB2 cells (a) and NIC
Supplementary Figure 1. Characterization of NMuMG-ErbB2 and NIC breast cancer cells expressing shrnas targeting LPP. NMuMG-ErbB2 cells (a) and NIC cells (b) were engineered to stably express either a LucA-shRNA
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b 350 300 250 200 150 100 50 0 E0 E10 E50 E0 E10 E50 E0 E10 E50 E0 E10 E50 Number of organoids per well 350 300 250 200 150 100 50 0 R0 R50 R100 R500 1st 2nd 3rd Noggin 100 ng/ml Noggin 10 ng/ml Noggin
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DOI: 10.1038/ncb2638 Figure S1 Morphological characteristics of fetal testes and ovaries from 6.5-20 developmental weeks. Representative images of Hematoxylin and Eosin staining of testes and ovaries over
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supplementary information
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Supplementary Figure 1: TSLP receptor skin expression in dcssc. A: Healthy control (HC) skin with TSLP receptor expression in brown (10x magnification). B: Second HC skin stained for TSLP receptor in brown
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c Ischemia (30 min) Reperfusion (8 w) Supplementary Figure bp 300 bp Ischemia (30 min) Reperfusion (4 h) Dox 20 mg/kg i.p.
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DOI: 10.1038/ncb2610 Figure S1 FSMCs derived from MSLN CLN transgenic mice express smooth muscle-specific proteins. Beta-galactosidase is ubiquitously expressed within cultured FSMCs derived from MSLN
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effects on organ development. a-f, Eye and wing discs with clones of ε j2b10 show no
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Supplemental Figure 1
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Supplementary Figure 1. The CagA-dependent wound healing or transwell migration of gastric cancer cell. AGS cells transfected with vector control or 3xflag-CagA expression vector were wounded using a pipette
marker. DAPI labels nuclei. Flies were 20 days old. Scale bar is 5 µm. Ctrl is
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Figure S1 Induction of non-apoptotic death of SV40-transformed and primary DKO MEFs, and DKO thymocytes. (A-F) STS-induced non-apoptotic death of DKO MEF. (A, B) Reduced viability of DKO MEFs after exposure
Nature Structural and Molecular Biology: doi: /nsmb Supplementary Figure 1
Supplementary Figure 1 Mutational analysis of the SA2-Scc1 interaction in vitro and in human cells. (a) Autoradiograph (top) and Coomassie stained gel (bottom) of 35 S-labeled Myc-SA2 proteins (input)
File Name: Supplementary Information Description: Supplementary Figures and Supplementary Table. File Name: Peer Review File Description:
File Name: Supplementary Information Description: Supplementary Figures and Supplementary Table File Name: Peer Review File Description: Supplementary Table 1 Primers and taqman probes used were the following:
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-. -. SUPPLEMENTARY INFORMATION DOI: 1.1/ncb86 a WAT-1 WAT- BAT-1 BAT- sk-muscle-1 sk-muscle- mir-133b mir-133a mir-6 mir-378 mir-1 mir-85 mir-378 mir-6a mir-18 mir-133a mir- mir- mir-341 mir-196a mir-17
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File Name: Supplementary Information Description: Supplementary Figures and Supplementary Tables File Name: Peer Review File Description: Primer Name Sequence (5'-3') AT ( C) RT-PCR USP21 F 5'-TTCCCATGGCTCCTTCCACATGAT-3'
Supplemental Figures:
Supplemental Figures: Figure 1: Intracellular distribution of VWF by electron microscopy in human endothelial cells. a) Immunogold labeling of LC3 demonstrating an LC3-positive autophagosome (white arrow)
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Supplementary Figure 1 Expression of Crb3 in mouse sciatic nerve: biochemical analysis (a) Schematic of Crb3 isoforms, ERLI and CLPI, indicating the
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Supplementary Figure 1: Digitoxin induces apoptosis in primary human melanoma cells but not in normal melanocytes, which express lower levels of the
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DOI: 10.1038/ncb2988 Supplementary Figure 1 Kif7 L130P encodes a stable protein that does not localize to cilia tips. (a) Immunoblot with KIF7 antibody in cell lysates of wild-type, Kif7 L130P and Kif7
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Supplemental Table S1
Supplemental Table S. Tumorigenicity and metastatic potential of 44SQ cell subpopulations a Tumorigenicity b Average tumor volume (mm ) c Lung metastasis d CD high /4 8. 8/ CD low /4 6./ a Mice were injected
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TMA-VARESE COHORT-1 TMA-BERN COHORT-2
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Supplementary Information
Supplementary Information Astrocytes regulate adult hippocampal neurogenesis through ephrin-b signaling Randolph S. Ashton, Anthony Conway, Chinmay Pangarkar, Jamie Bergen, Kwang-Il Lim, Priya Shah, Mina
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Supplementary Fig. S1. Schematic diagram of minigenome segments.
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Supplementary Figure 1. Nature Neuroscience: doi: /nn.4547
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SUPPLEMENTARY INFORMATION
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Supplementary Figures
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