Journal: Cell reports

Article Title: Chromatin-associated orphan snoRNA regulates DNA damage-mediated differentiation via a non-canonical complex.

doi: 10.1016/j.celrep.2022.110421

Figure Lengend Snippet: Figure 1. High-throughput microarray profiles casnoRNAs and identifies the redistribution of orphan SNORA73 in response to DNA damage (A) Schematic illustration of the subcellular fractionation followed by high-throughput customized snoRNA-microarray analyses. (B) Heatmap and cluster analysis of snoRNAome microarray showing the subset of orphan casnoRNAs in NB4. (C) Venn diagram showing the overlapping of identified casnoRNA (red) with downregulated snoRNAs (blue) in AML obtained from two published datasets. (D) Volcano plot showing the chromatin enrichment changes of orphan casnoRNAs in non-treated groups (control) versus doxorubicin-treated (2 mM for 4 h) groups (Dox). The significantly downregulated (blue, log2 fold change < 0.5, p <0.05) or upregulated (red, log2 fold change > 0.5, p < 0.05) casnoRNAs were shown. Vertical dashed lines indicate cut-off of log2 fold change (0.5 or 0.5), whereas the horizontal dashed lines indicate the cut-off of p value (0.05).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RIPA lysis buffer Beyotime Biotechnology Cat#P0013B RNasin Ribonuclease Inhibitor Promega Cat#N2115 13complete ULTRA protease inhibitor Roche Cat#5892970001 HaltTM Protease Inhibitor Cocktail, EDTA-Free (100X) Thermo Fisher Scientific Cat#78425 PierceTM Protein A/G Magnetic Beads Thermo Fisher Scientific Cat#88803 DynabeadsTM MyOneTM Streptavidin T1 Thermo Fisher Scientific Cat#65601 ProteinIso GST Resin TransGen Biotech Cat#GF101-01 Ni Sepharose 6 Fast Flow GE Healthcare Cat#GE17-5318-01 Amylose Resin NEB Cat#E8021L Human Methylcellulose Complete Media R&D systems Cat#HSC003 Recombinant human PARP1 protein abcam Cat#ab79663 Critical commercial assays RT reagent Kit Takara Cat#RR047A SYBR Premix ExTaq real-time PCR Kit Takara Cat#RR861A Mut Express II Fast Mutagenesis Kit V2 Vazyme Cat#C214-01 ClonExpress Ultra One Step Cloning Kit Vazyme Cat#C115 pEASY -Blunt Simple Cloning Kit TransGen Biotech Cat#CB111-01 Magna RIP RNA-Binding Protein Immunoprecipitation Kit Millipore Cat#17-700 TranscriptAid T7 High Yield Transcription Kit Thermo Fisher Scientific Cat#K0441 GeneJET RNA Purification Kit Thermo Fisher Scientific Cat#K0731 PierceMagnetic RNA-Protein Pull-Down Kit Thermo Fisher Scientific Cat#20164 Deposited data snoRNA microarray This paper GEO: GSE173730 Raw image data Mendeley data Mendeley Data: https://doi.org/10.17632/ nx2mxjbh7w.1 Experimental models: Cell lines NB4 DSMZ Cat#ACC-207; RRID: CVCL_0005 HL-60 ATCC Cat#CCL-240; RRID: CVCL_0002 U937 ATCC Cat#CRL-1593.2; RRID: CVCL_0007 293T ATCC Cat#CRL-1573; RRID: CVCL_0045 Experimental models: Organisms/strains Mouse: NOD.CB17-Prkdcscid/NcrCrl Charles river Cat#406 Xenograft AML mouse model This paper N/A Oligonucleotides Primers This paper See Table S3 siRNA This paper See Table S4 shRNA sequence This paper See Table S4 sgRNA sequence This paper See Table S4 Recombinant DNA pGreenPuro-shRNA System Biosciences Cat#SI505A-1 pCDH-CMV-MCS-EF1-Puro System Biosciences Cat#CD510B-1 pPACKH1-GAG System Biosciences Cat#LV500A-1 pPACKH1-REV System Biosciences Cat#LV500A-1 pVSV-G System Biosciences Cat#LV500A-1 pMAL-c2X Walker et al., 2010 Addgene#75286; RRID: Addgene_75286 dCas13d-GFP Konermann et al., 2018 Addgene#109050; RRID: Addgene_109050 (Continued on next page) Cell Reports 38, 110421, March 29, 2022 e2

Techniques: High Throughput Screening Assay, Microarray, Fractionation, Control

Journal: Cell reports

Article Title: Chromatin-associated orphan snoRNA regulates DNA damage-mediated differentiation via a non-canonical complex.

doi: 10.1016/j.celrep.2022.110421

Figure Lengend Snippet: Figure 2. SNORA73 downregulation limits DNA damage level and contributes to differentiation block and AML development (A) Both SNORA73A and SNORA73B are significantly downregulated in preliminary patients with AML (n = 53) vs. healthy control (n = 5). Relative expression was used to quantify its expression relative to that of 5S rRNA. Mann-Whitney test. (B and C) The levels of g-H2AX shown by immunofluorescence (B) and Western blot (C) in control and SNORA73-depleted cells. Scale bar: 5 mm. (D) Representative Wright-Giemsa staining images of control and SNORA73-depleted NB4, HL-60 and U937 cells treated with 1 mM ATRA for 24 h. Scale bar: 25 mm. (E) Flow cytometry quantification of CD11b levels in control and SNORA73-depleted NB4, HL-60, and U937 cells; cells were treated with 1 mM ATRA for 24 h. (F) Representative images (left) and quantification (right) of colony formation assay for control and SNORA73-depleted NB4, HL-60, and U937 cells. The cells were treated with 1 mM ATRA for 24 h before seeding. Scale bar: 50 mm.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RIPA lysis buffer Beyotime Biotechnology Cat#P0013B RNasin Ribonuclease Inhibitor Promega Cat#N2115 13complete ULTRA protease inhibitor Roche Cat#5892970001 HaltTM Protease Inhibitor Cocktail, EDTA-Free (100X) Thermo Fisher Scientific Cat#78425 PierceTM Protein A/G Magnetic Beads Thermo Fisher Scientific Cat#88803 DynabeadsTM MyOneTM Streptavidin T1 Thermo Fisher Scientific Cat#65601 ProteinIso GST Resin TransGen Biotech Cat#GF101-01 Ni Sepharose 6 Fast Flow GE Healthcare Cat#GE17-5318-01 Amylose Resin NEB Cat#E8021L Human Methylcellulose Complete Media R&D systems Cat#HSC003 Recombinant human PARP1 protein abcam Cat#ab79663 Critical commercial assays RT reagent Kit Takara Cat#RR047A SYBR Premix ExTaq real-time PCR Kit Takara Cat#RR861A Mut Express II Fast Mutagenesis Kit V2 Vazyme Cat#C214-01 ClonExpress Ultra One Step Cloning Kit Vazyme Cat#C115 pEASY -Blunt Simple Cloning Kit TransGen Biotech Cat#CB111-01 Magna RIP RNA-Binding Protein Immunoprecipitation Kit Millipore Cat#17-700 TranscriptAid T7 High Yield Transcription Kit Thermo Fisher Scientific Cat#K0441 GeneJET RNA Purification Kit Thermo Fisher Scientific Cat#K0731 PierceMagnetic RNA-Protein Pull-Down Kit Thermo Fisher Scientific Cat#20164 Deposited data snoRNA microarray This paper GEO: GSE173730 Raw image data Mendeley data Mendeley Data: https://doi.org/10.17632/ nx2mxjbh7w.1 Experimental models: Cell lines NB4 DSMZ Cat#ACC-207; RRID: CVCL_0005 HL-60 ATCC Cat#CCL-240; RRID: CVCL_0002 U937 ATCC Cat#CRL-1593.2; RRID: CVCL_0007 293T ATCC Cat#CRL-1573; RRID: CVCL_0045 Experimental models: Organisms/strains Mouse: NOD.CB17-Prkdcscid/NcrCrl Charles river Cat#406 Xenograft AML mouse model This paper N/A Oligonucleotides Primers This paper See Table S3 siRNA This paper See Table S4 shRNA sequence This paper See Table S4 sgRNA sequence This paper See Table S4 Recombinant DNA pGreenPuro-shRNA System Biosciences Cat#SI505A-1 pCDH-CMV-MCS-EF1-Puro System Biosciences Cat#CD510B-1 pPACKH1-GAG System Biosciences Cat#LV500A-1 pPACKH1-REV System Biosciences Cat#LV500A-1 pVSV-G System Biosciences Cat#LV500A-1 pMAL-c2X Walker et al., 2010 Addgene#75286; RRID: Addgene_75286 dCas13d-GFP Konermann et al., 2018 Addgene#109050; RRID: Addgene_109050 (Continued on next page) Cell Reports 38, 110421, March 29, 2022 e2

Techniques: Blocking Assay, Control, Expressing, MANN-WHITNEY, Western Blot, Staining, Flow Cytometry, Colony Assay

Journal: Cell reports

Article Title: Chromatin-associated orphan snoRNA regulates DNA damage-mediated differentiation via a non-canonical complex.

doi: 10.1016/j.celrep.2022.110421

Figure Lengend Snippet: Figure 4. SNORA73 regulates DNA damage-mediated differentiation in AML (A and B) The levels of g-H2AX shown by immunofluorescence (A) and Western blot (B) in control and SNORA73-depleted cells with 10 mM olaparib treatment for 24 h. Scale bar: 5 mm. (C) Western blot shows the levels of g-H2AX in control and SNORA73-depleted primary AML patient cells with or without 10 mM olaparib treatment for 24 h. (D and E) Representative Wright-Giemsa staining images (D) and flow cytometry quantification of CD11b levels (E) for control and SNORA73-depleted NB4 and U937 cells with 10 mM olaparib treatment for 24 h. Scale bar: 25 mm. (F) Illustration for ascites tumor development in NOD/SCID mice transplanted with sh-NC or sh-SNORA73 NB4 cells. (G) Representative flow cytometry plots and quantification of CD11b levels by flow cytometry for sh-NC and sh-SNORA73 NB4 cells isolated from ascites fluid. The cells were treated with 1 mM ATRA for 24 h. (H) Western blot for g-H2AX levels in sh-NC and sh-SNORA73 NB4 cells isolated from mouse ascites fluid (three mice each group). The cells were treated with or without 10 mM olaparib for 24 h. Values are mean ± SD (n = 3), Student’s t test. ***p < 0.001. See also Figure S5.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RIPA lysis buffer Beyotime Biotechnology Cat#P0013B RNasin Ribonuclease Inhibitor Promega Cat#N2115 13complete ULTRA protease inhibitor Roche Cat#5892970001 HaltTM Protease Inhibitor Cocktail, EDTA-Free (100X) Thermo Fisher Scientific Cat#78425 PierceTM Protein A/G Magnetic Beads Thermo Fisher Scientific Cat#88803 DynabeadsTM MyOneTM Streptavidin T1 Thermo Fisher Scientific Cat#65601 ProteinIso GST Resin TransGen Biotech Cat#GF101-01 Ni Sepharose 6 Fast Flow GE Healthcare Cat#GE17-5318-01 Amylose Resin NEB Cat#E8021L Human Methylcellulose Complete Media R&D systems Cat#HSC003 Recombinant human PARP1 protein abcam Cat#ab79663 Critical commercial assays RT reagent Kit Takara Cat#RR047A SYBR Premix ExTaq real-time PCR Kit Takara Cat#RR861A Mut Express II Fast Mutagenesis Kit V2 Vazyme Cat#C214-01 ClonExpress Ultra One Step Cloning Kit Vazyme Cat#C115 pEASY -Blunt Simple Cloning Kit TransGen Biotech Cat#CB111-01 Magna RIP RNA-Binding Protein Immunoprecipitation Kit Millipore Cat#17-700 TranscriptAid T7 High Yield Transcription Kit Thermo Fisher Scientific Cat#K0441 GeneJET RNA Purification Kit Thermo Fisher Scientific Cat#K0731 PierceMagnetic RNA-Protein Pull-Down Kit Thermo Fisher Scientific Cat#20164 Deposited data snoRNA microarray This paper GEO: GSE173730 Raw image data Mendeley data Mendeley Data: https://doi.org/10.17632/ nx2mxjbh7w.1 Experimental models: Cell lines NB4 DSMZ Cat#ACC-207; RRID: CVCL_0005 HL-60 ATCC Cat#CCL-240; RRID: CVCL_0002 U937 ATCC Cat#CRL-1593.2; RRID: CVCL_0007 293T ATCC Cat#CRL-1573; RRID: CVCL_0045 Experimental models: Organisms/strains Mouse: NOD.CB17-Prkdcscid/NcrCrl Charles river Cat#406 Xenograft AML mouse model This paper N/A Oligonucleotides Primers This paper See Table S3 siRNA This paper See Table S4 shRNA sequence This paper See Table S4 sgRNA sequence This paper See Table S4 Recombinant DNA pGreenPuro-shRNA System Biosciences Cat#SI505A-1 pCDH-CMV-MCS-EF1-Puro System Biosciences Cat#CD510B-1 pPACKH1-GAG System Biosciences Cat#LV500A-1 pPACKH1-REV System Biosciences Cat#LV500A-1 pVSV-G System Biosciences Cat#LV500A-1 pMAL-c2X Walker et al., 2010 Addgene#75286; RRID: Addgene_75286 dCas13d-GFP Konermann et al., 2018 Addgene#109050; RRID: Addgene_109050 (Continued on next page) Cell Reports 38, 110421, March 29, 2022 e2

Techniques: Western Blot, Control, Staining, Cytometry, Isolation

Journal: Biomolecules

Article Title: Rapid Detection of PML::RARA Fusions in Acute Promyelocytic Leukemia: CRISPR/Cas9 Nanopore Sequencing with Adaptive Sampling

doi: 10.3390/biom14121595

Figure Lengend Snippet: Standard-of-care genetic tests and CENAS-based nanopore sequencing in this study.

Article Snippet: The NB4 cell line was obtained from Accegen (Fairfield, NJ, USA) and the GM12878 cell line from Coriell Institute (Camden, NJ, USA).

Techniques: Nanopore Sequencing, Fluorescence In Situ Hybridization, Microarray, Single Vesicle Fusion Assay