plex307 Search Results


plasmid plex307 dpp4 puro  (Addgene inc)
93
Addgene inc plasmid plex307 dpp4 puro
Plasmid Plex307 Dpp4 Puro, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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human slc7a5  (Addgene inc)
94
Addgene inc human slc7a5
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Human Slc7a5, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plex307 ace2 puro  (Addgene inc)
93
Addgene inc plex307 ace2 puro
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Plex307 Ace2 Puro, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plasmid plex307 furin puro  (Addgene inc)
91
Addgene inc plasmid plex307 furin puro
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Plasmid Plex307 Furin Puro, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plex307 apn puro  (Addgene inc)
93
Addgene inc plex307 apn puro
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Plex307 Apn Puro, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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pentr 2b gfp kif26b c plasmid  (Addgene inc)
90
Addgene inc pentr 2b gfp kif26b c plasmid
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Pentr 2b Gfp Kif26b C Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plex307 tmprss2 blast  (Addgene inc)
93
Addgene inc plex307 tmprss2 blast
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Plex307 Tmprss2 Blast, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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cd13 aminopeptidase n sequence  (Addgene inc)
91
Addgene inc cd13 aminopeptidase n sequence
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Cd13 Aminopeptidase N Sequence, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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lentiviruses  (Addgene inc)
91
Addgene inc lentiviruses
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Lentiviruses, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plex 307 sod1e133instt  (Addgene inc)
88
Addgene inc plex 307 sod1e133instt
Figure 1. A functional genomics screen identifies <t>SLC7A5</t> as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.
Plex 307 Sod1e133instt, supplied by Addgene inc, used in various techniques. Bioz Stars score: 88/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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cmv rras plasmid  (Addgene inc)
91
Addgene inc cmv rras plasmid
a Violin plots show decreased R-RAS mRNA in EC of IPF ( n = 4 lungs) compared to donor ( n = 8 lungs), GSE 122960 datasets. R-RAS expression was log normalized. b R-RAS mRNA was decreased in FACS-sorted ECs from IPF lungs ( n = 6) compared to donor lungs ( n = 3) as shown by qRT-PCR. Data presented as mean ± SD, * p = 0.238, Mann–Whitney Two-tailed test. c Immunostaining for R-RAS, FOXF1 and CD31 shows co-localization of FOXF1 and R-RAS in ECs of donor lungs ( n = 5). Neither R-RAS nor FOXF1 are detected in ECs within IPF fibrotic lesions ( n = 5). Nuclei are counterstained with DAPI. Bar = 20 μm. d qRT-PCR shows that shRNA-mediated knockdown of FOXF1 (sh FOXF1 ) in HUVECs decreased R-RAS mRNA compared to control (Scr). ACTB mRNA was used for normalization ( n = 3), ** p < 0.01 and *** p < 0.001 by Student’s T test (two-tailed). e qRT-PCR shows that siRNA-mediated knockdown of Foxf1 in mouse MFLM-91U ECs decreased <t>Rras</t> mRNA. Actb mRNA was used for normalization ( n = 3). f ChIP-seq shows direct binding of FOXF1 protein to Rras promoter region in MFLM-91U cells. Binding of FOXF1 to Rras promoter is associated with H3K4me3 marks but not H3K27me3 marks. g Schematic drawing of the pGL2-Rras-Luc construct with the −762/+13 bp Rras promoter region containing the FOXF1-binding site (top panel). In co-transfection experiments, <t>CMV-FOXF1</t> expression vector increased transcriptional activity of the −762/+13 Rras promoter region compared to CMV-empty vector (bottom panel), n = 3, *** p < 0.001. h Overexpression of R-Ras decreased Ccl2 and TNFα mRNAs in mock-transfected cells and prevented upregulation of Ccl2 and TNFα in cells transfected with Foxf1-specific siRNA. MFLM-91U cells were transfected with non-targeting siRNA (siControl) or si Foxf1 , and EEV-empty vector or EEV- Rras . Actb mRNA was used for normalization (siControl+EEV-empty Foxf1 , n = 6; siControl+EEV-empty Rras , n = 6; siControl+EEV-empty Ccl2 , n = 3; siControl+EEV-empty Tnf , n = 6; siControl+EEV-Rras Foxf1 , n = 6; siControl+EEV-Rras Rras , n = 6; siControl+EEV-Rras Ccl2 , n = 6; siControl+EEV-Rras Tnf , n = 3; siFoxf1+EEV-empty Foxf1 , n = 6; siFoxf1+EEV-empty Rras , n = 3; siFoxf1+EEV-empty Ccl2 , n = 3; siFoxf1+EEV-empty Tnf , n = 3; siFoxf1+EEV-Rras Foxf1 , n = 6; siFoxf1l+EEV-Rras Rras , n = 3; siFoxf1l+EEV-Rras Ccl2 , n = 6; siFoxf1+EEV-Rras Tnf , n = 3), * p < 0.05, ** p < 0.01, *** p < 0.001 by Student’s t test (two tailed). Source data are provided as a Source Data file.
Cmv Rras Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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plex307 ace2 blast  (Addgene inc)
93
Addgene inc plex307 ace2 blast
a Violin plots show decreased R-RAS mRNA in EC of IPF ( n = 4 lungs) compared to donor ( n = 8 lungs), GSE 122960 datasets. R-RAS expression was log normalized. b R-RAS mRNA was decreased in FACS-sorted ECs from IPF lungs ( n = 6) compared to donor lungs ( n = 3) as shown by qRT-PCR. Data presented as mean ± SD, * p = 0.238, Mann–Whitney Two-tailed test. c Immunostaining for R-RAS, FOXF1 and CD31 shows co-localization of FOXF1 and R-RAS in ECs of donor lungs ( n = 5). Neither R-RAS nor FOXF1 are detected in ECs within IPF fibrotic lesions ( n = 5). Nuclei are counterstained with DAPI. Bar = 20 μm. d qRT-PCR shows that shRNA-mediated knockdown of FOXF1 (sh FOXF1 ) in HUVECs decreased R-RAS mRNA compared to control (Scr). ACTB mRNA was used for normalization ( n = 3), ** p < 0.01 and *** p < 0.001 by Student’s T test (two-tailed). e qRT-PCR shows that siRNA-mediated knockdown of Foxf1 in mouse MFLM-91U ECs decreased <t>Rras</t> mRNA. Actb mRNA was used for normalization ( n = 3). f ChIP-seq shows direct binding of FOXF1 protein to Rras promoter region in MFLM-91U cells. Binding of FOXF1 to Rras promoter is associated with H3K4me3 marks but not H3K27me3 marks. g Schematic drawing of the pGL2-Rras-Luc construct with the −762/+13 bp Rras promoter region containing the FOXF1-binding site (top panel). In co-transfection experiments, <t>CMV-FOXF1</t> expression vector increased transcriptional activity of the −762/+13 Rras promoter region compared to CMV-empty vector (bottom panel), n = 3, *** p < 0.001. h Overexpression of R-Ras decreased Ccl2 and TNFα mRNAs in mock-transfected cells and prevented upregulation of Ccl2 and TNFα in cells transfected with Foxf1-specific siRNA. MFLM-91U cells were transfected with non-targeting siRNA (siControl) or si Foxf1 , and EEV-empty vector or EEV- Rras . Actb mRNA was used for normalization (siControl+EEV-empty Foxf1 , n = 6; siControl+EEV-empty Rras , n = 6; siControl+EEV-empty Ccl2 , n = 3; siControl+EEV-empty Tnf , n = 6; siControl+EEV-Rras Foxf1 , n = 6; siControl+EEV-Rras Rras , n = 6; siControl+EEV-Rras Ccl2 , n = 6; siControl+EEV-Rras Tnf , n = 3; siFoxf1+EEV-empty Foxf1 , n = 6; siFoxf1+EEV-empty Rras , n = 3; siFoxf1+EEV-empty Ccl2 , n = 3; siFoxf1+EEV-empty Tnf , n = 3; siFoxf1+EEV-Rras Foxf1 , n = 6; siFoxf1l+EEV-Rras Rras , n = 3; siFoxf1l+EEV-Rras Ccl2 , n = 6; siFoxf1+EEV-Rras Tnf , n = 3), * p < 0.05, ** p < 0.01, *** p < 0.001 by Student’s t test (two tailed). Source data are provided as a Source Data file.
Plex307 Ace2 Blast, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Image Search Results


Figure 1. A functional genomics screen identifies SLC7A5 as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.

Journal: Cell reports

Article Title: SLC7A5 is required for cancer cell growth under arginine-limited conditions.

doi: 10.1016/j.celrep.2024.115130

Figure Lengend Snippet: Figure 1. A functional genomics screen identifies SLC7A5 as required for growth on citrulline (A) Schematic of citrulline synthesis into arginine. (B) Concentrations of arginine and citrulline in mouse serum (mean ± SD, n = 5). (C) Rates of appearance of arginine and citrulline as determined by steady-state labeling fraction upon defined infusion (mean ± SD, n = 5). (D) Media conditions used for the CRISPR screen. (E) Growth of A375mASS1-OE cells in Low Arg, High Arg, and Low Arg minus Cit media. Data are expressed in terms of relative growth, where the readings on day 0 are normalized to 1 (mean ± SD, n = 4). (F) Schematic of the CRISPR-based screen to identify metabolic genes required for growth under low-arginine conditions. (G) Gene scores from cells grown under Low Arg vs. High Arg. SLC7A5, the top hit, is highlighted in red. SLC7A1 (arginine transporter) and ASL (arginosuccinate lyase) are also highlighted. The red line is the equation y = x passing though (0,0) to highlight differential essentiality. (H) Top 25 genes scoring as selectively essential in Low Arg vs. High Arg. Genes linked to glycosylation are shown in blue, the urea cycle in red, transport in purple, reactive oxygen species (ROS) metabolism in orange, and other genes in green.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Oligonucleotides sgRNA for CRISPR KO and qPCR Oligonucleotides This study Table S1 Recombinant DNA Human ASS1 in pLX304 DNASU Cat#HSCD00438196 Human SLC7A5 in pDONR221 DNASU Cat#HSCD00042452 pLX307 Empty Vector Addgene Cat#41392 Human SLC7A5 in pLX307 This study N/A pSpCas9(BB)-2A-GFP (PX458)-ASS1_exon11 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0-SLC7A5_exon2 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0 Addgene Cat#62988 pSpCas9(BB)-2A-GFP (PX458) Addgene Cat#48138 Human CRISPR Metabolic Gene Knockout Library Addgene Cat#110066 psPAX2 Addgene Cat#12260 PMD2.G Addgene Cat#12259 Software and algorithms MAGeCK GitHub https://github.com/liulab-dfci/MAGeCK GraphPad Prism 10 GraphPad Software https://www.graphpad.com/ R Version 4.4.0 R https://cran.r-project.org/ EL-MAVEN Software Elucidata Agrawal et al.79 AccuCor GitHub https://github.com/XiaoyangSu/AccuCor Biorender Biorender https://www.biorender.com/ Other Countess 3 Automated Cell Counter Invitrogen Cat#AMQAX2000 BioTek Synergy Neo2 Hybrid Multimode Reader Agilent N/A BioRad ChemiDoc MP Imaging System BioRad Cat#12003154 Cryomill Retsch N/A LC480 PCR Lightcycler Roche Cat#05015278001

Techniques: Functional Assay, Labeling, CRISPR, Glycoproteomics

Figure 2. SLC7A5 is required for citrulline uptake, metabolism, and growth in arginine-free media (A) Immunoblot showing SLC7A5 and ASS1 expression in two SLC7A5 KO clones in the A375mASS1-OE (melanoma) cell line. WT, SLC7A5 WT. b-Actin was used as a loading control. (B) Growth of A375mASS1-OE and SLC7A5 KO cells in the indicated media. The base medium was RPMI without arginine supplemented with 110 mM arginine or citrulline as indicated. Media were refreshed on day 2 (mean ± SD, n = 4). (C) Immunoblot showing SLC7A5 expression from two SLC7A5 KO clones in the MCF7 (breast cancer) cell line. b-Actin was used as a loading control. (D) Growth of MCF7 SLC7A5 WT and KO cells in the indicated media. Media were refreshed on day 2. (mean ± SD, n = 4). (E) Citrulline consumption from the media over a 48-h period; A375mASS1-OE and SLC7A5 KO. Media: RPMI + 110 mM Arg + 110 mM Cit (mean ± SD, n = 4). **p < 0.01, unpaired two-tailed t test. (F) Citrulline consumption from the media with SLC7A5 WT and KO over a 48-h period; MCF7 WT and SLC7A5 KO. Media: RPMI + 110 mM Arg + 110 mM Cit (mean ± SD, n = 4). ***p < 0.001, unpaired two-tailed t test. (G) Uptake of [1-13C]-citrulline over a 15-min period in A375m WT and SLC7A5 KO cells. Media: RPMI + 110 mM Arg + 40 mM Cit (mean ± SD, n = 3). ***p < 0.001, unpaired two-tailed t test. (H) Ion count of unlabeled (M+0) and labeled (M+1) argininosuccinate. Cells were plated in RPMI + 110 mM [1-13C]-citrulline for 16 h, and then metabolites were extracted and analyzed using LC-MS (mean ± SD, n = 4).

Journal: Cell reports

Article Title: SLC7A5 is required for cancer cell growth under arginine-limited conditions.

doi: 10.1016/j.celrep.2024.115130

Figure Lengend Snippet: Figure 2. SLC7A5 is required for citrulline uptake, metabolism, and growth in arginine-free media (A) Immunoblot showing SLC7A5 and ASS1 expression in two SLC7A5 KO clones in the A375mASS1-OE (melanoma) cell line. WT, SLC7A5 WT. b-Actin was used as a loading control. (B) Growth of A375mASS1-OE and SLC7A5 KO cells in the indicated media. The base medium was RPMI without arginine supplemented with 110 mM arginine or citrulline as indicated. Media were refreshed on day 2 (mean ± SD, n = 4). (C) Immunoblot showing SLC7A5 expression from two SLC7A5 KO clones in the MCF7 (breast cancer) cell line. b-Actin was used as a loading control. (D) Growth of MCF7 SLC7A5 WT and KO cells in the indicated media. Media were refreshed on day 2. (mean ± SD, n = 4). (E) Citrulline consumption from the media over a 48-h period; A375mASS1-OE and SLC7A5 KO. Media: RPMI + 110 mM Arg + 110 mM Cit (mean ± SD, n = 4). **p < 0.01, unpaired two-tailed t test. (F) Citrulline consumption from the media with SLC7A5 WT and KO over a 48-h period; MCF7 WT and SLC7A5 KO. Media: RPMI + 110 mM Arg + 110 mM Cit (mean ± SD, n = 4). ***p < 0.001, unpaired two-tailed t test. (G) Uptake of [1-13C]-citrulline over a 15-min period in A375m WT and SLC7A5 KO cells. Media: RPMI + 110 mM Arg + 40 mM Cit (mean ± SD, n = 3). ***p < 0.001, unpaired two-tailed t test. (H) Ion count of unlabeled (M+0) and labeled (M+1) argininosuccinate. Cells were plated in RPMI + 110 mM [1-13C]-citrulline for 16 h, and then metabolites were extracted and analyzed using LC-MS (mean ± SD, n = 4).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Oligonucleotides sgRNA for CRISPR KO and qPCR Oligonucleotides This study Table S1 Recombinant DNA Human ASS1 in pLX304 DNASU Cat#HSCD00438196 Human SLC7A5 in pDONR221 DNASU Cat#HSCD00042452 pLX307 Empty Vector Addgene Cat#41392 Human SLC7A5 in pLX307 This study N/A pSpCas9(BB)-2A-GFP (PX458)-ASS1_exon11 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0-SLC7A5_exon2 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0 Addgene Cat#62988 pSpCas9(BB)-2A-GFP (PX458) Addgene Cat#48138 Human CRISPR Metabolic Gene Knockout Library Addgene Cat#110066 psPAX2 Addgene Cat#12260 PMD2.G Addgene Cat#12259 Software and algorithms MAGeCK GitHub https://github.com/liulab-dfci/MAGeCK GraphPad Prism 10 GraphPad Software https://www.graphpad.com/ R Version 4.4.0 R https://cran.r-project.org/ EL-MAVEN Software Elucidata Agrawal et al.79 AccuCor GitHub https://github.com/XiaoyangSu/AccuCor Biorender Biorender https://www.biorender.com/ Other Countess 3 Automated Cell Counter Invitrogen Cat#AMQAX2000 BioTek Synergy Neo2 Hybrid Multimode Reader Agilent N/A BioRad ChemiDoc MP Imaging System BioRad Cat#12003154 Cryomill Retsch N/A LC480 PCR Lightcycler Roche Cat#05015278001

Techniques: Western Blot, Expressing, Clone Assay, Control, Two Tailed Test, Labeling, Liquid Chromatography with Mass Spectroscopy

Figure 4. SLC7A5 and ASS1 are upregu- lated in response to arginine starvation (A) Relative transcript levels of ATF4, ASS1, SLC7A5, and SLC7A1 in A375m WT cells cultured in the indicated media as determined by RT- qPCR. Ct values were normalized to RPS2 (mean ± SD, n = 4). **p < 0.01, ***p < 0.001, ****p < 0.0001; 2-way ANOVA and multiple com- parisons were corrected using the Holm-Sidak method. (B) Immunoblots of SLC7A5 and ASS1 from A375m, HCT116, and LN229 WT cells plated for 48 h in the indicated media. (C) Relative amino acid levels of cells plated for 24 h in either RPMI + 110 mM Cit (Cit, red), RPMI + 110 mM Arg (Arg, blue), RPMI + 110 mM Cit + 1 mM GCN2iB (Cit + GCN2iB, gray) or RPMI + 110 mM Arg + 1 mM GCN2iB (Arg + GCN2iB, gold) (mean ± SD, n = 3–4). ns, not significant; **p < 0.01, ***p < 0.001, ****p < 0.0001, multiple comparisons- corrected 2-way ANOVA. (D) Immunoblot of SLC7A5 and ATF4 in A375m WT cells plated for 48 h with and without GCN2iB (1 mM) in the indicated media containing combi- nations of RPMI + 110 mM Cit or RPMI + 110 mM Arg. (E) Correlation between transcript levels of SLC7A5 versus ASS1 in breast cancer (BRCA) tumor transcript data taken from TCGA. The gray area around the linear regression line indicates the 95% confidence interval. (F) Correlation between transcript levels of SLC7A5 versus ASS1 in the luminal B (LumB) subtype of BRCA tumor transcript data taken from TCGA. The gray area around the linear regression line indicates the 95% confidence interval.

Journal: Cell reports

Article Title: SLC7A5 is required for cancer cell growth under arginine-limited conditions.

doi: 10.1016/j.celrep.2024.115130

Figure Lengend Snippet: Figure 4. SLC7A5 and ASS1 are upregu- lated in response to arginine starvation (A) Relative transcript levels of ATF4, ASS1, SLC7A5, and SLC7A1 in A375m WT cells cultured in the indicated media as determined by RT- qPCR. Ct values were normalized to RPS2 (mean ± SD, n = 4). **p < 0.01, ***p < 0.001, ****p < 0.0001; 2-way ANOVA and multiple com- parisons were corrected using the Holm-Sidak method. (B) Immunoblots of SLC7A5 and ASS1 from A375m, HCT116, and LN229 WT cells plated for 48 h in the indicated media. (C) Relative amino acid levels of cells plated for 24 h in either RPMI + 110 mM Cit (Cit, red), RPMI + 110 mM Arg (Arg, blue), RPMI + 110 mM Cit + 1 mM GCN2iB (Cit + GCN2iB, gray) or RPMI + 110 mM Arg + 1 mM GCN2iB (Arg + GCN2iB, gold) (mean ± SD, n = 3–4). ns, not significant; **p < 0.01, ***p < 0.001, ****p < 0.0001, multiple comparisons- corrected 2-way ANOVA. (D) Immunoblot of SLC7A5 and ATF4 in A375m WT cells plated for 48 h with and without GCN2iB (1 mM) in the indicated media containing combi- nations of RPMI + 110 mM Cit or RPMI + 110 mM Arg. (E) Correlation between transcript levels of SLC7A5 versus ASS1 in breast cancer (BRCA) tumor transcript data taken from TCGA. The gray area around the linear regression line indicates the 95% confidence interval. (F) Correlation between transcript levels of SLC7A5 versus ASS1 in the luminal B (LumB) subtype of BRCA tumor transcript data taken from TCGA. The gray area around the linear regression line indicates the 95% confidence interval.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Oligonucleotides sgRNA for CRISPR KO and qPCR Oligonucleotides This study Table S1 Recombinant DNA Human ASS1 in pLX304 DNASU Cat#HSCD00438196 Human SLC7A5 in pDONR221 DNASU Cat#HSCD00042452 pLX307 Empty Vector Addgene Cat#41392 Human SLC7A5 in pLX307 This study N/A pSpCas9(BB)-2A-GFP (PX458)-ASS1_exon11 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0-SLC7A5_exon2 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0 Addgene Cat#62988 pSpCas9(BB)-2A-GFP (PX458) Addgene Cat#48138 Human CRISPR Metabolic Gene Knockout Library Addgene Cat#110066 psPAX2 Addgene Cat#12260 PMD2.G Addgene Cat#12259 Software and algorithms MAGeCK GitHub https://github.com/liulab-dfci/MAGeCK GraphPad Prism 10 GraphPad Software https://www.graphpad.com/ R Version 4.4.0 R https://cran.r-project.org/ EL-MAVEN Software Elucidata Agrawal et al.79 AccuCor GitHub https://github.com/XiaoyangSu/AccuCor Biorender Biorender https://www.biorender.com/ Other Countess 3 Automated Cell Counter Invitrogen Cat#AMQAX2000 BioTek Synergy Neo2 Hybrid Multimode Reader Agilent N/A BioRad ChemiDoc MP Imaging System BioRad Cat#12003154 Cryomill Retsch N/A LC480 PCR Lightcycler Roche Cat#05015278001

Techniques: Cell Culture, Quantitative RT-PCR, Western Blot

Figure 5. A small-molecule inhibitor of SLC7A5 sensitizes cells to arginine deprivation (A) Growth of cancer cell lines cultured in RPMI + Arg, RPMI + Cit, with and without 1 mM JPH203. Media were refreshed on day 2 (mean ± SD, n = 4). ***p > 0.001, ****p < 0.0001, 2-way ANOVA. (B) Rescue of growth in the Cit + JPH203 condition with either 1 mM citrulline or 110 mM arginine. Media were refreshed on day 2 (mean ± SD, n = 4). ****p < 0.0001, unpaired two-tailed t test.

Journal: Cell reports

Article Title: SLC7A5 is required for cancer cell growth under arginine-limited conditions.

doi: 10.1016/j.celrep.2024.115130

Figure Lengend Snippet: Figure 5. A small-molecule inhibitor of SLC7A5 sensitizes cells to arginine deprivation (A) Growth of cancer cell lines cultured in RPMI + Arg, RPMI + Cit, with and without 1 mM JPH203. Media were refreshed on day 2 (mean ± SD, n = 4). ***p > 0.001, ****p < 0.0001, 2-way ANOVA. (B) Rescue of growth in the Cit + JPH203 condition with either 1 mM citrulline or 110 mM arginine. Media were refreshed on day 2 (mean ± SD, n = 4). ****p < 0.0001, unpaired two-tailed t test.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Oligonucleotides sgRNA for CRISPR KO and qPCR Oligonucleotides This study Table S1 Recombinant DNA Human ASS1 in pLX304 DNASU Cat#HSCD00438196 Human SLC7A5 in pDONR221 DNASU Cat#HSCD00042452 pLX307 Empty Vector Addgene Cat#41392 Human SLC7A5 in pLX307 This study N/A pSpCas9(BB)-2A-GFP (PX458)-ASS1_exon11 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0-SLC7A5_exon2 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0 Addgene Cat#62988 pSpCas9(BB)-2A-GFP (PX458) Addgene Cat#48138 Human CRISPR Metabolic Gene Knockout Library Addgene Cat#110066 psPAX2 Addgene Cat#12260 PMD2.G Addgene Cat#12259 Software and algorithms MAGeCK GitHub https://github.com/liulab-dfci/MAGeCK GraphPad Prism 10 GraphPad Software https://www.graphpad.com/ R Version 4.4.0 R https://cran.r-project.org/ EL-MAVEN Software Elucidata Agrawal et al.79 AccuCor GitHub https://github.com/XiaoyangSu/AccuCor Biorender Biorender https://www.biorender.com/ Other Countess 3 Automated Cell Counter Invitrogen Cat#AMQAX2000 BioTek Synergy Neo2 Hybrid Multimode Reader Agilent N/A BioRad ChemiDoc MP Imaging System BioRad Cat#12003154 Cryomill Retsch N/A LC480 PCR Lightcycler Roche Cat#05015278001

Techniques: Cell Culture, Two Tailed Test

Figure 6. SLC7A5 regulates citrulline metabolism in an in vivo xenograft model (A) Immunoblot of SLC7A5 in tumors originating from A375m xenografts. b-Actin was used as a loading control. (B) Growth of bilateral A375m WT and SLC7A5 KO xenograft tumors. Mice were fed a high-protein chow diet (mean ± SEM, n = 10). **p < 0.01, paired two- tailed t test. (C) Labeling fraction of [1-13C]-citrulline in the serum of mice 1 h after intra- peritoneal injection (mean ± SD, n = 5). (D) Tumor [1-13C]-citrulline uptake normalized to serum citrulline. Mice were injected with 83 mM (0.07 g/kg) of [1-13C]-citrulline (mean ± SEM, n = 5). *p < 0.05, paired two-tailed t test.

Journal: Cell reports

Article Title: SLC7A5 is required for cancer cell growth under arginine-limited conditions.

doi: 10.1016/j.celrep.2024.115130

Figure Lengend Snippet: Figure 6. SLC7A5 regulates citrulline metabolism in an in vivo xenograft model (A) Immunoblot of SLC7A5 in tumors originating from A375m xenografts. b-Actin was used as a loading control. (B) Growth of bilateral A375m WT and SLC7A5 KO xenograft tumors. Mice were fed a high-protein chow diet (mean ± SEM, n = 10). **p < 0.01, paired two- tailed t test. (C) Labeling fraction of [1-13C]-citrulline in the serum of mice 1 h after intra- peritoneal injection (mean ± SD, n = 5). (D) Tumor [1-13C]-citrulline uptake normalized to serum citrulline. Mice were injected with 83 mM (0.07 g/kg) of [1-13C]-citrulline (mean ± SEM, n = 5). *p < 0.05, paired two-tailed t test.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Oligonucleotides sgRNA for CRISPR KO and qPCR Oligonucleotides This study Table S1 Recombinant DNA Human ASS1 in pLX304 DNASU Cat#HSCD00438196 Human SLC7A5 in pDONR221 DNASU Cat#HSCD00042452 pLX307 Empty Vector Addgene Cat#41392 Human SLC7A5 in pLX307 This study N/A pSpCas9(BB)-2A-GFP (PX458)-ASS1_exon11 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0-SLC7A5_exon2 This study N/A pSpCas9(BB)-2A-Puro (PX459) V2.0 Addgene Cat#62988 pSpCas9(BB)-2A-GFP (PX458) Addgene Cat#48138 Human CRISPR Metabolic Gene Knockout Library Addgene Cat#110066 psPAX2 Addgene Cat#12260 PMD2.G Addgene Cat#12259 Software and algorithms MAGeCK GitHub https://github.com/liulab-dfci/MAGeCK GraphPad Prism 10 GraphPad Software https://www.graphpad.com/ R Version 4.4.0 R https://cran.r-project.org/ EL-MAVEN Software Elucidata Agrawal et al.79 AccuCor GitHub https://github.com/XiaoyangSu/AccuCor Biorender Biorender https://www.biorender.com/ Other Countess 3 Automated Cell Counter Invitrogen Cat#AMQAX2000 BioTek Synergy Neo2 Hybrid Multimode Reader Agilent N/A BioRad ChemiDoc MP Imaging System BioRad Cat#12003154 Cryomill Retsch N/A LC480 PCR Lightcycler Roche Cat#05015278001

Techniques: In Vivo, Western Blot, Control, Two Tailed Test, Labeling, Injection

a Violin plots show decreased R-RAS mRNA in EC of IPF ( n = 4 lungs) compared to donor ( n = 8 lungs), GSE 122960 datasets. R-RAS expression was log normalized. b R-RAS mRNA was decreased in FACS-sorted ECs from IPF lungs ( n = 6) compared to donor lungs ( n = 3) as shown by qRT-PCR. Data presented as mean ± SD, * p = 0.238, Mann–Whitney Two-tailed test. c Immunostaining for R-RAS, FOXF1 and CD31 shows co-localization of FOXF1 and R-RAS in ECs of donor lungs ( n = 5). Neither R-RAS nor FOXF1 are detected in ECs within IPF fibrotic lesions ( n = 5). Nuclei are counterstained with DAPI. Bar = 20 μm. d qRT-PCR shows that shRNA-mediated knockdown of FOXF1 (sh FOXF1 ) in HUVECs decreased R-RAS mRNA compared to control (Scr). ACTB mRNA was used for normalization ( n = 3), ** p < 0.01 and *** p < 0.001 by Student’s T test (two-tailed). e qRT-PCR shows that siRNA-mediated knockdown of Foxf1 in mouse MFLM-91U ECs decreased Rras mRNA. Actb mRNA was used for normalization ( n = 3). f ChIP-seq shows direct binding of FOXF1 protein to Rras promoter region in MFLM-91U cells. Binding of FOXF1 to Rras promoter is associated with H3K4me3 marks but not H3K27me3 marks. g Schematic drawing of the pGL2-Rras-Luc construct with the −762/+13 bp Rras promoter region containing the FOXF1-binding site (top panel). In co-transfection experiments, CMV-FOXF1 expression vector increased transcriptional activity of the −762/+13 Rras promoter region compared to CMV-empty vector (bottom panel), n = 3, *** p < 0.001. h Overexpression of R-Ras decreased Ccl2 and TNFα mRNAs in mock-transfected cells and prevented upregulation of Ccl2 and TNFα in cells transfected with Foxf1-specific siRNA. MFLM-91U cells were transfected with non-targeting siRNA (siControl) or si Foxf1 , and EEV-empty vector or EEV- Rras . Actb mRNA was used for normalization (siControl+EEV-empty Foxf1 , n = 6; siControl+EEV-empty Rras , n = 6; siControl+EEV-empty Ccl2 , n = 3; siControl+EEV-empty Tnf , n = 6; siControl+EEV-Rras Foxf1 , n = 6; siControl+EEV-Rras Rras , n = 6; siControl+EEV-Rras Ccl2 , n = 6; siControl+EEV-Rras Tnf , n = 3; siFoxf1+EEV-empty Foxf1 , n = 6; siFoxf1+EEV-empty Rras , n = 3; siFoxf1+EEV-empty Ccl2 , n = 3; siFoxf1+EEV-empty Tnf , n = 3; siFoxf1+EEV-Rras Foxf1 , n = 6; siFoxf1l+EEV-Rras Rras , n = 3; siFoxf1l+EEV-Rras Ccl2 , n = 6; siFoxf1+EEV-Rras Tnf , n = 3), * p < 0.05, ** p < 0.01, *** p < 0.001 by Student’s t test (two tailed). Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Lung endothelial cells regulate pulmonary fibrosis through FOXF1/R-Ras signaling

doi: 10.1038/s41467-023-38177-2

Figure Lengend Snippet: a Violin plots show decreased R-RAS mRNA in EC of IPF ( n = 4 lungs) compared to donor ( n = 8 lungs), GSE 122960 datasets. R-RAS expression was log normalized. b R-RAS mRNA was decreased in FACS-sorted ECs from IPF lungs ( n = 6) compared to donor lungs ( n = 3) as shown by qRT-PCR. Data presented as mean ± SD, * p = 0.238, Mann–Whitney Two-tailed test. c Immunostaining for R-RAS, FOXF1 and CD31 shows co-localization of FOXF1 and R-RAS in ECs of donor lungs ( n = 5). Neither R-RAS nor FOXF1 are detected in ECs within IPF fibrotic lesions ( n = 5). Nuclei are counterstained with DAPI. Bar = 20 μm. d qRT-PCR shows that shRNA-mediated knockdown of FOXF1 (sh FOXF1 ) in HUVECs decreased R-RAS mRNA compared to control (Scr). ACTB mRNA was used for normalization ( n = 3), ** p < 0.01 and *** p < 0.001 by Student’s T test (two-tailed). e qRT-PCR shows that siRNA-mediated knockdown of Foxf1 in mouse MFLM-91U ECs decreased Rras mRNA. Actb mRNA was used for normalization ( n = 3). f ChIP-seq shows direct binding of FOXF1 protein to Rras promoter region in MFLM-91U cells. Binding of FOXF1 to Rras promoter is associated with H3K4me3 marks but not H3K27me3 marks. g Schematic drawing of the pGL2-Rras-Luc construct with the −762/+13 bp Rras promoter region containing the FOXF1-binding site (top panel). In co-transfection experiments, CMV-FOXF1 expression vector increased transcriptional activity of the −762/+13 Rras promoter region compared to CMV-empty vector (bottom panel), n = 3, *** p < 0.001. h Overexpression of R-Ras decreased Ccl2 and TNFα mRNAs in mock-transfected cells and prevented upregulation of Ccl2 and TNFα in cells transfected with Foxf1-specific siRNA. MFLM-91U cells were transfected with non-targeting siRNA (siControl) or si Foxf1 , and EEV-empty vector or EEV- Rras . Actb mRNA was used for normalization (siControl+EEV-empty Foxf1 , n = 6; siControl+EEV-empty Rras , n = 6; siControl+EEV-empty Ccl2 , n = 3; siControl+EEV-empty Tnf , n = 6; siControl+EEV-Rras Foxf1 , n = 6; siControl+EEV-Rras Rras , n = 6; siControl+EEV-Rras Ccl2 , n = 6; siControl+EEV-Rras Tnf , n = 3; siFoxf1+EEV-empty Foxf1 , n = 6; siFoxf1+EEV-empty Rras , n = 3; siFoxf1+EEV-empty Ccl2 , n = 3; siFoxf1+EEV-empty Tnf , n = 3; siFoxf1+EEV-Rras Foxf1 , n = 6; siFoxf1l+EEV-Rras Rras , n = 3; siFoxf1l+EEV-Rras Ccl2 , n = 6; siFoxf1+EEV-Rras Tnf , n = 3), * p < 0.05, ** p < 0.01, *** p < 0.001 by Student’s t test (two tailed). Source data are provided as a Source Data file.

Article Snippet: In vitro rescue experiments were performed by dual transfection of si Foxf1 and CMV- Rras plasmid (Addgene plasmid # 102864) into MFLM-91U cells using Dharmafect Duo transfection reagent (Dharmacon).

Techniques: Expressing, Quantitative RT-PCR, MANN-WHITNEY, Two Tailed Test, Immunostaining, shRNA, Knockdown, Control, ChIP-sequencing, Binding Assay, Construct, Cotransfection, Plasmid Preparation, Activity Assay, Over Expression, Transfection