centrin2 Search Results


centrin2  (Novus Biologicals)
93
Novus Biologicals centrin2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Centrin2, supplied by Novus Biologicals, 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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Average 93 stars, based on 1 article reviews
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hek293 cells  (Sino Biological)
92
Sino Biological hek293 cells
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Hek293 Cells, supplied by Sino Biological, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti centrin 2  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology anti centrin 2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Anti Centrin 2, supplied by Santa Cruz Biotechnology, 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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Average 93 stars, based on 1 article reviews
anti centrin 2 - by Bioz Stars, 2026-04
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goat anti cetn2  (Proteintech)
93
Proteintech goat anti cetn2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Goat Anti Cetn2, supplied by Proteintech, 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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Average 93 stars, based on 1 article reviews
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gfp  (Addgene inc)
93
Addgene inc gfp
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Gfp, 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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Average 93 stars, based on 1 article reviews
gfp - by Bioz Stars, 2026-04
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rabbit anti centrin2  (Cell Signaling Technology Inc)
93
Cell Signaling Technology Inc rabbit anti centrin2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Rabbit Anti Centrin2, supplied by Cell Signaling Technology 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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Average 93 stars, based on 1 article reviews
rabbit anti centrin2 - by Bioz Stars, 2026-04
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mcherry centrin2 n 10  (Addgene inc)
93
Addgene inc mcherry centrin2 n 10
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Mcherry Centrin2 N 10, 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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mcherry centrin2 n 10 - by Bioz Stars, 2026-04
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centrin 2  (Santa Cruz Biotechnology)
88
Santa Cruz Biotechnology centrin 2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Centrin 2, supplied by Santa Cruz Biotechnology, 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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mouse anti centrin 2  (Proteintech)
94
Proteintech mouse anti centrin 2
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Mouse Anti Centrin 2, supplied by Proteintech, 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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tdgtm1psch mice  (Addgene inc)
91
Addgene inc tdgtm1psch mice
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Tdgtm1psch Mice, 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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centrin 2 antobodies  (Santa Cruz Biotechnology)
85
Santa Cruz Biotechnology centrin 2 antobodies
Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for <t>Centrin2</t> and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.
Centrin 2 Antobodies, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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mouse antibodies against α-tubulin  (Merck & Co)
90
Merck & Co mouse antibodies against α-tubulin
A , Depletion of DONSON induced four separated centrioles in HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies <t>against</t> <t>centrin-2</t> (green) and Cep192 (red). B, Histograms represent frequency of interphase cells with the indicated phenotypes observed in A. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). C, Depletion of DONSON did not affect the number of centrin foci in G1 phase. HeLa cells were treated with siControl or siDONSON for 48 h, and 10 μM EdU for 30 min before fixation, followed by immunostaining with antibodies against centrin-2 (green) and CENP-F (red). S phase cells were monitored by EdU staining (red). CENPF and EdU double-negative cells were classified as G1 phase cells, and others as S or G2 phase cells. D, Histograms represent frequency of interphase cells with the indicated phenotype observed in C. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). E, Recruitment of C-Nap1 to separated centrioles in DONSON-depleted HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies against centrin-2 (green) and C-Nap1 (red). F, Histograms represent frequency of cells with the indicated number of C-Nap1 positive centrioles among cells with four separated centrioles observed in E. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). G, Time-lapse observation of precocious centriole disengagement in interphase in DONSON-depleted cells. HeLa GFP-centrin-1 cells were visualized every 20 min for 48 h after 24 h treatment of siControl or siDONSON and 3 h treatment of 100 nM SiR-DNA. Left– right arrows indicate precociously disengaged centrioles. H, Schematic model of the phenotypes of DONSON depletion. Depletion of DONSON causes precocious centriole disengagement in interphase. All scale bars, 5 μm. Two-tailed, unpaired Student’s t-test was used in B and D to obtain P value. **, p < 0.01; N.S., not significantly different (p > 0.05).
Mouse Antibodies Against α Tubulin, supplied by Merck & Co, 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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Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for Centrin2 and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.

Journal: Frontiers in Molecular Biosciences

Article Title: Brain microvascular endothelial cells possess a second cilium that arises from the daughter centriole

doi: 10.3389/fmolb.2023.1250016

Figure Lengend Snippet: Brain ECs possess a second cilium from the daughter centriole. (A) . Brain endothelial cells from frontal cortex of one-week-old Tie2Cre·Pkd2 WT/WT (with Cre activation; control wild-type group) and Tie2Cre·Pkd2 flox/flox (with Cre activation; experimental Pkd2 group) mice were collected and immunostained for primary cilia with acetylated-α-tubulin and nuclei with DAPI using the immunofluorescence method. Scale bar = 20 μm. (B) . The table represents the number of brain endothelial cells with a two-cilia phenotype. A minimum of 50 cells were assessed in triplicates for two-cilia phenotype detection. Scale bar = 10 μm. (C) . Human primary brain microvascular endothelial cells (HBMECs) were immunostained for mother centriole distal and subdistal appendage markers, CEP164 and NINEIN, respectively (red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother centriole and arrowheads represent second cilia from the daughter centriole. (D) . HBMECs stained for Centrin2 and HsSAS, markers for distal lumen and procentriolar markers of the mother and daughter centriole (Red), nuclei stained with DAPI, and primary cilia stained with ARL13B (green). White arrows indicate cilia from the mother and daughter centriole and their respective cilia. Scale bar = 10 μm. (E) . Primary cilia size varies based on ciliogenesis from the mother and daughter centrioles of HBMECs. N = 25 double cilia cells were quantified for their respective size. (F) . Pictorial representation of the centriole markers used in the study. All the experiments were performed in triplicates. Results are presented as mean ± SEM. SEM, standard error to the mean. p < 0.05 was considered significant. Statistics were performed using paired t-tests. (G) . HBMECs were contact inhibited, arrested at the G0/G1 phase of the cell cycle for 24 h, treated with PDGF-BB ligand for 60 min, immunostained for ARL13B cilia (green) and CENTRIN 2 (red) centriole antibodies, and imaged at 100X using a Keyence BZ-X700 fluorescent microscope (Japan). Scale bar = 5 μm.

Article Snippet: This was followed by blocking in 4% BSA in PBS and overnight incubation with primary antibodies of ARL13B (1:500), IFT88 (1:100), CEP164 (1:100), CENTRIN2 (1:200), NINEIN (Novus biologicals, Cat#NBP2-13657) (1:100), and HsSAS (Santa Cruz Biotechnology, Cat#SC-81431) (1:100).

Techniques: Activation Assay, Control, Immunofluorescence, Staining, Microscopy

CEP164 knockdown in HBMECs. (A) . HBMECs were knocked down for CEP164 siRNA and control siRNA and immunostained for CENTRIN2, a centriole antibody. (B) . CEP164 knockdown was performed in HBMECs and protein expression of CEP164 for knockdown efficiency and CENTRIN2 and ARL13B was performed. (C) . Quantification of the Western blot was performed using ImageJ software. n = 3 in each experimental group. Results are presented as mean ± SEM. SEM, standard error to the mean. Statistics were performed using paired t-tests.

Journal: Frontiers in Molecular Biosciences

Article Title: Brain microvascular endothelial cells possess a second cilium that arises from the daughter centriole

doi: 10.3389/fmolb.2023.1250016

Figure Lengend Snippet: CEP164 knockdown in HBMECs. (A) . HBMECs were knocked down for CEP164 siRNA and control siRNA and immunostained for CENTRIN2, a centriole antibody. (B) . CEP164 knockdown was performed in HBMECs and protein expression of CEP164 for knockdown efficiency and CENTRIN2 and ARL13B was performed. (C) . Quantification of the Western blot was performed using ImageJ software. n = 3 in each experimental group. Results are presented as mean ± SEM. SEM, standard error to the mean. Statistics were performed using paired t-tests.

Article Snippet: This was followed by blocking in 4% BSA in PBS and overnight incubation with primary antibodies of ARL13B (1:500), IFT88 (1:100), CEP164 (1:100), CENTRIN2 (1:200), NINEIN (Novus biologicals, Cat#NBP2-13657) (1:100), and HsSAS (Santa Cruz Biotechnology, Cat#SC-81431) (1:100).

Techniques: Knockdown, Control, Expressing, Western Blot, Software

A , Depletion of DONSON induced four separated centrioles in HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). B, Histograms represent frequency of interphase cells with the indicated phenotypes observed in A. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). C, Depletion of DONSON did not affect the number of centrin foci in G1 phase. HeLa cells were treated with siControl or siDONSON for 48 h, and 10 μM EdU for 30 min before fixation, followed by immunostaining with antibodies against centrin-2 (green) and CENP-F (red). S phase cells were monitored by EdU staining (red). CENPF and EdU double-negative cells were classified as G1 phase cells, and others as S or G2 phase cells. D, Histograms represent frequency of interphase cells with the indicated phenotype observed in C. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). E, Recruitment of C-Nap1 to separated centrioles in DONSON-depleted HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies against centrin-2 (green) and C-Nap1 (red). F, Histograms represent frequency of cells with the indicated number of C-Nap1 positive centrioles among cells with four separated centrioles observed in E. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). G, Time-lapse observation of precocious centriole disengagement in interphase in DONSON-depleted cells. HeLa GFP-centrin-1 cells were visualized every 20 min for 48 h after 24 h treatment of siControl or siDONSON and 3 h treatment of 100 nM SiR-DNA. Left– right arrows indicate precociously disengaged centrioles. H, Schematic model of the phenotypes of DONSON depletion. Depletion of DONSON causes precocious centriole disengagement in interphase. All scale bars, 5 μm. Two-tailed, unpaired Student’s t-test was used in B and D to obtain P value. **, p < 0.01; N.S., not significantly different (p > 0.05).

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , Depletion of DONSON induced four separated centrioles in HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). B, Histograms represent frequency of interphase cells with the indicated phenotypes observed in A. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). C, Depletion of DONSON did not affect the number of centrin foci in G1 phase. HeLa cells were treated with siControl or siDONSON for 48 h, and 10 μM EdU for 30 min before fixation, followed by immunostaining with antibodies against centrin-2 (green) and CENP-F (red). S phase cells were monitored by EdU staining (red). CENPF and EdU double-negative cells were classified as G1 phase cells, and others as S or G2 phase cells. D, Histograms represent frequency of interphase cells with the indicated phenotype observed in C. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). E, Recruitment of C-Nap1 to separated centrioles in DONSON-depleted HeLa cells. HeLa cells were treated with siControl or siDONSON and immunostained with antibodies against centrin-2 (green) and C-Nap1 (red). F, Histograms represent frequency of cells with the indicated number of C-Nap1 positive centrioles among cells with four separated centrioles observed in E. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). G, Time-lapse observation of precocious centriole disengagement in interphase in DONSON-depleted cells. HeLa GFP-centrin-1 cells were visualized every 20 min for 48 h after 24 h treatment of siControl or siDONSON and 3 h treatment of 100 nM SiR-DNA. Left– right arrows indicate precociously disengaged centrioles. H, Schematic model of the phenotypes of DONSON depletion. Depletion of DONSON causes precocious centriole disengagement in interphase. All scale bars, 5 μm. Two-tailed, unpaired Student’s t-test was used in B and D to obtain P value. **, p < 0.01; N.S., not significantly different (p > 0.05).

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques: Immunostaining, Staining, Two Tailed Test

The phenotype induced by DONSON depletion was confirmed by using another siRNAHeLa cells were treated with siRNAs targeting against different sequences of DONSON ORF and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci. Scale bar, 5 μm.

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: The phenotype induced by DONSON depletion was confirmed by using another siRNAHeLa cells were treated with siRNAs targeting against different sequences of DONSON ORF and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci. Scale bar, 5 μm.

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques:

A , Aberrant acquisition of PCM components by precociously disengaged daughter centrioles in DONSON-depleted cells. HeLa cells were immunostained with antibodies against centrin-2 (green) and PCNT (red). B, Histograms represent frequency of DONSON-depleted cells with the indicated number of PCNT foci among cells with four separated centrioles in interphase. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). C, Microtubule regrowth assay after 10 μM Nocodazole treatment for 3 h and subsequent ice-cold treatment for 1 h in DONSON-depleted cells. HeLa cells were immunostained with antibodies against EB1 (green) and Cep192 (red). White arrowheads indicate centrosomes with MTOC activity. Arrowheads with white outline indicate centrosomes without microtubule nucleation. D, Centriole reduplication after depletion of DONSON in HeLa cells. HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with the indicated antibodies. E, Histograms represent frequency of interphase cells with the indicated number of HsSAS-6 foci among cells with four separated centrioles observed in D. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). F, HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). G, Histograms represent frequency of interphase cells with the indicated phenotypes observed in F. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). Two-tailed, unpaired Student’s t-test was used to obtain P value. *, p < 0.05. H, Schematic model of the phenotype of DONSON depletion observed in A, D and F. Depletion of DONSON led to centriole disengagement, centriole-to-centrosome conversion, acquisition of ectopic MTOC activity and centriole amplification in interphase. All scale bars, 5 μm.

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , Aberrant acquisition of PCM components by precociously disengaged daughter centrioles in DONSON-depleted cells. HeLa cells were immunostained with antibodies against centrin-2 (green) and PCNT (red). B, Histograms represent frequency of DONSON-depleted cells with the indicated number of PCNT foci among cells with four separated centrioles in interphase. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). C, Microtubule regrowth assay after 10 μM Nocodazole treatment for 3 h and subsequent ice-cold treatment for 1 h in DONSON-depleted cells. HeLa cells were immunostained with antibodies against EB1 (green) and Cep192 (red). White arrowheads indicate centrosomes with MTOC activity. Arrowheads with white outline indicate centrosomes without microtubule nucleation. D, Centriole reduplication after depletion of DONSON in HeLa cells. HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with the indicated antibodies. E, Histograms represent frequency of interphase cells with the indicated number of HsSAS-6 foci among cells with four separated centrioles observed in D. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). F, HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). G, Histograms represent frequency of interphase cells with the indicated phenotypes observed in F. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). Two-tailed, unpaired Student’s t-test was used to obtain P value. *, p < 0.05. H, Schematic model of the phenotype of DONSON depletion observed in A, D and F. Depletion of DONSON led to centriole disengagement, centriole-to-centrosome conversion, acquisition of ectopic MTOC activity and centriole amplification in interphase. All scale bars, 5 μm.

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques: Regrowth Assay, Activity Assay, Two Tailed Test, Amplification

A , Ectopic acquisition of PCM component after depletion of DONSON in HeLa cells. HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with antibodies against centrin-2 (green) and γ-Tubulin (red). B, Cells with aberrant number of diplosomes after DONSON depletion. HeLa cells were treated with siRNA for 48 h and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). All scale bars, 5 μm.

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , Ectopic acquisition of PCM component after depletion of DONSON in HeLa cells. HeLa cells were treated with siControl or siDONSON for 48 h and immunostained with antibodies against centrin-2 (green) and γ-Tubulin (red). B, Cells with aberrant number of diplosomes after DONSON depletion. HeLa cells were treated with siRNA for 48 h and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). All scale bars, 5 μm.

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques:

A , Centriole disengagement caused by DONSON depletion in HeLa and U2OS cells. The cells were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). B, Histograms represent frequency of interphase cells with the indicated phenotypes observed in A. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). C, p53-dependent centrosome clustering in DONSON-depleted U2OS cells. U2OS cells were treated with the indicated siRNAs for 48 h and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). D, Histograms represent frequency of interphase cells with the indicated phenotypes observed in C. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). E, Disruption of cortical actin cytoskeleton reduced centrosome clustering in U2OS cells. U2OS cells were treated with siControl or siDONSON for 48 h and DMSO or Crenolanib for 48 h. Arrowheads indicate Cep192 foci (centrosomes). F, Histograms represent frequency of interphase cells with the indicated phenotypes observed in E. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). G, Schematic model of p53-dependent centrosome clustering after ectopic centrosome conversion in DONSON-depleted cells. All scale bars, 5 μm. All the squares with white dotted lines are 2.5 μm square. Tukey’s multiple comparison test was used in B, D, and F to obtain P value. *, p < 0.05; **, p < 0.01.

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , Centriole disengagement caused by DONSON depletion in HeLa and U2OS cells. The cells were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). B, Histograms represent frequency of interphase cells with the indicated phenotypes observed in A. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). C, p53-dependent centrosome clustering in DONSON-depleted U2OS cells. U2OS cells were treated with the indicated siRNAs for 48 h and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate Cep192 foci (centrosomes). D, Histograms represent frequency of interphase cells with the indicated phenotypes observed in C. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). E, Disruption of cortical actin cytoskeleton reduced centrosome clustering in U2OS cells. U2OS cells were treated with siControl or siDONSON for 48 h and DMSO or Crenolanib for 48 h. Arrowheads indicate Cep192 foci (centrosomes). F, Histograms represent frequency of interphase cells with the indicated phenotypes observed in E. Values are mean percentages ± s.d. from three independent experiments (n = 50 for each experiment). G, Schematic model of p53-dependent centrosome clustering after ectopic centrosome conversion in DONSON-depleted cells. All scale bars, 5 μm. All the squares with white dotted lines are 2.5 μm square. Tukey’s multiple comparison test was used in B, D, and F to obtain P value. *, p < 0.05; **, p < 0.01.

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques:

A , The distance between disengaged centrioles was different in interphase, depending on the cell lines. RPE-1 and PANC-1 cells were treated with siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate precociously disengaged centrioles. B, Histograms represent frequency of interphase cells with the indicated phenotype observed in A. (two independent experiments, N=50 for each experiment). C, The phenotype induced by p53 depletion was confirmed by using another siRNA. U2OS cells were treated with DONSON siRNA and siRNAs targeting against different sequences of TP53 ORF and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate precociously disengaged centrioles. All scale bars, 5 μm.

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , The distance between disengaged centrioles was different in interphase, depending on the cell lines. RPE-1 and PANC-1 cells were treated with siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate precociously disengaged centrioles. B, Histograms represent frequency of interphase cells with the indicated phenotype observed in A. (two independent experiments, N=50 for each experiment). C, The phenotype induced by p53 depletion was confirmed by using another siRNA. U2OS cells were treated with DONSON siRNA and siRNAs targeting against different sequences of TP53 ORF and were immunostained with antibodies against centrin-2 (green) and Cep192 (red). Arrowheads indicate precociously disengaged centrioles. All scale bars, 5 μm.

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques:

A , Chromosome segregation errors observed in DONSON-depleted HeLa cells. The cells were treated with siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). B, Histograms represent frequency of mitotic cells with the indicated phenotype observed in A. Values are mean percentages ± s.d. from three independent experiments (N=30 for each experiment). C, Crenolanib treatment increased the number of mitotic cells with multi-polar spindles in DONSON-depleted U2OS cells. U2OS cells were treated with siRNA, followed by treatment with DMSO or Crenolanib (500 nM) for 6 h. The cells were immunostained as in A. D, Histograms represent frequency of mitotic cells with the indicated phenotypes observed in C. Values are mean percentages ± s.d. from three independent experiments (N=30 for each experiment). All scale bars, 5 μm. Two-tailed, unpaired Student’s t-test was used in B, and Tukey’s multiple comparison test was used in D to obtain P value. *, p < 0.05; **, p < 0.01; N.S., not significantly different (p > 0.05).

Journal: bioRxiv

Article Title: DONSON , a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

doi: 10.1101/2020.05.10.086777

Figure Lengend Snippet: A , Chromosome segregation errors observed in DONSON-depleted HeLa cells. The cells were treated with siDONSON and immunostained with antibodies against centrin-2 (green) and Cep192 (red). B, Histograms represent frequency of mitotic cells with the indicated phenotype observed in A. Values are mean percentages ± s.d. from three independent experiments (N=30 for each experiment). C, Crenolanib treatment increased the number of mitotic cells with multi-polar spindles in DONSON-depleted U2OS cells. U2OS cells were treated with siRNA, followed by treatment with DMSO or Crenolanib (500 nM) for 6 h. The cells were immunostained as in A. D, Histograms represent frequency of mitotic cells with the indicated phenotypes observed in C. Values are mean percentages ± s.d. from three independent experiments (N=30 for each experiment). All scale bars, 5 μm. Two-tailed, unpaired Student’s t-test was used in B, and Tukey’s multiple comparison test was used in D to obtain P value. *, p < 0.05; **, p < 0.01; N.S., not significantly different (p > 0.05).

Article Snippet: The following primary antibodies were used in this study: rabbit antibodies against Cep192 (Bethyl Laboratories, A302–324 A, IF 1:1000), CP110 (Proteintech, 12780-1-AP, IF 1:500), C-Nap1 (a gift from Erich Nigg IF 1:1000), CENP-F (Bethyl Laboratories, A301–617 A IF 1:1000), Cep152 (Bethyl Laboratories, A302–480A, IF 1:1000), PCNT (Abcam, ab4448, IF 1:2000), and γ-Tubulin (Merck, T5192, IF 1:1000),; mouse antibodies against centrin-2 (Merck, 20H5, IF 1:1000), HsSAS-6 (Santa Cruze Biotechnology, sc-81431, IF 1:1000), EB1 (BD Transduction Laboratories, 610534, IF 1:500), α-Tubulin (Merck, DM1A, IF 1:1000), GFP (Invitrogen, A-11120, IF 1:1000), and FLAG-tag (Merck, F1804, IF 1:1000).

Techniques: Two Tailed Test