polyclonal rabbit anti rpap3 antibodies Search Results


anti rpap3  (Proteintech)
92
Proteintech anti rpap3
Anti Rpap3, supplied by Proteintech, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/anti rpap3/product/Proteintech
Average 92 stars, based on 1 article reviews
anti rpap3 - by Bioz Stars, 2026-03
92/100 stars
  Buy from Supplier
rpap3 (sab1411438) rabbit 1/1000  (Millipore)
90
Millipore rpap3 (sab1411438) rabbit 1/1000
(A) Schematic representation of R2TP with its four subunits in different tones of blue <t>(RPAP3,</t> PIH1D1 and the RUVBL1/2 hetero-hexamer). RPAP3 is the core subunit that contacts directly HSP90, PIH1D1 and RUVBL1/2. (B) β galactosidase activity in Rpap3 wtsi/+ small intestines (top) and colon (bottom), as compared to negative controls (n=2). Scale bars are identical for all images. (C) Schematic representation of the experimental setup. Littermates aged of 8 weeks receive two sequential intra-peritoneal injections of tamoxifen at the indicated time. (D) Depletion of Rpap3 after tamoxifen injection. Western blots were revealed with antibodies against the indicated proteins in extracts of the jejunum and the colonic epitheliums of Rpap3 flox/flox controls (blue) or VilCreER T2 ; Rpap3 flox/flox animals (red), 5 days after the first tamoxifen injection. Each lane was loaded with the lysate obtained from a single animal. Molecular sizes are indicated on the right. (E) Individual weight variations in females (top panel,) or males (bottom panel) for VilCreER T2 ; Rpap3 flox/flox animals (red curves, n= 6 females and n=5 males) and Rpap3 flox/flox controls (blue curves, n=8 females and n=7 males). Individual weight was set at 100 for each animal at day 0. (F) Length of small intestines and colons from VilCreER T2 ; Rpap3 flox/flox mice (red bars) and controls (blue bars). Females were measured at day 8 (top, n=3), and males at day 10 (bottom, n=4). (G) Small intestines (white arrowheads) from VilCreER T2 ; Rpap3 flox/flox animals were filled with liquid (left panel) or blood (right panel) from day 8 to 10; a control animal is shown above.
Rpap3 (Sab1411438) Rabbit 1/1000, supplied by Millipore, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rpap3 (sab1411438) rabbit 1/1000/product/Millipore
Average 90 stars, based on 1 article reviews
rpap3 (sab1411438) rabbit 1/1000 - by Bioz Stars, 2026-03
90/100 stars
  Buy from Supplier
polyclonal rabbit anti rpap3 antibodies  (Bethyl)
93
Bethyl polyclonal rabbit anti rpap3 antibodies
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Polyclonal Rabbit Anti Rpap3 Antibodies, supplied by Bethyl, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/polyclonal rabbit anti rpap3 antibodies/product/Bethyl
Average 93 stars, based on 1 article reviews
polyclonal rabbit anti rpap3 antibodies - by Bioz Stars, 2026-03
93/100 stars
  Buy from Supplier
ab9485  (Abcam)
99
Abcam ab9485
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Ab9485, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/ab9485/product/Abcam
Average 99 stars, based on 1 article reviews
ab9485 - by Bioz Stars, 2026-03
99/100 stars
  Buy from Supplier
anti dpcd  (Proteintech)
92
Proteintech anti dpcd
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Anti Dpcd, supplied by Proteintech, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/anti dpcd/product/Proteintech
Average 92 stars, based on 1 article reviews
anti dpcd - by Bioz Stars, 2026-03
92/100 stars
  Buy from Supplier
rabbit monoclonal anti tsc1  (Cell Signaling Technology Inc)
95
Cell Signaling Technology Inc rabbit monoclonal anti tsc1
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Rabbit Monoclonal Anti Tsc1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rabbit monoclonal anti tsc1/product/Cell Signaling Technology Inc
Average 95 stars, based on 1 article reviews
rabbit monoclonal anti tsc1 - by Bioz Stars, 2026-03
95/100 stars
  Buy from Supplier
rabbit monoclonal anti tsc2  (Cell Signaling Technology Inc)
96
Cell Signaling Technology Inc rabbit monoclonal anti tsc2
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Rabbit Monoclonal Anti Tsc2, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rabbit monoclonal anti tsc2/product/Cell Signaling Technology Inc
Average 96 stars, based on 1 article reviews
rabbit monoclonal anti tsc2 - by Bioz Stars, 2026-03
96/100 stars
  Buy from Supplier
mouse anti-gst  (EASY BIO Inc)
90
EASY BIO Inc mouse anti-gst
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Mouse Anti Gst, supplied by EASY BIO Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/mouse anti-gst/product/EASY BIO Inc
Average 90 stars, based on 1 article reviews
mouse anti-gst - by Bioz Stars, 2026-03
90/100 stars
  Buy from Supplier
anti cp110  (Proteintech)
95
Proteintech anti cp110
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Anti Cp110, supplied by Proteintech, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/anti cp110/product/Proteintech
Average 95 stars, based on 1 article reviews
anti cp110 - by Bioz Stars, 2026-03
95/100 stars
  Buy from Supplier
8599s primary antibodies  (Cell Signaling Technology Inc)
95
Cell Signaling Technology Inc 8599s primary antibodies
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
8599s Primary Antibodies, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/8599s primary antibodies/product/Cell Signaling Technology Inc
Average 95 stars, based on 1 article reviews
8599s primary antibodies - by Bioz Stars, 2026-03
95/100 stars
  Buy from Supplier
1c12 cst2524s mouse  (Cell Signaling Technology Inc)
96
Cell Signaling Technology Inc 1c12 cst2524s mouse
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
1c12 Cst2524s Mouse, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/1c12 cst2524s mouse/product/Cell Signaling Technology Inc
Average 96 stars, based on 1 article reviews
1c12 cst2524s mouse - by Bioz Stars, 2026-03
96/100 stars
  Buy from Supplier
rabbit anti ruvbl2  (Proteintech)
93
Proteintech rabbit anti ruvbl2
Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, <t>RPAP3,</t> MIA3, EIF5AL1, and TRMT6.
Rabbit Anti Ruvbl2, 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
https://www.bioz.com/result/rabbit anti ruvbl2/product/Proteintech
Average 93 stars, based on 1 article reviews
rabbit anti ruvbl2 - by Bioz Stars, 2026-03
93/100 stars
  Buy from Supplier

Image Search Results


(A) Schematic representation of R2TP with its four subunits in different tones of blue (RPAP3, PIH1D1 and the RUVBL1/2 hetero-hexamer). RPAP3 is the core subunit that contacts directly HSP90, PIH1D1 and RUVBL1/2. (B) β galactosidase activity in Rpap3 wtsi/+ small intestines (top) and colon (bottom), as compared to negative controls (n=2). Scale bars are identical for all images. (C) Schematic representation of the experimental setup. Littermates aged of 8 weeks receive two sequential intra-peritoneal injections of tamoxifen at the indicated time. (D) Depletion of Rpap3 after tamoxifen injection. Western blots were revealed with antibodies against the indicated proteins in extracts of the jejunum and the colonic epitheliums of Rpap3 flox/flox controls (blue) or VilCreER T2 ; Rpap3 flox/flox animals (red), 5 days after the first tamoxifen injection. Each lane was loaded with the lysate obtained from a single animal. Molecular sizes are indicated on the right. (E) Individual weight variations in females (top panel,) or males (bottom panel) for VilCreER T2 ; Rpap3 flox/flox animals (red curves, n= 6 females and n=5 males) and Rpap3 flox/flox controls (blue curves, n=8 females and n=7 males). Individual weight was set at 100 for each animal at day 0. (F) Length of small intestines and colons from VilCreER T2 ; Rpap3 flox/flox mice (red bars) and controls (blue bars). Females were measured at day 8 (top, n=3), and males at day 10 (bottom, n=4). (G) Small intestines (white arrowheads) from VilCreER T2 ; Rpap3 flox/flox animals were filled with liquid (left panel) or blood (right panel) from day 8 to 10; a control animal is shown above.

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) Schematic representation of R2TP with its four subunits in different tones of blue (RPAP3, PIH1D1 and the RUVBL1/2 hetero-hexamer). RPAP3 is the core subunit that contacts directly HSP90, PIH1D1 and RUVBL1/2. (B) β galactosidase activity in Rpap3 wtsi/+ small intestines (top) and colon (bottom), as compared to negative controls (n=2). Scale bars are identical for all images. (C) Schematic representation of the experimental setup. Littermates aged of 8 weeks receive two sequential intra-peritoneal injections of tamoxifen at the indicated time. (D) Depletion of Rpap3 after tamoxifen injection. Western blots were revealed with antibodies against the indicated proteins in extracts of the jejunum and the colonic epitheliums of Rpap3 flox/flox controls (blue) or VilCreER T2 ; Rpap3 flox/flox animals (red), 5 days after the first tamoxifen injection. Each lane was loaded with the lysate obtained from a single animal. Molecular sizes are indicated on the right. (E) Individual weight variations in females (top panel,) or males (bottom panel) for VilCreER T2 ; Rpap3 flox/flox animals (red curves, n= 6 females and n=5 males) and Rpap3 flox/flox controls (blue curves, n=8 females and n=7 males). Individual weight was set at 100 for each animal at day 0. (F) Length of small intestines and colons from VilCreER T2 ; Rpap3 flox/flox mice (red bars) and controls (blue bars). Females were measured at day 8 (top, n=3), and males at day 10 (bottom, n=4). (G) Small intestines (white arrowheads) from VilCreER T2 ; Rpap3 flox/flox animals were filled with liquid (left panel) or blood (right panel) from day 8 to 10; a control animal is shown above.

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Activity Assay, Injection, Western Blot

(A) Micrographs are tissue sections stained for Olfm, in the crypts from controls animals (left), or VilCreER T2 ; Rpap3 flox/flox animals 7 days after the first tamoxifen injection (right). (B) Staining for Olfm4 in the jejunum from controls (top panel) and VilCreER T2 ; Rpap3 flox/flox animals 6 to 8 days after the first tamoxifen injection. Scale is identical in all images (n=2 to 4 animals/time point from at least two independent experiments). (C) Schematic representation of a crypt from the small intestine, with CBC stem cells (in green) sandwiched between Paneth cells (in brown) and progenitors forming the TA on top (purple). (D) Micrographs are tissue sections stained for lysozyme, a specific marker of Paneth cells, in the jejunum of controls (top) and VilCreER T2 ; Rpap3 flox/flox mice from day 6 to day 8 (black arrows). Panels are representative for 2-3 animals/time-point from two independent experiments. The scale bar is identical in all pictures. (E) Micrographs are tissue sections stained for cleaved caspase 3 (cleaved cas3), a marker of apoptosis, in the jejunum. In control animals, cleaved cas3 + cells (brown arrows) are mainly detected at the tip of the villi, as a result of epithelium turnover (top panel). In the jejunum from VilCreER T2 ; Rpap3 flox/flox animals, at day 6 and 7, cleaved caspase 3 + cells were detected within the crypts(brown arrows). Panels are representative from 2 to 4 animals/time point, from two independent experiments. The graph on top of the column shows the total number of apoptotic cells at day 6, divided by the surface of the jejunum for each mouse analyzed. Mean values with S.E.M are indicated for each experimental group. Unpaired two-tailed t-test with Welch’s correction indicates significant difference between controls and VilCreER T2 ; Rpap3 flox/flox animals (p=0, 0384; t=3,538, df=3, n=4). (F) Micrographs are tissue sections stained for p53 by immunofluorescence in controls (left) and VilCreER T2 ; Rpap3 flox/flox mice (right) at day 6 (n=4). Dapi: nuclei.

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) Micrographs are tissue sections stained for Olfm, in the crypts from controls animals (left), or VilCreER T2 ; Rpap3 flox/flox animals 7 days after the first tamoxifen injection (right). (B) Staining for Olfm4 in the jejunum from controls (top panel) and VilCreER T2 ; Rpap3 flox/flox animals 6 to 8 days after the first tamoxifen injection. Scale is identical in all images (n=2 to 4 animals/time point from at least two independent experiments). (C) Schematic representation of a crypt from the small intestine, with CBC stem cells (in green) sandwiched between Paneth cells (in brown) and progenitors forming the TA on top (purple). (D) Micrographs are tissue sections stained for lysozyme, a specific marker of Paneth cells, in the jejunum of controls (top) and VilCreER T2 ; Rpap3 flox/flox mice from day 6 to day 8 (black arrows). Panels are representative for 2-3 animals/time-point from two independent experiments. The scale bar is identical in all pictures. (E) Micrographs are tissue sections stained for cleaved caspase 3 (cleaved cas3), a marker of apoptosis, in the jejunum. In control animals, cleaved cas3 + cells (brown arrows) are mainly detected at the tip of the villi, as a result of epithelium turnover (top panel). In the jejunum from VilCreER T2 ; Rpap3 flox/flox animals, at day 6 and 7, cleaved caspase 3 + cells were detected within the crypts(brown arrows). Panels are representative from 2 to 4 animals/time point, from two independent experiments. The graph on top of the column shows the total number of apoptotic cells at day 6, divided by the surface of the jejunum for each mouse analyzed. Mean values with S.E.M are indicated for each experimental group. Unpaired two-tailed t-test with Welch’s correction indicates significant difference between controls and VilCreER T2 ; Rpap3 flox/flox animals (p=0, 0384; t=3,538, df=3, n=4). (F) Micrographs are tissue sections stained for p53 by immunofluorescence in controls (left) and VilCreER T2 ; Rpap3 flox/flox mice (right) at day 6 (n=4). Dapi: nuclei.

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Staining, Injection, Marker, Two Tailed Test, Immunofluorescence

(A) Schematic representation of the experimental setting: 8-week-old mice of the indicated genotype received 2 sequential injections of tamoxifen 24h apart, and were analyzed 5 to 8 days after the first injection. 2 hours before each sacrifice, BrdU was injected intra-peritoneally to detect cells in S-phase (thin arrows). (B) Pictures of jejunum tissue sections stained with HE (left) or by IHC with anti-Ki67 antibodies (right panels – Ki67 signal is brown) at different days after the first tamoxifen injection. Black arrowheads indicate remnants of crypt glands with Ki67 + cells observed at day 8. Scale bar is shown in control HE panel. Each panel is representative of 8 to 12 animals from three independent experiments. (C) Pictures are jejunum tissue section of controls (boxed in blue) and VilCreER T2 ; Rpap3 flox/flox animals (boxed in red), stained by IHC with anti-BrdU antibodies (n=3-6 animals/ time point from two independent experiments). Scale bar is shown in control panel.

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) Schematic representation of the experimental setting: 8-week-old mice of the indicated genotype received 2 sequential injections of tamoxifen 24h apart, and were analyzed 5 to 8 days after the first injection. 2 hours before each sacrifice, BrdU was injected intra-peritoneally to detect cells in S-phase (thin arrows). (B) Pictures of jejunum tissue sections stained with HE (left) or by IHC with anti-Ki67 antibodies (right panels – Ki67 signal is brown) at different days after the first tamoxifen injection. Black arrowheads indicate remnants of crypt glands with Ki67 + cells observed at day 8. Scale bar is shown in control HE panel. Each panel is representative of 8 to 12 animals from three independent experiments. (C) Pictures are jejunum tissue section of controls (boxed in blue) and VilCreER T2 ; Rpap3 flox/flox animals (boxed in red), stained by IHC with anti-BrdU antibodies (n=3-6 animals/ time point from two independent experiments). Scale bar is shown in control panel.

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Injection, Staining

(A) Western blot analysis of preparations enriched for epithelial crypt cells from the jejunum of animals, sacrificed 6 days after the first tamoxifen injection. NOP58, PRP8, ATR and mTOR and TRRAP were detected with specific antibodies. Tubulin and GAPDH were used as loading controls. Each lane was loaded with the lysate obtained from one animal of the indicated genotype (6 animals per gel). Molecular weights are indicated on the right. See Supplemental Figure 4 for an expanded view of the membranes and signal quantification. (B) Images are tissue sections of small intestines stained by immunohistochemistry (IHC) for Rpb1, the large subunit of RNA polymerase II, from control Rpap3 flox/flox mice (blue frame, left panel), or VilCreER T2; Rpap3 flox/flox animals at day 6 (red frame, right panel). Note that the staining in stromal cells is nuclear in both wild-type and VilCreER T2 ; Rpap3 flox/flox animals (stromal cells do not express the Cre), while it becomes cytoplasmic in the mutant epithelium. The scale is identical for both pictures. Panels are representative for n=6 animals from three independent experiments. (C) Magnification of crypts from (B). The scale bar is identical for both images. (D) Schematic interpretation of the IHC in (B, C). In control epithelial cells, R2TP incorporates Rpb1 into RNA PolII, which is then imported into the nucleus. In the absence of Rpap3, neo-synthesized Rpb1 accumulates in the cytoplasm.

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) Western blot analysis of preparations enriched for epithelial crypt cells from the jejunum of animals, sacrificed 6 days after the first tamoxifen injection. NOP58, PRP8, ATR and mTOR and TRRAP were detected with specific antibodies. Tubulin and GAPDH were used as loading controls. Each lane was loaded with the lysate obtained from one animal of the indicated genotype (6 animals per gel). Molecular weights are indicated on the right. See Supplemental Figure 4 for an expanded view of the membranes and signal quantification. (B) Images are tissue sections of small intestines stained by immunohistochemistry (IHC) for Rpb1, the large subunit of RNA polymerase II, from control Rpap3 flox/flox mice (blue frame, left panel), or VilCreER T2; Rpap3 flox/flox animals at day 6 (red frame, right panel). Note that the staining in stromal cells is nuclear in both wild-type and VilCreER T2 ; Rpap3 flox/flox animals (stromal cells do not express the Cre), while it becomes cytoplasmic in the mutant epithelium. The scale is identical for both pictures. Panels are representative for n=6 animals from three independent experiments. (C) Magnification of crypts from (B). The scale bar is identical for both images. (D) Schematic interpretation of the IHC in (B, C). In control epithelial cells, R2TP incorporates Rpb1 into RNA PolII, which is then imported into the nucleus. In the absence of Rpap3, neo-synthesized Rpb1 accumulates in the cytoplasm.

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Western Blot, Injection, Staining, Immunohistochemistry, Mutagenesis, Synthesized

(A) Schematic representation of the experimental setting. Eight-week-old Lgr5-GFP-IRES-CreER T2 ; Rpap3 flox/flox mice received five sequential intra-peritoneal injections of tamoxifen and were analyzed 7 or 10 days after the first injection. In this genetic model, the Cre is expressed in the Lgr5 + CBC stem cells labeled by GFP (see scheme on the right). (B) Images are tissue sections labelled by immunofluorescence with antibodies against GFP (green) and Rpb1 (red), with dapi counter-staining of nuclei (blue). White arrow: GFP+ crypts. Asterisks: GFP-crypts. At day 7, control Lgr5-GFP-IRES-CreER T2 ; Rpap3 +/+ animals do not show detectable Rpb1 signal in the cytoplasm (0/29 GFP+ and 0/96 GFP - crypts; n=2). In the jejunum of Lgr5-GFP-IRES-CreER T2 ; Rpap3 flox/flox animals (n=3), GFP + crypts show detectable Rpb1 signal in the cytoplasm (50/50). In contrast, adjacent GFP - crypts, which do not express CreER T2 , do not show cytoplasmic Rpb1 staining (0/136, white asterisk). At day 10, in the colons of Lgr5-GFP-IRES-CreER T2 ; RPAP3 flox/flox animals (n=2), Rpb1 is detected in the cytoplasm of CBC stem cells and progenitors in the GFP + crypts (white arrow; 11/11). In contrast, Rpb1 was strictly nuclear in the neighboring GFP - crypts (white asterisk, 0/15). Scale bars are identical for matching panels. (C) Images are intestine tissue section of wild-type animals stained for BrdU. Animals received one BrdU injection and were sacrificed at the indicated time point. Scale bars are identical in each row. The experiment was repeated twice (2 to 4 animals/time-point/ experiment).

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) Schematic representation of the experimental setting. Eight-week-old Lgr5-GFP-IRES-CreER T2 ; Rpap3 flox/flox mice received five sequential intra-peritoneal injections of tamoxifen and were analyzed 7 or 10 days after the first injection. In this genetic model, the Cre is expressed in the Lgr5 + CBC stem cells labeled by GFP (see scheme on the right). (B) Images are tissue sections labelled by immunofluorescence with antibodies against GFP (green) and Rpb1 (red), with dapi counter-staining of nuclei (blue). White arrow: GFP+ crypts. Asterisks: GFP-crypts. At day 7, control Lgr5-GFP-IRES-CreER T2 ; Rpap3 +/+ animals do not show detectable Rpb1 signal in the cytoplasm (0/29 GFP+ and 0/96 GFP - crypts; n=2). In the jejunum of Lgr5-GFP-IRES-CreER T2 ; Rpap3 flox/flox animals (n=3), GFP + crypts show detectable Rpb1 signal in the cytoplasm (50/50). In contrast, adjacent GFP - crypts, which do not express CreER T2 , do not show cytoplasmic Rpb1 staining (0/136, white asterisk). At day 10, in the colons of Lgr5-GFP-IRES-CreER T2 ; RPAP3 flox/flox animals (n=2), Rpb1 is detected in the cytoplasm of CBC stem cells and progenitors in the GFP + crypts (white arrow; 11/11). In contrast, Rpb1 was strictly nuclear in the neighboring GFP - crypts (white asterisk, 0/15). Scale bars are identical for matching panels. (C) Images are intestine tissue section of wild-type animals stained for BrdU. Animals received one BrdU injection and were sacrificed at the indicated time point. Scale bars are identical in each row. The experiment was repeated twice (2 to 4 animals/time-point/ experiment).

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Injection, Labeling, Immunofluorescence, Staining

(A) The graph depicts transcript levels of R2TP components in human primary colorectal tumor samples (n = 380; COADREAD cohort), as compared to normal solid tissues (n = 51). y-axis: Log2 normalized counts for the indicated transcript. Distributions are presented as box-and-whisker plots (center line: median; box limits, first and third quartiles; whiskers, 10th and 90th percentiles). Statistical significance was determined by one-way ANOVA (asterisk: P < 0.001). (B) Kaplan–Meier analysis of disease-free survival among 177 CRC patients according to the proportion of RPAP3-expressing cells in tumor tissues (C, P = 0.037). Solid green line and dashed blue line indicate High and Low proportion of RPAP3-expressing tumoral cells, respectively. Right panels show examples of CRC tissues with low (top) or high (bottom) RPAP3 expression. (C) Proposed model for R2TP activity in the small and large intestine. R2TP assembles cellular machineries such as RNA polymerases, snoRNPs, snRNPs and PIKKs-complexes in CBCs and progenitors in the proliferative compartment (blue cells). Differentiated cells (including Paneth cells from the small intestine crypts, in pink) mostly rely on the complexes assembled during the proliferative phase. A defect in R2TP activity induces client dysfunction, cell cycle arrest and apoptosis via p53, and eventually, epithelium degradation.

Journal: bioRxiv

Article Title: The R2TP chaperone assembles cellular machineries in intestinal CBC stem cells and progenitors

doi: 10.1101/2019.12.19.882712

Figure Lengend Snippet: (A) The graph depicts transcript levels of R2TP components in human primary colorectal tumor samples (n = 380; COADREAD cohort), as compared to normal solid tissues (n = 51). y-axis: Log2 normalized counts for the indicated transcript. Distributions are presented as box-and-whisker plots (center line: median; box limits, first and third quartiles; whiskers, 10th and 90th percentiles). Statistical significance was determined by one-way ANOVA (asterisk: P < 0.001). (B) Kaplan–Meier analysis of disease-free survival among 177 CRC patients according to the proportion of RPAP3-expressing cells in tumor tissues (C, P = 0.037). Solid green line and dashed blue line indicate High and Low proportion of RPAP3-expressing tumoral cells, respectively. Right panels show examples of CRC tissues with low (top) or high (bottom) RPAP3 expression. (C) Proposed model for R2TP activity in the small and large intestine. R2TP assembles cellular machineries such as RNA polymerases, snoRNPs, snRNPs and PIKKs-complexes in CBCs and progenitors in the proliferative compartment (blue cells). Differentiated cells (including Paneth cells from the small intestine crypts, in pink) mostly rely on the complexes assembled during the proliferative phase. A defect in R2TP activity induces client dysfunction, cell cycle arrest and apoptosis via p53, and eventually, epithelium degradation.

Article Snippet: Primary antibodies from Cell Signaling: ATM D2E2 (CST 2873S) rabbit at 1/1000, ATR E1S3S (CST 13934S) rabbit at 1/1000, mTOR (CST 2972S) rabbit 1/1000, p53 1C12 (CST2524S) mouse 1/1000; PRP8 (sc-30207, Santa Cruz) rabbit 1/200; from ABCAM: EFTUD2 (ab72456) rabbit 1/2000, GAPDH (ab8245 6C5) mouse 1/10000; and from Sigma: NOP58 (HPA018472) rabbit 1/100, RPAP3 (SAB1411438) rabbit 1/1000, Tubulin (I2G10) mouse 1/500, TRRAP 2TRR-1B3 (MABE1008) mouse 1/1000.

Techniques: Whisker Assay, Expressing, Activity Assay

Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, RPAP3, MIA3, EIF5AL1, and TRMT6.

Journal: Marine Drugs

Article Title: Target Identification of the Marine Natural Products Dictyoceratin-A and -C as Selective Growth Inhibitors in Cancer Cells Adapted to Hypoxic Environments

doi: 10.3390/md17030163

Figure Lengend Snippet: Binding of probe A ( 3 ) to the candidate proteins in cell lysates. Cell lysates (100 μg protein each), prepared from DU145 cells cultured under hypoxic conditions (H, lanes 2, 4, and 6) or normoxic conditions (N, lanes 1, 3, and 5) were incubated with probe A ( 3 ). The probe with bound protein was then separated from the supernatant by the addition of streptavidin-conjugated Dynabeads. The resulting supernatants and Dynabeads were used for western blotting analysis to detect RBM28, RPAP3, MIA3, EIF5AL1, and TRMT6.

Article Snippet: Polyclonal rabbit anti-RPAP3 antibodies were obtained from Bethyl Laboratories, Inc. (Montgomery, TX, USA).

Techniques: Binding Assay, Cell Culture, Incubation, Western Blot

Growth of RBM28-, RPAP3-, and MIA30-knockdown DU145 cells under normoxic and hypoxic conditions. ( a ) The expression levels of the indicated proteins in each knockdown cell line were analyzed by western blotting. The expression level of each protein was calculated from its relative density, which was normalized to that of β-actin. ( b ) The proliferation rates of MIA3-, RBM28-, and RPAP3-knockdown DU145 cells were investigated under normoxic and hypoxic conditions. The growth inhibition rate was calculated as a percentage relative to the negative control cells. Differences were considered significant at * p < 0.01.

Journal: Marine Drugs

Article Title: Target Identification of the Marine Natural Products Dictyoceratin-A and -C as Selective Growth Inhibitors in Cancer Cells Adapted to Hypoxic Environments

doi: 10.3390/md17030163

Figure Lengend Snippet: Growth of RBM28-, RPAP3-, and MIA30-knockdown DU145 cells under normoxic and hypoxic conditions. ( a ) The expression levels of the indicated proteins in each knockdown cell line were analyzed by western blotting. The expression level of each protein was calculated from its relative density, which was normalized to that of β-actin. ( b ) The proliferation rates of MIA3-, RBM28-, and RPAP3-knockdown DU145 cells were investigated under normoxic and hypoxic conditions. The growth inhibition rate was calculated as a percentage relative to the negative control cells. Differences were considered significant at * p < 0.01.

Article Snippet: Polyclonal rabbit anti-RPAP3 antibodies were obtained from Bethyl Laboratories, Inc. (Montgomery, TX, USA).

Techniques: Knockdown, Expressing, Western Blot, Inhibition, Negative Control

Expression of hypoxia-related proteins in RPAP3-knockdown DU145 cells. The expression levels of the indicated hypoxia-related proteins in RPAP3-knockdown DU145 cells were compared with those of wild-type DU145 cells treated with/without dictyoceratin-A ( 1 ) or -C ( 2 ) (10 µM each) using western blotting analysis. The expression level of each protein was calculated from its relative density, which was normalized to that of β-actin.

Journal: Marine Drugs

Article Title: Target Identification of the Marine Natural Products Dictyoceratin-A and -C as Selective Growth Inhibitors in Cancer Cells Adapted to Hypoxic Environments

doi: 10.3390/md17030163

Figure Lengend Snippet: Expression of hypoxia-related proteins in RPAP3-knockdown DU145 cells. The expression levels of the indicated hypoxia-related proteins in RPAP3-knockdown DU145 cells were compared with those of wild-type DU145 cells treated with/without dictyoceratin-A ( 1 ) or -C ( 2 ) (10 µM each) using western blotting analysis. The expression level of each protein was calculated from its relative density, which was normalized to that of β-actin.

Article Snippet: Polyclonal rabbit anti-RPAP3 antibodies were obtained from Bethyl Laboratories, Inc. (Montgomery, TX, USA).

Techniques: Expressing, Knockdown, Western Blot

Effect of dictyoceratin-A ( 1 ) on RPAP3-overexpressing DU145 cells. ( a ) The expression levels of RPAP3 protein in stable RPAP3-overexpressing cells were compared with those in wild-type DU145 cells using western blotting. The expression levels of RPAP3 were calculated based on the relative density of bands, normalized to that of β-actin. ( b ) Antiproliferative activity of dictyoceratin-A ( 1 ) in wild-type and RPAP3-overexpressing DU145 cells. The growth inhibition rate was calculated as a percentage relative to that in control cells. Differences were considered significant at * p < 0.01.

Journal: Marine Drugs

Article Title: Target Identification of the Marine Natural Products Dictyoceratin-A and -C as Selective Growth Inhibitors in Cancer Cells Adapted to Hypoxic Environments

doi: 10.3390/md17030163

Figure Lengend Snippet: Effect of dictyoceratin-A ( 1 ) on RPAP3-overexpressing DU145 cells. ( a ) The expression levels of RPAP3 protein in stable RPAP3-overexpressing cells were compared with those in wild-type DU145 cells using western blotting. The expression levels of RPAP3 were calculated based on the relative density of bands, normalized to that of β-actin. ( b ) Antiproliferative activity of dictyoceratin-A ( 1 ) in wild-type and RPAP3-overexpressing DU145 cells. The growth inhibition rate was calculated as a percentage relative to that in control cells. Differences were considered significant at * p < 0.01.

Article Snippet: Polyclonal rabbit anti-RPAP3 antibodies were obtained from Bethyl Laboratories, Inc. (Montgomery, TX, USA).

Techniques: Expressing, Western Blot, Activity Assay, Inhibition, Control