Figure 1
Ribosome biogenesis defects in RPS19+/p.Q126X iPSCs. (A) Western blot showing RPS19 protein in RPS19+/p.Q126X iPSCs and a WT (WT1) control line. Lane 2 represents the mutant iPSC line examined throughout this manuscript. Lane 3 represents an iPSC line that did not fulfill all of the pluripotency criteria and therefore was not examined further. Ratio of RPS19:actin determined by densitometry is shown relative to the value for the WT1 sample, which was assigned an arbitrary value of 1.0. (B) Sucrose gradient polysome profiles showing reduced 40S:60S ratio in RPS19+/p.Q126X iPSCs compared with a WT control line. Arrows show direction of the sucrose gradient from less to more dense. (C) Histogram summarizing 3 independent polysome profiling experiments examining RPS19+/p.Q126X iPSCs and 2 different WT lines. The 40S:60S ratio was significantly reduced in the mutated clone (0.55 vs 0.66, *P < .05). (D) Diagram showing rRNA maturation and Northern blot probes complementary to ITS1 (for the 40S unit) and ITS2 (for the 60S unit), used in Figures 1E, 4G, and 6D. The probe sequences are shown in supplemental Table 3. (E) Northern blot analysis of iPSCs using the ITS1 probe. RPS19+/p.Q126X iPSCs exhibit relative accumulation of 21S pre-rRNA compared with WT iPSCs. The 21S:18SE pre-rRNA ratios determined by densitometry scanning is shown at the bottom of the panels. ITS, internal transcribed spacer.

Ribosome biogenesis defects in RPS19+/p.Q126X iPSCs. (A) Western blot showing RPS19 protein in RPS19+/p.Q126X iPSCs and a WT (WT1) control line. Lane 2 represents the mutant iPSC line examined throughout this manuscript. Lane 3 represents an iPSC line that did not fulfill all of the pluripotency criteria and therefore was not examined further. Ratio of RPS19:actin determined by densitometry is shown relative to the value for the WT1 sample, which was assigned an arbitrary value of 1.0. (B) Sucrose gradient polysome profiles showing reduced 40S:60S ratio in RPS19+/p.Q126X iPSCs compared with a WT control line. Arrows show direction of the sucrose gradient from less to more dense. (C) Histogram summarizing 3 independent polysome profiling experiments examining RPS19+/p.Q126X iPSCs and 2 different WT lines. The 40S:60S ratio was significantly reduced in the mutated clone (0.55 vs 0.66, *P < .05). (D) Diagram showing rRNA maturation and Northern blot probes complementary to ITS1 (for the 40S unit) and ITS2 (for the 60S unit), used in Figures 1E, 4G, and 6D. The probe sequences are shown in supplemental Table 3. (E) Northern blot analysis of iPSCs using the ITS1 probe. RPS19+/p.Q126X iPSCs exhibit relative accumulation of 21S pre-rRNA compared with WT iPSCs. The 21S:18SE pre-rRNA ratios determined by densitometry scanning is shown at the bottom of the panels. ITS, internal transcribed spacer.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal