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. 2019 Oct 3;76(1):27–43.e11. doi: 10.1016/j.molcel.2019.07.010

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© 2019 The Author(s)

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

PMC Copyright notice

Loss of SLX4IP and SLX4 Causes a Synthetic Growth Defect

(A) SLX4IP^−/− U2OS cells transfected with siSLX4 were fixed and processed for PML immunofluorescence followed by telomeric PNA (TelG) FISH. DNA was counterstained with DAPI. Scale bar represents 10 μm. Insets represent 3× magnifications of the indicated fields.

(B) Quantification of (A). At least 30 mitotic cells per condition were counted. Data are represented as mean ± SD; n = 3; ^∗p < 0.01 and ^∗∗p < 0.001, one-way ANOVA; ns, not significant.

(C) U2OS cells transfected with the indicated siRNAs were fixed and processed for PICH immunofluorescence followed by telomeric PNA (TelG) FISH. DNA was counterstained with DAPI. Scale bar represents 10 μm. Insets represent 3× magnifications of the indicated fields.

(D) Quantification of (C). At least 30 mitotic cells per condition were counted. Data are represented as mean ± SD; n = 3; ^∗∗∗∗p < 0.00001, one-way ANOVA; ns, not significant.

(E) Quantification of (C). At least 30 mitotic cells per condition were counted. Data are represented as mean ± SD (n = 3).

(F) U2OS cells transfected with the indicated siRNAs were fixed and processed for Na^+/ K⁺ ATPase α1 immunofluorescence followed by telomeric PNA (TelG) FISH. DNA was counterstained with DAPI. Scale bar represents 10 μm. Insets represent 3× magnifications of the indicated fields.

(G) Quantification of (F). At least 100 cells per condition were counted. Data are presented as mean ± SD; n = 3; ^∗∗∗p < 0.0001, one-way ANOVA; ns, not significant.

(H) U2OS cells transfected with the indicated siRNAs were fixed, stained with propidium iodide, and analyzed using FACS. At least 10,000 cells per condition were counted.

(I) Quantitation of (H). Data are presented as 5th–95th percentiles; n = 5; ^∗∗∗∗p < 0.00001, one-way ANOVA; ns, not significant.

(J) U2OS cells were transfected with the indicated siRNAs. After 72 h of knockdown, cells were re-seeded and were then permitted to grow for 11 days before fixation and staining.

(K) Quantitation of (I). The surviving fraction was normalized to SLX4IP^+/+ siCTRL, which was arbitrarily assigned a value of 1. Data are represented as mean ± SD; n = 5; ^∗∗∗∗p < 0.00001, Student’s t test; ns, not significant.

See also Figure S6.