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

Figure 5.

Figure 5

SLX4IP Interacts with BLM Helicase

(A) U2OS cells transfected with the indicated siRNAs were fixed and processed for BLM immunofluorescence followed by telomeric PNA (TelG) FISH. Scale bar represents 10 μm. Dashed lines indicate nucleus outlines (as determined using DAPI staining; not shown). Insets represent 3× magnifications of the indicated fields.

(B) Quantification of (A). At least 100 cells per condition were counted. Data are presented as 5th–95th percentiles; n = 3; ∗∗∗∗p < 0.00001, one-way ANOVA; ns, not significant.

(C) U2OS whole-cell extracts were separated using SDS-PAGE and analyzed using BLM immunoblotting. Tubulin was used as loading control. Numbers on the right denote molecular weight (kDa). Numbers below indicate protein levels. Protein levels were normalized to SLX4IP+/+, which was arbitrarily assigned a value of 1.

(D) Whole-cell extracts from HEK293 cells transiently expressing GFP constructs were subjected to GFP-trap co-immunoprecipitation (IP). Input and IP samples were separated using SDS-PAGE and analyzed using GFP, SLX4IP, and RMI2 immunoblotting. Numbers denote molecular weight (kDa).

(E) Whole-cell extracts from HEK293 cells transiently expressing GFP constructs were subjected to GFP-trap co-immunoprecipitation (IP). Input and IP samples were separated using SDS-PAGE and analyzed using GFP and BLM immunoblotting. Numbers denote molecular weight (kDa).

(F) Whole-cell extracts from HEK293 cells transfected with the indicated siRNAs and transiently expressing GFP constructs were subjected to GFP-trap co-immunoprecipitation (IP). Input and IP samples were separated using SDS-PAGE and analyzed using GFP, SLX4, BLM, and XPF immunoblotting. Tubulin was used as loading control. Numbers denote molecular weight (kDa).

(G) Recombinant Flag-BLM and SLX4IP proteins were subjected to BLM co-immunoprecipitation (IP). Normal IgGs were used as negative IP control. Input and IP samples were separated using SDS-PAGE and analyzed using BLM and SLX4IP immunoblotting. Numbers denote molecular weight (kDa).

See also Figure S7.