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. 2020 Apr 13;23(5):101058. doi: 10.1016/j.isci.2020.101058

Figure 6.

Figure 6

Nwd1 and Paics Are Localized in Purinosomes

(A–L) HeLa cells were transfected with FLAG-Nwd1 (red) together with Fgams-EGFP (A–F) or Paics-EGFP (G–L) and cultured in complete medium (A–C and G–I) or in purine-depleted medium (D–F, J–L). Nwd1 was co-clustered with Fgams or Paics as purinosomes in purine-depleted medium (arrows). Nuclei were stained with Hoechst dye (blue). Insets in (D–F) and (J–L) showed the magnified view of the individual purinosomes formed in cytoplasm. (M) Co-IP showing the interaction between Nwd1 and Fgams. HEK293 cells expressing FLAG-Nwd1 and/or Fgams-EGFP were subjected to immunoprecipitation with an anti-FLAG antibody, followed by immunoblotting with anti-EGFP or anti-FLAG antibodies.

(N–P) HeLa cells co-transfected with EGFP-Nwd1 (N) and Hsp90-HA (O) were cultured under purine-depleted conditions. (P) Merged view. The arrows indicate the colocalization of Nwd1 and Hsp90 in purinosomes. Insets showed the high power photomicrograph of the individual purinosomes.

(Q) Interaction between Nwd1 and Hsp90. HEK293 cells expressing Hsp90-HA and/or FLAG-Nwd1 were subjected to immunoprecipitation using an anti-FLAG antibody, followed by immunoblotting with an anti-HA or anti-FLAG antibody. Scale bars, 15 μm.