Figure 5

MMS22L and ASF1B trafficking to the nucleus is mediated by IPO4 and is impaired in CDAI patients because of a defective interaction between IPO4 and CDAN1. (A) Quantification of PLA signals between MMS22L and CDAN1 in HeLa cells displayed as the mean number ±SD of dots per nucleus and cytoplasm. Analysis was performed using the ImageJ software. (B) Western blot analysis showing the expression of MMS22L and ASF1B in the nuclear (NE) extracts of shCTRL and shCDAN1‐transduced UT‐7 cells. Protein levels were compared to TBP expression. Data are representative of three independent experiments. (C) Western blot analysis showing the expression of MMS22L, CDAN1, and ASF1B in the nuclear (NE) extracts of shCTRL and shIPO4‐transduced UT‐7 cells. Protein levels were compared to TBP expression. Data are representative of two independent experiments. (D) Upper panel: western blot analysis showing the expression of MMS22L and ASF1B in the nuclear extract (NE) of B‐LCLs established from healthy controls (CTRL 1, 2, 3) and CDAI patients (CDAI 1, 2, 3). Protein levels were compared to TBP expression. Data are representative of two independent experiments. Lower panel: Quantification of relative protein expression level of MMS22L (left panel) and ASF1B (right panel) in CDAI patients. Results are represented as the mean ± SD of the three controls versus the mean ± SD of the three CDAI patients, all normalized to the controls. (E) Quantification of PLA signals between CDAN1 and MMS22L performed in B‐LCLs established from healthy controls (CTRL 1, 2) and CDAI patients (CDAI 1, 2, 3). Results are displayed as the mean percentage ±SD of red dots located in the nucleus and in the cytoplasm. Data from the 2 CTRL B‐LCLs and from the 3 CDAI B‐LCLs were pooled together. Analysis was performed using the ImageJ software. (F) Quantification of PLA signals between CDAN1 and IPO4 performed in B‐LCLs established from healthy controls (CTRL 1, 2) and CDAI patients (CDAI 1, 2, 3). Results are displayed as the mean percentage ±SD of red dots locating in the nucleus and in the cytoplasm. Data from the 2 CTRL B‐LCLs and from the 3 CDAI B‐LCLs were pooled together. Analysis was performed using the ImageJ software. P‐values are determined by a two‐tailed t‐test. ns, not significant, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.