Figure 6. CFAP52 interacts with SPATA6 and regulates its expression.
(A) Expression of nine ASS-associated proteins was analysed by western blotting in testis lysates from WT mice and Cfap52-KO mice. β-actin served as a loading control. Western blotting experiments were repeated three times with similar results. ASS, acephalic spermatozoa syndrome. Bar graph representing band intensities of SPATA6 blots, and data represent the mean ± SEM of three biological replicates. Student’s t test, **p<0.01. (B) Coimmunofluorescence staining of SPATA6 (green) and ac-Tub (red) in spermatozoa from the CFAP52-mutant patient and a normal control. Scale bars, 5 μm. The staining experiments were repeated three times with similar results. (C) A coimmunoprecipitation assay showed that FLAG-tagged CFAP52 could be immunoprecipitated with Myc-tagged SPATA6 in HEK293T cell extracts. (D) An interaction between endogenous SPATA6 and CFAP52 was identified in mouse testis lysates. Rabbit IgG served as the negative control. Co-IP experiments were repeated three times with similar results. (E, F) The degradation of FLAG-tagged SPATA6 in HEK293T cells with or without Myc-tagged CFAP52. Protein samples were harvested at the indicated times after treatment with 100 μg/ml cycloheximide (CHX) to block new protein synthesis. β-actin served as a loading control. Western blotting experiments were repeated three times with similar results.
Figure 6—figure supplement 1. SUN5 staining in spermatozoa from the CFAP52-mutant patient and a healthy control.
