Fig. 4.

CFAP70 regulates the cytoplasmic preassembly of the CSC components CFAP61 and CFAP91. (a) Myc-tagged plasmids of twelve axoneme proteins were constructed by PCR using testis cDNA template and confirmed by sequencing. Flag-tagged CFAP70 was immunoprecipitated with Myc-tagged DNAI1, DNAI2, CFAP61, and CFAP91 but not Myc-tagged ODAD3, DNALI1, SPAG6, RSPH1, RSPH3, RSPH9, DRC2, and DRC4 in HEK293T cells using co-IP assays. The co-IP experiments were repeated for three times and representative blots were shown. (b) Endogenous immunoprecipitation of DNAI2 from testis extracts reveals no defects in DNAI2 association with its heterodimeric partner DNAI1 in Cfap70-KO mice. (c) Endogenous immunoprecipitation of CFAP61 from testis extracts revealed partial disruption in its association with CFAP91 in Cfap70-KO mice compared to WT mice, as revealed by the intensities of the coprecipated bands of CFAP91. The grey values of the protein bands in b and c were measured by ImageJ software (https://imagej.en.softonic.com/). (d) The relative amount of CFAP91 immunoprecipitated by CFAP61 in the testis protein lysates from Cfap70-KO mice and their littermate WT mice. Data were presented as the mean ± SD (n = 3 each group). Statistical significance was determined by two-tailed, unpaired Student's t test; p < 0.0001.