Figure 3.

Functional capacity of α-repeat arrays in recruiting the centromere proteins CENPA and CENPB can be assessed by PCR-based assays of centromeric DNA. ChIP was performed on LNCaP prostate cancer cells using CENPA and CENPB antibodies or control mouse IgG antibody. The arrays to which centromere proteins bind were measured by qPCR in immunoprecipitated chromatin and compared to the input. (A) Occupancy of CENPA on specific centromere arrays. At least one array in the centromere of each human chromosome recruited CENPA. CENPA antibody precipitated between 0.1 and 0.4% of every centromere array other than the array DYZ3 in Chromosome Y (0.7%). Asterisks indicate dominant arrays in each chromosome that recruit CENPA; these arrays also represent the largest centromere core arrays in every chromosome. We do not yet have an assay to discriminate between the array D14Z1/D22Z1, which is present in both Chr 14 and 22. A PCR assay that measures both arrays shows that D14Z1/D22Z1 indeed dominates the recruitment of CENPA to centromeres 14 and 22. We do not have an assay for the larger array D19Z3 in Chr 19, which resembles D1Z7. Given that the latter recruits CENPA, it is likely that D19Z3 similarly recruits CENPA to centromere 19. (B) Occupancy of CENPB on centromeric arrays. At least one array in the centromere of each human chromosome (other than Y) recruited CENPB. CENPB did not bind the array DYZ3 in Y. There is a significant difference between the amounts of D7Z2 immunoprecipitated with CENPB antibody as compared to the IgG control (P < 0.001). The existence of only one CENPB box in a sequence of 16 alphoid repeats (Waye et al. 1987a) might explain the relatively low binding of CENPB to this array. Asterisks indicate arrays that contain CENPB box sequences.