Fig. 4: Bmc1 is recruited to the active telomerase holoenzyme by Pof8.

A Northern blot and semi-quantitative RT-PCR of TER1 and U6 in PrA immunoprecipitates for an untagged wild type (wt), PrA-tagged (Bmc1 PrA), and PrA-tagged Pof8 knockout strain (Bmc1 PrA ∆Pof8). Bmc1 PrA was detected in input and immunoprecipitated samples by western blots probing for PrA (bottom panel). Possible cleavage products are indicated with an asterisk. B qRT-PCR of TER1 and U6 in Bmc1 PrA immunoprecipitates from wild-type and pof8∆ strains normalized to input RNA. Relative TER1 and U6 IP was calculated by comparing percent immunoprecipitation of TER1 or U6 to immunoprecipitation from an untagged strain (mean ± standard error, two-tailed unpaired t test *P  <  0.05, **P  <  0.01, ***P  <  0.001, and ****P < 0.0001) (n = 3 biological replicates). C Northern blot of mature and intron-containing U6 in PrA- and myc-tagged immunoprecipitated RNA. D Telomerase assay of PrA-tagged Bmc1 in a wild-type and pof8∆ strain. A ³²P-labeled 100-mer oligonucleotide was used as a loading control. Telomerase extension products were compared to a terminal transferase ladder, with +1 and +4 extension products indicated. Western blot probing for PrA following PrA immunoprecipitation is shown in the panel below. E Proposed model of the fission yeast telomerase holoenzyme. TER1 structure and binding locations are based on models constructed by Hu et al.¹⁶ and Mennie et al.¹⁴. Source data are provided as a Source Data file.