Binding of hPTTG to Ku-70
in intact cells. (A) Expression of hPTTG in differentiated
and undifferentiated HL-60 cells. Equal amounts of total extracts from
HL-60 cells treated with PMA (40 ng/ml), with the vehicle DMSO
or untreated, were separated by SDS–PAGE, transferred to
nitrocellulose filters and incubated with anti-hPTTG. HL-60, extract
from 106 HL-60 cells. (B) Co-immunoprecipitation
of hPTTG with Ku-70 and Ku-80 in undifferentiated HL-60 cells. Anti-hPTTG and
preimmune (PI) sera were used to immunoprecipitate from NP-40 extracts
of HL-60 cells (3 × 107 cells
each). Complexes were resolved by SDS–PAGE, transferred
to nitrocellulose filters and incubated with anti-Ku-70, anti-Ku-80
and anti-hPTTG. HL-60, extract from 106 HL-60 cells.
(C) Study of the relevance of the hPTTG–Ku-70
interaction. Anti-hPTTG was used to immunoprecipitate from extracts
of HL-60 cells (3 × 107)
treated with PMA (40 ng/ml), the vehicle DMSO or untreated.
Nitrocellulose filters were incubated with anti-Ku-70 and anti-hPTTG.
HL-60, extract from 106 HL-60 cells. Asterisks indicate
the IgG heavy and light chains.