Modulation of M2-PK activity by HPV-16 E7.
(A) Overexpression of M2-PK in 14/2 cells. Expression of
E7 was induced in 14/2 cells by addition of dexamethasone for 4 hr.
In a control experiment, asynchronously growing NRK cells were either
mock-treated or treated with dexamethasone for 4 hr. Extracts were
prepared from these cells as indicated, and the abundance of M2-PK in
the cellular extracts was analyzed by direct immunoblotting. (B
Upper) Gel filtration analysis. Extracts from E7-expressing or
control 14/2 cells were subjected to gel filtration, followed by
determination of M2-PK activity in individual fractions, by using a PEP
concentration of 2 mM. The peaks of main activities also are indicated
for GAPDH, enolase (EN), and lactate dehydrogenase (LDH), assayed in
parallel. (Lower) The abundance of M2-PK in each
fraction was determined by direct immunoblotting. (C and
D) Isoelectric focusing. Extracts from E7-expressing or
control 14/2 cells were subjected to isoelectric focusing, and M2-PK
activity was determined in the presence of 2 mM PEP (C) or
0.2 mM PEP (D) as indicated. The peak of main activity also
is indicated for GAPDH, assayed in parallel. pI values of selected
fractions are given for reference.