N-terminal 281 residues in human cyclin T support
Tat transactivation. Reciprocal substitutions of the cysteine and
tyrosine at position 261 inactivate human cyclin T and activate mouse
cyclin T, respectively, in CHO cells. The HIV-1 long terminal repeat
was expressed alone [pHIVSCAT (lane 1)] or together with Tat
[pcDNA3Tat (lane 2)]. To Tat were added human cyclin T [hT726 (lane
3)], truncated human cyclin T containing the substitution of lysine to
asparagine at position 277 [hT281K277N (lane 4)], truncated human
cyclin T containing the substitution of the alanine to proline at
position 269 [hT372A269P (lane 5)], human cyclin T containing the
substitution of the lysine to methionine at position 265 [hT726K265M
(lane 6)], truncated human cyclin T containing the substitution of the
cysteine to tyrosine at position 261 [hT281C261Y (lane 7)], truncated
human cyclin T to position 227 [hT227 (lane 8)], and mouse cyclin T
containing the substitution of the tyrosine at position 261 to cysteine
[mT724Y261C (lane 9)]. Solid and striped bars denote wild-type or
truncated and mutant cyclin T proteins, respectively. The value of the
CAT activity of pHIVSCAT alone was set to 1. Standard errors of the
mean from three independent transfections are presented. Western
blotting revealed that levels of expression of the full length and
truncated human cyclin T proteins (hT281K277N and hT281C261Y) were
similar. Numbers above the lanes correspond to lanes from coexpression
assays. NS, nonspecific bands.