Tristetraprolin (
TTP or ZFP36) is a tandem CCCH zinc-finger
RNA-binding protein that regulates the stability of certain AU-rich element (ARE) mRNAs. Recent work suggests that
TTP is deficient in
cancer cells when compared with normal cell types. In this study we found that
TTP expression was lower in invasive
breast cancer cells (MDAMB231) compared with normal breast cell lines MCF12A and MCF-10.
TTP targets were probed using a novel approach by expressing the C124R zinc-finger
TTP mutant that functions as dominant negative and increases target
mRNA expression. In contrast to wild-type
TTP, C124R
TTP was able to increase certain ARE-
mRNA expressions in serum-stimulated
breast cancer cells. Using an ARE-gene microarray, novel targets of
TTP regulation were identified, namely,
urokinase plasminogen activator (uPA), uPA receptor and
matrix metalloproteinase-1, all known to have prominent roles in
breast cancer invasion and
metastasis. Expression of these targets was upregulated in tumorigenic types, particularly in highly invasive MDAMB231. The
mRNA half-lives of these
TTP-regulated genes were increased in
TTP-knockout embryonic mouse fibroblasts, as assessed using real-time polymerase chain reaction, whereas forced restoration of
TTP by transfection led to a reduction in their
mRNA levels.
RNA immunoprecipitation confirmed an association of
TTP, but not C124R, with these target transcripts. Moreover,
TTP reduced, whereas the mutant C124R
TTP increased, the activity of reporter constructs fused to target ARE. As a result of
TTP regulation, invasiveness of MDAMB231 cells was reduced. The data suggest that
TTP, in a
3' untranslated region-and ARE-dependent manner, regulates an important subset of
cancer-related genes that are involved in cellular growth, invasion and
metastasis.