Interactions of CCCH zinc finger proteins with mRNA - Binding of tristetraprolin-related zinc finger proteins to Au-rich elements and destabilizationof mRNA

Macrophages derived from tristetraprolin (TTP)-deficient mice exhibited inc reased tumor necrosis factor alpha (TNF alpha) release as a consequence of increased stability of TNF alpha mRNA, TTP was then shown to destabilize TN F alpha mRNA after binding directly to the AU-rich region (ARE) of the 3'-u ntranslated region of the TNF alpha mRNA In mammals and in Xenopus, TTP is the prototype of a small family of three known zinc finger proteins contain ing two CCCH zinc fingers spaced 18 amino acids apart; a fourth more distan tly related family member has been identified in Xenopus and fish. We show here that representatives of all four family members were able to bind to t he TNF alpha ARE in a cell-free system and, in most cases, promote the brea kdown of TNF alpha mRNA in intact cells. Because the primary sequences of t hese CCCH proteins are most closely related in their tandem zinc finger dom ains, we tested whether various fragments of TTP that contained both zinc f ingers resembled the intact protein in these assays. We found that amino- a nd carboxyl-terminal truncated forms of TTP, as well as a 77 amino acid fra gment that contained both zinc fingers, could bind to the TNF alpha ARE in cell-free cross-linking and gel shift assays. In addition, these truncated forms of TTP could also stimulate the apparent deadenylation and/or breakdo wn of TNF alpha mRNA in intact cells. Alignments of the tandem zinc finger domains from all four groups of homologous proteins have identified invaria nt residues as well as group-specific signature amino acids that presumably contribute to ARE binding and protein-specific activities, respectively.