Rapid Proteasomal Degradation of Posttranscriptional Regulators of the TIS11/Tristetraprolin Family Is Induced by an Intrinsically Unstructured Region Independently of Ubiquitination

Abstract

The TIS11/tristetraprolin (TTP) CCCH tandem zinc finger proteins are major effectors in the destabilization of mRNAs bearing AU-rich elements (ARE) in their 3= untranslated regions. In this report, we demonstrate that the Drosophila melanogaster dTIS11 protein is short-lived due to its rapid ubiquitin-independent degradation by the proteasome. Our data indicate that this mechanism is tightly associated with the intrinsically unstructured, disordered N-and C-terminal domains of the protein. Furthermore, we show that TTP, the mammalian TIS11/TTP protein prototype, shares the same three-dimensional characteristics and is degraded by the same proteolytic pathway as dTIS11, thereby indicating that this mechanism has been conserved across evolution. Finally, we observed a phosphorylation-dependent inhibition of dTIS11 and TTP degradation by the proteasome in vitro, raising the possibility that such modifications directly affect proteasomal recognition for these proteins. As a group, RNAbinding proteins (RNA-BPs) have been described as enriched in intrinsically disordered regions, thus raising the possibility that the mechanism that we uncovered for TIS11/TTP turnover is widespread among other RNA-BPs.

The control of inflammation via the phosphorylation and dephosphorylation of tristetraprolin: a tale of two phosphatases

“…Unexpectedly, ubiquitination of TTP could not be detected, nor was TTP protected from proteasomal degradation by mutation of all five lysine residues [82]. These observations suggest an atypical mode of degradation of TTP protein.…”

“…These observations suggest an atypical mode of degradation of TTP protein. The only 3D structure of TTP protein solved to date is for the highly conserved central zinc finger region [83], and secondary structure prediction programmes all fail to identify any stable structure in the N-terminal and C-terminal domains [82] (JLED, unpublished data). The unstructured nature of the majority of TTP protein is consistent with proteasomal degradation via a default degradation pathway shared with other largely unstructured proteins [84].…”

“…Since a common feature of 14-3-3 proteins is that they impose secondary structure upon client phosphoproteins [48,49], an obvious hypothesis is that 14-3-3-mediated imposition of stable structure allows TTP protein to escape degradation-by-default. Gueydan and colleagues report that the addition of recombinant 14-3-3 proteins failed to prevent degradation of purified TTP in vitro [82]. It would arguably be more relevant (though less easy) to ask whether 14-3-3 proteins are necessary for the stabilisation of phosphorylated TTP.…”

Tristetraprolin and Its Role in Regulation of Airway Inflammation

“…This complex also serves to protect TTP from dephosphorylation in a protein phosphatase 2A (PP2A)-dependent manner (Sun et al, 2007) and prevent degradation of TTP via the proteasome (Deleault et al, 2008;Pfeiffer and Brooks, 2012). There are, however, some data to suggest that the role of 14-3-3 proteins might be limited to chaperoning TTP and only preventing its dephosphorylation (Marchese et al, 2010;Ngoc et al, 2014). The additional role as a block on proteasomal degradation is challenged, as recombinant 14-3-3 in the presence of TTP did not affect TTP stability and degradation by the proteasome (Ngoc et al, 2014).…”

“…There are, however, some data to suggest that the role of 14-3-3 proteins might be limited to chaperoning TTP and only preventing its dephosphorylation (Marchese et al, 2010;Ngoc et al, 2014). The additional role as a block on proteasomal degradation is challenged, as recombinant 14-3-3 in the presence of TTP did not affect TTP stability and degradation by the proteasome (Ngoc et al, 2014). This is in line with data that showed that TTP levels increased when cells were in the presence of a proteasome inhibitor.…”