Chorioallantoic Fusion Defects and Embryonic Lethality Resulting from Disruption of Zfp36L1, a Gene Encoding a CCCH Tandem Zinc Finger Protein of the Tristetraprolin Family

Abstract

The mouse gene Zfp36L1 encodes zinc finger protein 36-like 1 (Zfp36L1), a member of the tristetraprolin (TTP) family of tandem CCCH finger proteins. TTP can bind to AU-rich elements within the 3-untranslated regions of the mRNAs encoding tumor necrosis factor (TNF) and granulocyte-macrophage colony-stimulating factor (GM-CSF), leading to accelerated mRNA degradation. TTP knockout mice exhibit an inflammatory phenotype that is largely due to increased TNF secretion. Zfp36L1 has activities similar to those of TTP in cellular RNA destabilization assays and in cell-free RNA binding and deadenylation assays, suggesting that it may play roles similar to those of TTP in mammalian physiology. To address this question we disrupted Zfp36L1 in mice. All knockout embryos died in utero, most by approximately embryonic day 11 (E11). Failure of chorioallantoic fusion occurred in about two-thirds of cases. Even when fusion occurred, by E10.5 the affected placentas exhibited decreased cell division and relative atrophy of the trophoblast layers. Although knockout embryos exhibited neural tube abnormalities and increased apoptosis within the neural tube and also generalized runting, these and other findings may have been due to deficient placental function. Embryonic expression of Zfp36L1 at E8.0 was greatest in the allantois, consistent with a potential role in chorioallantoic fusion. Fibroblasts derived from knockout embryos had apparently normal levels of fully polyadenylated compared to deadenylated GM-CSF mRNA and normal rates of turnover of this mRNA species, both sensitive markers of TTP deficiency in cells. We postulate that lack of Zfp36L1 expression during mid-gestation results in the abnormal stabilization of one or more mRNAs whose encoded proteins lead directly or indirectly to abnormal placentation and fetal death.The tristetraprolin (TTP) family of CCCH tandem zinc finger proteins consists of three known members in mammals and a fourth more distantly related protein in frogs and fish (reviewed in reference 2). The best-studied family member is TTP, which is the product of the immediate-early response gene Zfp36 in the mouse (ZFP36 in humans). TTP appears to act in normal physiology to bind to AU-rich elements (ARE) in certain mRNAs and to destabilize those transcripts. Specifically, the mRNAs encoding tumor necrosis factor (TNF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) are stabilized in TTP knockout (KO) mice and in cells derived from them. This results in excessive secretion of these cytokines from the appropriate cell types, resulting in a severe systemic inflammatory phenotype with myeloid hyperplasia in the TTP KO mice (37).The two other known mammalian TTP family members contain CCCH tandem zinc finger (TZF) domains that are very similar to that of TTP; these TZF domains represent the ARE-binding domains of the proteins. One of these proteins (Zfp36L1, for Zfp36-like 1) was actually the first member of this family in which the TZF domain was recognized (15). Its cDNA was originally cloned ...

Phosphorylation site analysis of the anti-inflammatory and mRNA-destabilizing protein tristetraprolin

“…A third related protein (ZFP36L3) was recently identified that seems to be expressed only in rodents as a placentaspecific protein [31]. Recent studies have shown that mice deficient in ZFP36L1 develop chorioallantoic fusion defects and embryonic lethality [73], and mice with decreased levels of an amino-terminal truncated form of ZFP36L2 exhibit female infertility and disrupted early embryonic development [74]. Some of the phosphorylation sites reported here are located in relatively conserved sequence blocks among all three proteins of the TTP family, including T 106 , Y 158 , S 184 , S 186 , S 217 , S 218 , S 273 , S 276 , Y 284 , S 294 and S 296 in hTTP (Figure 4).…”

A Cassette System to Study Embryonic Stem Cell Differentiation by Inducible RNA Interference

“…We reported recently that the mRNA decay promoting activity of Brf1 is negatively regulated by phosphorylation via protein kinase B, which promotes complex formation to 14-3-3 [39]. Target mRNAs of Brf1 are not known but play a role in development, as mice lacking both alleles die at day 11 [40]. That Brf1 is controlled at least in part by Stat3 was suggested by the LIF removal experiment where Brf1 levels dropped (Fig.…”

“…If, as may well be the case, Brf1 also regulates cardiomyocyte formation in human ES cells, the protocol described here may prove useful in regenerative cardiology for replacement therapy. In addition, transgenic mice eventually generated from the lines described here might circumvent the lethality of Brf1 knockouts [40] and allow the downregulation of Brf1 in adult tissue to reveal further functional aspects of this post-transcriptional regulator.…”

A novel concept in antiangiogenic and antitumoral therapy: multitarget destabilization of short-lived mRNAs by the zinc finger protein ZFP36L1

“…All four proteins bind and destabilize ARE-containing mRNAs in vitro (Baou et al, 2009). However, gene knockout studies have provided evidence for their unique role in vivo (Carballo et al, 1998;Stumpo et al, 2004Stumpo et al, , 2009). ZFP36L1 knockout mice are embryonic lethal because of abnormal placentation and major vascular defects.…”