An A + U-rich element RNA-binding factor regulates c-myc mRNA stability in vitro.

Abstract

Transient expression of some proto-oncogenes, cytokines, and transcription factors occurs as a cellular response to growth factors, 12-O-tetradecanoylphorbol-13-acetate, antigen stimulation, or inflammation. Expression of these genes is mediated in part by the rapid turnover of their mRNAs. A+U-rich elements in the 3' untranslated regions of these mRNAs serve as one recognition signal targeting the mRNAs for rapid degradation. I report the identification of a cytosolic factor that both binds to the proto-oncogene c-myc A+U-rich element and specffically destabilizes c-myc mRNA in a cell-free mRNA decay system which reconstitutes mRNA decay processes found in cells. Proteinase K treatment of the factor abolishes its c-myc mRNA degradation activity without affecting its RNA-binding capacity. Thus, RNA substrate binding and degradation appear to be separable functions. These findings should aid in understanding how the cell selectively targets mRNAs for rapid turnover.mRNA stability plays an important role in regulating gene expression (9, 10, 34). The rapid turnover of an mRNA ensures that it is maintained at relatively low steady-state levels and that changes in the turnover rate can affect its steady-state level over a short period of time. This type of regulation allows transient alterations in the expression of some cytokines, transcription factors, and the c-myc and c-fos proto-oncogenes in response to growth factors (7, 37, 46), 12-O-tetradecanoylphorbol-13-acetate (23, 37), antigen stimulation (24), or inflammation (45). A common feature of the mRNAs encoded by all of these genes is an A+U-rich element (ARE) in the 3' untranslated region (3'UTR) (14). These AREs serve as one signal targeting the mRNAs for rapid turnover (18,39,46), although sequences in the c-myc 5'UTR (18,32,33) and the c-fos coding region (19,40) can, in some instances, influence mRNA turnover.Although much is known about the cis-acting elements controlling mRNA stability, the trans-acting factors remain largely undefined. The development of cell-free mRNA decay systems makes possible the identification, purification, and characterization of these trans-acting factors (5,31,35,41). Ross and colleagues developed a system that is based on earlier in vitro translation systems and includes polysomes from the erythroleukemia cell line K562, magnesium, potassium, ATP, GTP, an ATP-regenerating system, and a 130,000 x g postribosomal supernatant (S130) (13,35 (nt) from the 3' end, while the second product lacked 12 nt from the 3' end. In subsequent steps, the mRNA was degraded in a 3'-to-5' direction. The same degradation pathway appeared in cells during degradation of H4 histone mRNA (36). In addition, Brewer and Ross (11,12) found that c-myc mRNA was degraded in vitro by a sequential pathway, in which the A+U-rich sequences of the 3'UTR were cleaved only after most or all of the poly(A) was removed. The remainder of the mRNA was degraded in subsequent steps. This pathway has been observed in vivo for c-myc mRNA (42). Thus, the in vitro mRNA de...

Identification of an RNA element that confers post-transcriptional repression of connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene: Similarities to retroviral RNA-protein interactions

“…As shown in Figure 2a, a few short sequences that are signi®cantly homologous to reported cis-repressive elements could be found in the 200 bp area (Brewer, 1991;Sterling and Bresnick, 1996;Ogbourne and Antalis, 1998). Assuming that these putative elements might be involved in the observed repressive e ect, two deletion mutants of SA3 were prepared.…”

ABINs: A20 binding inhibitors of NF-κB and apoptosis signaling

“…In contrast with ABIN-1 and A20, ABIN-2 mRNA expression is not modulated upon stimulation of cells with TNF, LPS or interferon- [29], and is therefore believed to be NF-B independent However, ABIN-2 mRNA expression is augmented in adult mouse liver after partial hepatectomy, and like A20, in progesterone-stimulated progesterone receptor B expressing cells [32,33]. Furthermore, the 3'-untranslated region of ABIN-2 mRNA contains two binding sequences for 'AU-rich element binding factor 1' (AUF1), which has been suggested to regulate mRNA turnover [34]. In the case of rat uterus ABIN-2 mRNA, these sequences were indeed shown to bind AUF1, and ABIN-2 mRNA was 9 upregulated with the same kinetics as cytoplasmic AUF1 protein isoforms in ovariectomized rat uteri stimulated with 17-estradiol [35].…”

Post‐transcriptional regulation of gene expression by degradation of messenger RNAs

“…More recent studies have identified other factors capable of binding AREs with various degrees of avidity. These include selected nuclear ribonucleic proteins (hnRNP A1, hnRNP C (Hamilton et al, 1993), hnRNP D, also known as AUF-1 (Brewer, 1991)), RNA-binding proteins displaying enzymatic activities (GAPDH (Nagy and Rigby, 1995), AUH (Nakagawa et al, 1995)), other HuR-like proteins, belonging to the ELAV family (embryonic lethal abnormal vision) and related to the Drosophila neuron-specific RNA-binding proteins (Hel-N1, HuC, HuD, (Levine et al, 1993;Good, 1995;Ma et al, 1996) and HuR (Fan and Steitz, 1998;Peng et al, 1998), heat shock protein 70 (Hsp70) and others (AUBP (Malter and Hong, 1991), AU-A, AU-B, AU-C (Bohjanen et al, 1991(Bohjanen et al, , 1992, tristetraprolin (Taylor et al, 1996b), butyrate response factor-1 (BRF1), encoding a zinc finger protein homologous to tristetraprolin (Stoecklin et al, 2002), KSRP ), TIA-1 and TIAR (Piecyk et al, 2000)). …”

“…Normally, the p42 and p45 isoforms appear exclusively in the nucleus (Zhang et al, 1993;Arao et al, 2000). P37 AUF1 and/or p40 AUF1 are associated with the acceleration of mRNA turnover by ARE: a protein complex containing cytoplasmic p37 AUF1 and p40 AUF1 is able to destabilize polysomal c-myc mRNA in vitro (Brewer, 1991). The affinity of recombinant p37 AUF1 for the ARE motif is closely correlated with its potential to destabilize the mRNA .…”