Cloning and characterization of chicken YB-1: regulation of expression in the liver.

Abstract

A cDNA expression library constructed from day 9 embryonic liver was screened with a previously identified protein binding site in the flanking region of the liver-specific, estrogen-dependent avian apoVLDLII gene. Two of the clones isolated were shown to encode the chicken homolog of the Y-box binding protein, YB-1 (dbpb), which we have designated chkYB-1. This protein was originally identified in avian extracts by virtue of its ability to bind to two reverse CCAAT motifs in the Rous sarcoma virus enhancer. Since its identification, additional nucleic acid binding properties have been ascribed to its homologs, or closely related proteins, in other species. We have determined the sequence of chkYB-1, investigated its ability to bind to sites known to be involved in tissue-specific expression in the liver, and examined factors influencing its hepatic expression. These studies have demonstrated that the level of chkYB-1 mRNA in the liver decreases steadily throughout embryogenesis and for several weeks posthatching until adult levels are attained. We present several lines of evidence that YB-1 expression in the liver is positively associated with DNA synthesis or cell proliferation. Its binding characteristics indicate that the protein can interact specifically with a number of binding sites for liver!-enriched or specific factors. In addition, although it is not particularly asymmetric in terms of base composition, we find a marked preference in binding to the pyrimidine-rich strand of these sites regardless of the presence or polarity of an intact CCAAT box. The increased levels of expression of YB-1 during proliferation combined with its binding characteristics suggest that it may be involved in the reduced expression of liver-specific genes observed at early stages of development or during liver regeneration.A large number of elements involved in the regulation of eukaryotic genes contain a variation of the motif CCAAT as a subsequence (33). Sites containing such motifs have been shown to be recognized by a number of different trans-acting factors, some of which display restricted tissue distribution and others of which are ubiquitous (2, 44). A requirement for sequences in addition to the CCAAT motif has been demonstrated for some factors, but others display an ability to bind an array of sites with apparently little structural similarity other than the motif itself (9,18,19,29,39,61). A particularly striking example of the latter group is provided by the avian factor EFla, which was initially identified in crude nuclear protein extracts by virtue of its ability to bind to a site in the Rous sarcoma virus (RSV) long terminal repeat (LTR) (21,29,51,62). This protein has been purified and shown to bind to sites in the RSV LTR either as a homodimer or as a heterodimer formed with a second protein designated EFlb. Subsequently, EFla was shown to be capable of interacting with a number of binding sites for trans-acting factors that contain the sequence CCAAT in the noncoding DNA strand.DNA binding proteins that ha...

Targeting the Y/CCAAT box in cancer: YB-1 (YBX1) or NF-Y?

“…35 A similar strategy was reported in the cloning of rat EFIa, using a CCAAT oligo from the RSV LTR, 62 Xenopus FRG-1 63 and chicken YB-1. 64 This technique identifies phages producing a single polypeptide, and it could not have been used for NF-Y, as all three of its subunits are required for DNA binding. YB-1 was shown to be a protein with a known nucleic acid-binding domain, termed CSD (cold-shock domain), which is highly conserved in eukaryotes and prokaryotes ( Figure 1 and Mihailovich et al 65 ).…”

“…The chicken YB-1, subsequently cloned, also showed DNA binding with little dependence on the presence of a Y/CCAAT box. 64 (ii) The Xenopus FRG-1 gene was cloned with expression libraries 67 probed with antibodies directed against p54/p56, 68 one of the subunits of the mRNAbinding complex biochemically identified in the 1970s, and widely studied for its role in mRNA translation. 68 Thereafter, research on YB-1 proceeded, by and large, in two parallel fields: its role in the control of mRNA biology (splicing, stability, translation), in which it has taken a center-stage position (Bouvet and Wolffe 69 ; reviewed in Evdokimova et al 70 ), and its role in the control of transcriptional initiation, which has been more controversial.…”

The Y-box Binding Protein YB-1 Suppresses Collagen α1(I) Gene Transcription via an Evolutionarily Conserved Regulatory Element in the Proximal Promoter

“…COL1A1 Gene Promoter-Sequence analysis of the region Ϫ220 to ϩ115 of the mouse COL1A1 gene (19,22,23) revealed three putative YB-1 binding sites located between positions Ϫ83 and Ϫ72 (Site I), Ϫ103 and Ϫ92 (Site II), and Ϫ129 and Ϫ118 (Site III) (Fig. 1B).…”

“…Sizing of the CYE Binding Activities-Western blot analysis showed that YB-1, a protein with a calculated molecular size of 35 kDa and apparent size of 52 kDa in SDS-polyacrylamide gel (23), is expressed in nuclear extracts from quiescent and proliferating NRK cells (Fig. 4A).…”

YB-1 DNA-binding protein represses interferon gamma activation of class II major histocompatibility complex genes.

“…In addition to the CCAAT element, YB-1 also binds to several unrelated sequences as well as single-strand DNA and mRNA (47)(48)(49)(50)(51)(52)(53). There are many precedents for the ability of a DNA-binding protein to recognize unrelated sequences (54)(55)(56).…”