The fibronectin (FN) gene has become paradigmatic to illustrate genome evolution by exon shuffling, generation of protein diversity by alternative mRNA splicing, and topological coordination between transcription and splicing. Alternative splicing in three sites of the primary transcript gives rise to multiple FN polypeptides. This process is cell type-, development- and age-regulated. The different FN variants seem to play specific roles in FN dimer secretion, blood clotting, adhesion to lymphoid cells, skin wound healing, atherosclerosis, and liver fibrosis. This review focuses on function assignment to the alternatively spliced segments, as well as on the external signals and cis-acting sequences that control the mechanisms of alternative splicing. We also discuss FN transcriptional regulation in response to viral transformation, growth factors, and cyclic AMP in the light of promoter architecture and its interaction with specific transcription factors. The relevance of FN RNA "tracks" as assembly lines of coordinated transcription and RNA processing is also addressed.

We have previously proposed a molecular interaction between the liver factors that bind to the cyclic AMP response element (CRE) and CCAAT sites of the fibronectin (FN) gene based on the following evidence: (i) the close spacing of 20 base pairs between CRE and CCAAT elements is conserved in the FN genes from rats, mice, and humans; (ii) footprinting competitions showed that CRE oligonucleotides are able to detach both liver factors; (iii) CCAAT binding and transcriptional activity of liver extracts are reduced when the distance between the CRE and CCAAT elements is increased; and (iv) CCAAT-binding is stimulated by the addition of a liver extract fraction containing the CREbinding factor ATF-2. This report provides binding and immunochemical evidence that nuclear factor I (CTF/ NF-I) and CP1 (NF-Y or CBF) are the only liver factors that bind to the ؊150 CCAAT element of the FN gene, forming distinct complexes. We show that these factors bind less efficiently to the CCAAT site of a FN promoter in which the ؊170 CRE has been disrupted by site-directed mutagenesis and that each element contributes positively to the liver transcriptional activity assessed in vitro with a G-less cassette construct and in vivo by transfection of hepatoma cells with CAT constructs. Furthermore, using a method that combines UV crosslinking and immunoprecipitation, we show that antibodies specific to ATF-2 are able to specifically precipitate protein-protein-DNA complexes containing NF-I and CP1. This simple method preserves weak macromolecular interactions, avoiding the disruptive electrophoresis conditions of gel mobility shifts assays.Advances in our knowledge of the physiology of gene regulation will ultimately require the precise definition of the multimolecular processes that are built up on a given gene's DNA by both the well known and still unknown factors. Ideally, the model gene should be widely expressed and regulated at the levels of transcription and splicing. Besides fitting these requirements (1), the fibronectin (FN) 1 gene offers a suitable model because it has an essential role in development (2), and changes in its expression pattern are involved in pathology, particularly of the liver (3). The importance of studying FN regulation in liver is highlighted by the fact that although many tissues express FN, liver is the major source of the plasma form of FN (4), made of splicing variants different from those of the extracellular matrix forms. Moreover, liver fibrosis, a common pathological condition preceding cirrhosis, is accompanied by dramatic changes in FN expression (3), whose mechanisms are still poorly understood.The Ϫ170 cyclic AMP response element (CRE; 5Ј-TGACGTCA-3Ј) of the FN gene appears to be involved in different regulations of the promoter. Besides mediating cAMP stimulation (5-7), this element was unexpectedly shown to be essential for responsiveness to serum (8) in an additive way and therefore independent from cAMP. Viral induction by adenovirus E1a oncoprotein has also been shown to be mediated throug...

It is well known that uninfected mammalian cells contain DNA sequences which are closely related to retroviral genomic segments. However, these sequences seldom (if ever) have been found associated to highly repetitive (satellite) DNA. RPCS is a 348 bp monomer of a major satellite DNA from the South American rodents of the genus Ctenomys. It was found that RPCS contains several elements which are typical of the U3 region of retroviral LTRs. These elements are: a) a polypurine tract; b) two enhancer core sequences; c) two NF1 binding sites; d) two C/EBP binding sites; e) two CCAAT-motifs; f) a TATA box, and g) two putative polyadenylation motifs. Furthermore, the relative positions of these elements are as in the U3 retroviral regions.

Splitting and apparent splicing of ribosomal RNA, both previously unknown in vertebrates, were found in rodents of the genus Ctenomys. Instead of being formed by a single molecule of 4.4 kb, 28S rRNA is split in two molecules of 2.6 and 1.8 kb. A hidden break, mapping within a 106 bp 'intron' located in the D6 divergent region, is expressed in mature ribosomes of liver, lung, heart and spleen, as well as in primary fibroblast cultures. Testis-specific processing eliminates the intron and concomitantly the break site, producing non-split 28S rRNA molecules exclusively in this organ. The intron is flanked by two 9 bp direct repeats, revealing the acquisition by insertion of a novel rRNA processing strategy in the evolution of higher organisms.