Transforming growth factor-β1 (TGF-

Transforming growth factor 31 (TGFj81) -

No abstract

Several clones were isolated from a rat genonic library in order to further characterize a region of variability within the third membrane-spanning region of the fourth motif (IVS3) of the L-type voltage-dependent calcium channel. We report here that this diversity arises from alternative splicing of a primary transcript containing a single pair of adjacent exons each encoding a unique sequence for the IVS3 region. Definitive proof of a mutually exclusive splicing mechanism was obtained by genomic mapping of flanking upstream and downstream exons and by extensive sequence analysis of the relevant exon/intron boundaries. S1 nuclease protection experiments revealed that both variant forms of the IVS3 were equally expressed in newborn and fetal rat heart, whereas only a single isoform predominated in adult rat heart. The results demonstrate the existence of an important developmentally regulated switch mediated by alternatively spliced exons in cardiac tissue at a time when major changes in excitation occur.Voltage-dependent calcium channels (VDCCs) are a diverse class of molecules found in all excitable cells (1-4). These channels are essential for many cellular functions such as muscle contraction, propagation of action potentials, maintenance of electrical activity, excitation-secretion coupling, and neurotransmitter regulation (2, 3). The L-type VDCC from rabbit skeletal muscle has been purified and extensively characterized biochemically. The channel exists as a pentameric macromolecule consisting of a1, a<-8, 3,, and y subunits, all of which have been cloned and sequenced (4-7).Functionally, the a1 subunit alone can act as a VDCC (8, 9) and contains binding sites for calcium antagonists (10, 11), although native gating characteristics are lacking (9). The a2-8 and, in some cases, P subunits significantly increase current amplitude displayed by expressed a1 subunits (8,(12)(13)(14). More importantly, the 83 subunit modulates gating properties of the expressed skeletal muscle a1 subunit in mammalian L-cells (14). In Xenopus oocytes, the expression of the skeletal muscle 'y subunit determines the inactivation properties of the coexpressed cardiac a1 subunit (15). Thus, a normal functioning calcium channel probably requires the presence of all the other subunits.The a1 subunits from rabbit cardiac muscle and lung (8, 13, 16), rat aorta (17), and carp skeletal muscle (18) have been cloned and their primary structures have been described.Like the skeletal muscle a1 subunit, the cardiac and smooth muscle isoforms are large hydrophobic polypeptides comprising four repeating motifs (I-IV), each containing six putative transmembrane segments (S1-S6), flanked by Nand C-terminal cytoplasmic domains. Sequence analysis suggests that the cardiac and smooth muscle a1 subunits are encoded by the same gene, which is distinct from that encoding the skeletal muscle channel (9,12,17).In the cloning of the rat aorta a1 subunit, we described the existence of two cDNA species that were identical in all but the third membrane...

Cooperative DNA binding by transcription factors that bind to separate recognition sites is likely to require bending of intervening sequences and the appropriate orientation of transcription factor binding. We investigated DNA bending in complexes formed by the basic region-leucine zipper domains of Fos and Jun with the DNA binding region of nuclear factor of activated T cells 1 (NFAT1) at composite regulatory elements using gel electrophoretic phasing analysis. The NFAT1-Fos-Jun complex induced a bend at the ARRE2 site that was distinct from the sum of the bends induced by NFAT1 and Fos-Jun separately. We designate this difference DNA bending cooperativity. The bending cooperativity was directed toward the interaction interface between Fos-Jun and NFAT1. We also examined the influence of NFAT1 on the orientation of Fos-Jun heterodimer binding using a novel fluorescence resonance energy transfer assay. The interaction with NFAT1 could reverse the orientation of Fos-Jun heterodimer binding to the ARRE2 site. The principal determinants of both cooperative DNA bending and oriented heterodimer binding were localized to three amino acid residues at the amino-terminal ends of the leucine zippers of Fos and Jun. Consequently, interactions between transcription factors can remodel promoters by altering DNA bending and the orientation of heterodimer binding.