The first chordates appear in the fossil record at the time of the Cambrian explosion, nearly 550 million years ago. The modern ascidian tadpole represents a plausible approximation to these ancestral chordates. To illuminate the origins of chordate and vertebrates, we generated a draft of the protein-coding portion of the genome of the most studied ascidian, Ciona intestinalis. The Ciona genome contains ϳ16,000 protein-coding genes, similar to the number in other invertebrates, but only half that found in vertebrates. Vertebrate gene families are typically found in simplified form in Ciona, suggesting that ascidians contain the basic ancestral complement of genes involved in cell signaling and development. The ascidian genome has also acquired a number of lineage-specific innovations, including a group of genes engaged in cellulose metabolism that are related to those in bacteria and fungi.

Abstract. Twelve monoclonal antibodies have been raised against proteins in preparations of Z-disks isolated from Drosophila melanogaster flight muscle. The monoclonal antibodies that recognized Z-hand components were identified by immunofluorescence microscopy of flight muscle myofibrils. These antibodies have identified three Z-disk antigens on immunoblots of myofibrillar proteins. Monoclonal antibodies c~:1-4 recognize a 90-100-kD protein which we identify as o~-actinin on the basis of cross-reactivity with antibodies raised against honeybee and vertebrate ot-actinins. Monoclonal antibodies P:l-4 bind to the high molecular mass protein, projectin, a component of connecting filaments that link the ends of thick filaments to the Z-band in insect asynchronous flight muscles. The anti-projectin antibodies also stain synchronous muscle, but, surprisingly, the epitopes here are within the A-bands, not between the A-and Z-bands, as in flight muscle. Monoclonal antibodies Z(210):1-4 recognize a 210-kD protein that has not been previously shown to be a Z-band structural component. A fourth antigen, resolved as a doublet (,o400/600 kD) on immunoblots of Drosophila fibrillar proteins, is detected by a cross reacting antibody, Z(400):2, raised against a protein in isolated honeybee Z-disks. On Lowicryl sections of asynchronous flight muscle, indirect immunogold staining has localized ot-actinin and the 210-kD protein throughout the matrix of the Z-band, projectin between the Z: and A-bands, and the 400/600-kD components at the I-band/Z-band junction. Drosophila ol-actinin, projectin, and the 400/600-kD components share some antigenic determinants with corresponding honeybee proteins, but no honeybee protein interacts with any of the Z(210) antibodies.T HE Z-band is an electron-dense structural component of striated muscle. It serves as an attachment site for thin filaments and transmits tension between neighboring sarcomeres during contraction. Electron micrographs of both vertebrate muscle and insect fibrillar muscle show Z-bands with a highly ordered, almost crystalline, appearance in cross section (for reviews see 1, 4, 12, 40, 42). Several Z-band proteins have been identified from both vertebrate and insect species (2, 3, 6, 7, 18, 20-22, 25, 29, 30, 33-38); however, the manner in which these proteins are organized within the Z-band lattice is poorly understood. Moreover, the developmental programs that lead to the early organization of the Z-band are only beginning to be clarified.The study of insect Z-bands has been carried out primarily on the flight muscles of the honeybee (Apis) and the giant water bug (Lethocerus). These insects are particularly favorable for biochemical studies of muscle because their size and the predominance of the flight muscle permit isolation of reasonable amounts of homogeneous muscle tissue. The much smaller size of Drosophila presents obstacles for biochemical analyses but facilitates the genetic analyses that are proving to be another useful approach to the study of muscle structure ...

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We have characterized an unusual type of termination signal for T7 RNA polymerase that requires a conserved 7-base pair sequence in the DNA (ATCTGTT in the nontemplate strand). Each of the nucleotides within this sequence is critical for function, as any substitutions abolish termination. The primary site of termination occurs 7 nucleotides downstream from this sequence but is context-independent (that is, the sequence around the site of termination, and in particular the nucleotide at the site of termination, need not be conserved). Termination requires the presence of the conserved sequence and its complement in duplex DNA and is abolished or diminished if the signal is placed downstream of regions in which the non-template strand is missing or mismatched. Under the latter conditions, much of the RNA product remains associated with the template. The latter results suggest that proper resolution of the transcription bubble at its trailing edge and/or displacement of the RNA product are required for termination at this class of signal.A variety of signals have been found to modulate the process of transcript elongation. In general, these have been categorized as falling into the following three classes: pause sites, which temporarily halt the RNA polymerase (RNAP) 1 but subsequently allow resumption of transcription; termination signals, which cause release of the RNA and dissociation of the transcription complex; and arrest sites, at which the RNAP may be halted for a prolonged period but may escape by cleavage and subsequent elongation of the transcript (for review, see Refs. 1 and 2). Among the termination signals, the best characterized involve the formation of a stem-loop structure in the nascent RNA (3-5). Although there have been reports of pause, arrest, or termination signals that do not involve the formation of a structured RNA (see for example Ref. 6), these signals have been less well studied. In this work, we have characterized a sequence-specific pause/termination signal for T7 RNAP and have identified the elements that are required for its function.Two types of signals are known to cause pausing and/or termination by T7 RNAP (7,8). Class I terminators, typified by the signal that is present in the late region of T7 DNA (T⌽), encode RNAs that have the potential to form stable stem-loop structures followed by a run of U residues. These features are reminiscent of many intrinsic terminators utilized by Escherichia coli RNA polymerase, and a number of bacterial termination signals have been shown to cause T7 RNAP to terminate (8 -13). Although the members of this class encode RNAs that share a typical secondary structure, they exhibit little sequence homology.A second type of termination signal recognized by T7 RNAP was first identified in the cloned human prepro-parathyroid hormone (PTH) gene (8,14). These signals (class II signals) do not encode RNAs with an apparent consistent secondary structure but share a common sequence (ATCTGTT, in the nontemplate strand (8,15,16); this work). Additional membe...

Ascidians are simple chordates that are related to, and may resemble, vertebrate ancestors. Comparison of ascidian and vertebrate genomes is expected to provide insight into the molecular genetic basis of chordate/vertebrate evolution. We annotated muscle structural (contractile protein) genes in the completely determined genome sequence of the ascidian Ciona intestinalis, and examined gene expression patterns through extensive EST analysis. Ascidian muscle protein isoform families are generally of similar, or lesser, complexity in comparison with the corresponding vertebrate isoform families, and are based on gene duplication histories and alternative splicing mechanisms that are largely or entirely distinct from those responsible for generating the vertebrate isoforms. Although each of the three ascidian muscle types - larval tail muscle, adult body-wall muscle and heart - expresses a distinct profile of contractile protein isoforms, none of these isoforms are strictly orthologous to the smooth-muscle-specific, fast or slow skeletal muscle-specific, or heart-specific isoforms of vertebrates. Many isoform families showed larval-versus-adult differential expression and in several cases numerous very similar genes were expressed specifically in larval muscle. This may reflect different functional requirements of the locomotor larval muscle as opposed to the non-locomotor muscles of the sessile adult, and/or the biosynthetic demands of extremely rapid larval development.