Publisher Summary There exists a sequence of 8 nucleotides, highly conserved at the boundary between an exon and an intron, referred to as the 5′ splice site (5′ ss). The boundary between an intron and an exon, the 3′ splice site (3′ ss), also exhibits a highly conserved sequence of 4 nucleotides, preceded by a pyrimidine-rich region. To interpret the deluge of sequence information generated by the human genome project, it is important to be able to identify genes in uncharacterized sequences. Without this capability, the sequence information is of little value. For each sequence in GenBank, the splice sites were located using the information in the annotation preceding the sequence, a FEATURES line being followed by multiple lines indicating different features present in the gene. The percentages of lariat windows with high-scoring branch point sequences in all GenBank categories, with the exception of viruses, are higher than those of random sequences. A notable observation is that even the window that contains the second highest frequency of branch points among the five windows analyzed always had a lower frequency of high scoring branch points than expected for a window with a random sequence.

Splice junctions, branch point sites, and exons: sequence statistics, identification, and applications to genome project.

Drosophila doublesex gene controls somatic sexual differentiation by producing alternatively spliced mRNAs encoding related sex-specific polypeptides.

▪ Abstract Striated muscle is an intricate, efficient, and precise machine that contains complex interconnected cytoskeletal networks critical for its contractile activity. The individual units of the sarcomere, the basic contractile unit of myofibrils, include the thin, thick, titin, and nebulin filaments. These filament systems have been investigated intensely for some time, but the details of their functions, as well as how they are connected to other cytoskeletal elements, are just beginning to be elucidated. These investigations have advanced significantly in recent years through the identification of novel sarcomeric and sarcomeric-associated proteins and their subsequent functional analyses in model systems. Mutations in these cytoskeletal components account for a large percentage of human myopathies, and thus insight into the normal functions of these proteins has provided a much needed mechanistic understanding of these disorders. In this review, we highlight the components of striated muscle cyt...

Striated muscle cytoarchitecture: an intricate web of form and function.

Expression of the cardiac ventricular alpha- and beta-myosin heavy chain genes is developmentally and hormonally regulated.

Based on ultrastructural analysis of actively transcribing genes seen in electron micrographs, we present evidence that pre-mRNA splicing occurs with a reasonable frequency on the nascent transcripts of early Drosophila embryo genes and that splice site selection may generally precede polyadenylation. The details of the process observed are in agreement with results from in vitro splicing systems but differ in the more rapid completion of in vivo splicing. For those introns that are removed cotranscriptionally, a series of events is initiated following 3' splice site synthesis, beginning with ribonucleoprotein (RNP) particle formation at the 3' splice site within 48 sec, intron loop formation within 2 min, and splicing within 3 min. The initiation of the process is correlated with 3' splice site synthesis but is independent of 5' splice site synthesis, the position of the intron within the transcript, and the age or length of the transcript. In some cases, introns are removed from the 5' end of a transcript before introns are synthesized at the 3' end, supporting a possible role for the order of transcription in splice site pairing. In general, our observations are consistent with the 'first-come-first-served' principle of splice site selection, although an observed example of exon skipping indicates that alternative splicing possibilities can be accommodated within this general framework.

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Splice site selection, rate of splicing, and alternative splicing on nascent transcripts.

Drosophila muscle myosin heavy chain is encoded by a single-copy gene which is transcribed during both larval and adult development. This myosin gene maps to a chromosomal locus distant from any of the actin genes, but is within a cluster of flight muscle mutations.

Drosophila muscle myosin heavy chain encoded by a single gene in a cluster of muscle mutations.

Genomic and cDNA sequencing studies show that transcripts from the muscle myosin heavy-chain (MHC) gene of Drosophila melanogaster are alternatively spliced, producing RNAs that encode at least two MHC isoforms with different C termini. Transcripts encoding an MHC isoform with 27 unique C-terminal amino acids accumulate during both larval and adult muscle differentiation. Transcripts for the second isoform encode one unique C-terminal amino acid and accumulate almost exclusively in pupal and adult thoracic segments, the location of the indirect flight muscles. The 3' splice acceptor site preceding the thorax-specific exon is unusually purine rich and thus may serve as a thorax-specific splicing signal. We suggest that the alternative C termini of these two MHC isoforms control myofilament assembly and may play a role in generating the distinctive myofilament organizations of flight muscle and other muscle types.

Alternative RNA splicing generates transcripts encoding a thorax-specific isoform of Drosophila melanogaster myosin heavy chain.

Actin Heterogeneity in primary embryonic culture cells from Drosophila melanogaster.

We describe a method for preparing highly enriched cultures of Drosophila myoblasts from a heterogeneous cell population derived from gastrulating embryos. Enriched cultures are prepared by plating this heterogeneous population of cells in medium from which much of the free calcium is chelated by ethylene glycol-bis(beta-aminoethyl ether)N,N,N',N'-tetraacetate (EGTA). Adhesion of myoblasts to tissue culture plastic is better than that of other cell types when plated in this medium. Data concerning cell identity, timing of S phase, and fusion kinetics document the degree of enrichment for myogenic cells and illustrate their synchronous differentiation in vitro.

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J. James Donady

Papers

Drosophila muscle myosin heavy chain encoded by a single gene in a cluster of muscle mutations.

Alternative RNA splicing generates transcripts encoding a thorax-specific isoform of Drosophila melanogaster myosin heavy chain.

Actin Heterogeneity in primary embryonic culture cells from Drosophila melanogaster.

Isolation and partial characterization of Drosophila myoblasts from primary cultures of embryonic cells.

RNA synthesis and coding capacity of polyadenylated and nonpolyadenylated mRNA from cultures of differentiating Drosophila melanogaster myoblasts.