STRUCTURE AND ORGANIZATION OF THE HISTONE GENES OF THE NEWT NOTOPHTHALMUS VIRIDESCENS

I report the isolation and characterization of two classes of histone gene sequences of the newt Notophthalmus viridescens. The first class is the major family of histone genes. I isolated and characterized a cloned example of this family. Each histone gene was localized by hybridization, and its position verified and polarity determined by DNA sequencing. The 9000 base pair cloned DNA segment contains genes for each of the five histones in the order H1-H3-H2B-H2A-H4; all but the H2B gene are transcribed in the same direction. Sizes of the oocyte histone mRNAs were determined by hybridizing cloned newt histone genes to gel fractionated oocyte RNA.;I also studied a class of minor histone genes. I characterized two cloned segments; each contains a single H4 gene but no other histone gene sequences. These H4 genes comprise a minor fraction of the histone genes of the newt, since bands of the sizes predicted by the spacing of restriction enzyme sites in these cloned DNAs cannot be detected in whole genome Southern blots hybridized with H4 gene probes. One of the cloned DNAs was analyzed in detail by sequencing; the H4 gene encodes an aberrant, prematurely terminated peptide. Each of the cloned segments contains a moderately repetitive sequence; the sequences are not the same in the two clones, and neither is present in the major family of histone genes. I analyzed one family of repeated sequences and its relationship to the H4 gene. Surprisingly, the H4 gene interrupts a block of moderately repetitive DNA which is continuous in each of the other three examples studied. A nine base pair sequence occurs as a direct repeat at each of the H4 gene:middle repetitive sequence boundaries. These genes are compared to previously described pseudogenes and transposable elements, and their significance with respect to the evolution of the eukaryotic genome is discussed.;The haploid newt genome contains 600 to 800 clusters which are similar to the cloned segment in their restriction enzyme sites, suggesting that the histone genes comprise a large homogeneous gene family. Unlike other histone genes studied to date, newt histone gene clusters are not adjacent in the genome. Most neighboring clusters are separated by over 50 kilobases of DNA, and some are separated by as much as 100 kilobases. Most or all of the DNA which separates neighboring clusters is satellite DNA. Satellite sequences flank the cloned DNA and the genomic clusters, and must comprise a large part of the DNA which separates neighboring clusters. These findings are discussed with respect to the function of the newt histone genes and the evolution of repeated gene sequences.