Structure and expression of the overlapping ND4L and ND5 genes of Neurospora crassa mitochondria

Summary Genes homologous to the mammalian mitochondrial NADH dehydrogenase subunit genes ND4L and ND5 were identified in the mitochondrial genome of the filamentous fungus Neurospora crassa, and the structure and expression of these genes was examined. The ND4L gene (interrupted by one intervening sequence) potentially encodes an 89 residue long hydrophobic protein that shares about 26% homology (or 41% homology if conservative amino acid substitutions are allowed) with the analogous human mitochondrial protein. The ND5 gene (which contains two introns) encodes a 715 residue polypeptide that shares 23% homology with the human analogue; a 300 amino acid long region is highly conserved (50% homology) in the two ND5 proteins. The stop codon of the ND4L gene overlaps the initiation codon of the downstream ND5 gene, and the two genes are contranscribed and probably cotranslated. A presumed mature dicistronic (ND4L plus ND5) RNA was detected. The postulated mRNA (about 3.2 kb) contains 5 and 3 non-coding regions of about 86 and 730 nucleotides, respectively; this species is generated from very large precursor RNAs by a complex processing pathway. The ND4L and ND5 introns are all stable after their excision from the precursor species.Abbreviations bp base pairs - rRNA ribosomal RNA - ND NADH dehydrogenase - URF unidentified reading frame - kDal kilodaltons; a.a., amino acid     

The effect of the crossability loci Kr1 and Kr2 on fertilization frequency in hexaploid wheat x maize crosses

Summary Dominant alleles of the Kr1 and Kr2 genes reduce the crossability of hexaploid wheat with many alien species, including rye and Hordeum bulbosum, with Kr1 having the greater effect. However, a cytological study of wheat ovaries fixed 48 h after pollination showed that the wheat genotypes Highbury (kr1, Kr2) and Chinese Spring (Hope 5B) (kr1, kr2) were crossable with Seneca 60 maize, fertilization occurring in 14.4 and 30.7% of embryo sacs respectively. The latter figure was similar to the 29.7% fertilization found in Chinese Spring (kr1, kr2). Most embryo sacs in which fertilization occurred contained an embryo but lacked an endosperm and where an endosperm was formed it was usually highly aberrant. All three wheat x maize combinations were karyotypically unstable and rapidly eliminated maize chromosomes to produce haploid wheat embryos.     

Progeny testing for recessive genes: procedures and interpretations

Summary Methods for calculating the probability of detecting a carrier of a recessive gene by utilizing matings among related individuals are presented for single and litter bearing species. The confidence level for detection of heterozygosity depends upon: (1) the genetic relationship between mates, (2) the number of mates per male and the number of offspring per mate, (3) whether an estimate of recessive gene frequency before selection is available and (4) the magnitude of that frequency. Methods of computing probability of heterozygosity vs homozygosity utilizing Bayes theorem also are presented. In the conventional progeny test method, a sire initially is assumed heterozygous before calculations are made, but no prior information concerning his probable genotype is utilized. In the method using Bayes theorem, prior sources of information from relatives or from estimates of population allele frequency are utilized. This method gives the exact probability that a sire is not a carrier, given prior information and that he produces all normal offspring. These methods could be used in any sexually reproducing species to identify not only detrimental genes but beneficial genes as well.     

The physical map ofMus musculus chromosome 11 reveals evolutionary relationship with different syntenic groups of genes inHomo sapiens

Summary The physical localization of sequences homologous to three cloned genes was determined by in situ hybridization to metaphase chromosomes. Previous work had assigned the skeletal myosin heavy chain gene cluster (Myh), the functional locus for the cellular tumor antigen p53 (Trp53-1), and the cellular homologue of the viral erb-B oncogene (Erbb) toMus musculus chromosome 11 (MMU11). Our results provide regional assignments ofMyh andTrp53-1 to chromosome bands B2C, and ofErbb to bands A1A4. Taken together with in situ mapping of three other loci on MMU 11 (Hox-2 homeobox-containing gene cluster, theSparc protein, and theColla-1 collagen gene), which have been reported elsewhere, these data allowed us to construct a physical map of MMU11 and to compare it with the linkage map of this chromosome. The map positions of the homologous genes on human chromosomes suggest evolutionary relationships of distinct regions of MMU11 with six different human chromosome arms: 1p, 5q, 7p, 16p, 17p, and 17q. The delineation of conserved chromosome regions has important implications for the understanding of karyotype evolution in mammalian species and for the development of animal models of human genetic diseases.     

Mutations in the E1a gene of type 5 adenovirus result in oncogenic transformation of Fischer rat embryo cells

Transformation of a specific clone of Fischer rat embryo (CREF) cells with wild-type 5 adenovirus (Ad5) or the E1a plus E1b transforming gene regions of Ad5 results in epithelioid transformants that grow efficiently in agar but that do not induce tumors when inoculated into nude mice or syngeneic Fischer rats. In contrast, CREF cells transformed by a host-range Ad5 mutant, H5hrl, which contains a single base-pair deletion of nucleotide 1055 in E1a resulting in a 28-kd protein (calculated) in place of the wild-type 51-kd acidic protein, display a cold-sensitive transformation phenotype and an incomplete fibroblastic morphology but surprisingly do induce tumors in nude mice and syngeneic rats. Tumors develop in both types of animals following injection of CREF cells transformed by other cold-sensitive Ad5 E1a mutants (H5dl101 and H5in106), which contain alterations in their 13S mRNA and consequently truncated 289AA proteins. CREF cells transformed with only the E1a gene (0-4.5 m.u.) from H5hrl or H5dl101 also produce tumors in these animals. To directly determine the role of the 13S E1a encoded 289AA protein and the 12S E1a encoded 243AA protein in initiating an oncogenic phenotype in adenovirus-transformed CREF cells, we generated transformed cell lines following infection with the Ad2 mutant pm975, which synthesizes the 289AA E1a protein but not the 243AA protein, and the Ad5 mutant H5dl520 and the Ad2 mutant H2dl1500, which do not produce the 289AA E1a protein but synthesize the normal 243AA E1a protein. All three types of mutant adenovirus-transformed CREF cells induced tumors in nude mice and syngeneic rats. Tumor formation by these mutant adenovirus-transformed CREF cells was not associated with changes in the arrangement of integrated adenovirus DNA or in the expression of adenovirus early genes. These results indicate, therefore, that oncogenic transformation of CREF cells can occur in the presence of a wild-type 13S E1a protein or a wild-type 12S E1a protein when either protein is present alone, but does not occur when both wild-type E1a proteins are present.     

Fertile heteroallelic combinations of mutant alleles of the otu locus of Drosophila melanogaster

Summary This paper describes the ovarian pathologies observed when 108 different heteroallelic combinations were made involving 17 independent mutations at the ovarian tumor (otu) locus. Most of the mutant phenotypes can be explained as graded responses by individual germ cells to different levels of functionally active otu gene product (OGP) synthesized by the mutant cells themselves. The lowest and highest levels of OGP appear to be produced by otu ¹⁰ and otu ¹⁴, respectively. In most heteroallelic ovaries the alleles have additive effects, and hybrid germ cells reach a developmental stage more advanced than the weaker homozygote but less advanced than the stronger homozygote. However, examples of both positive and negative complementation also have been found, and these suggest that the products encoded by different mutant alleles can combine to form dimers or multimers which may be superior or inferior to the homodimers. In flies homozygous for otu ¹¹ most ovarioles contain tumors, but some germ cells are able to develop further than those in otu ¹⁴ homozygotes. This suggests that, while otu ¹¹ produces intermediate levels of OGP, it also produces a second product (which otu ¹⁴ cannot make) that is utilized at the period in oogenesis when development in cells homozygous for otu ¹⁴ is blocked. When otu ¹¹ is combined with any one of eight specific alleles, it allows oocyte/nurse cell syncytia to differentiate that can complete development and undergo embryogenesis, if fertilized. The endopolyploid nurse cells of these hybrids have giant polytene chromosomes, and the presence of GPCs in functionally active, germ-line derived cells provides an interesting new system for experimental study. Analysis of the characteristic ovarian pathologies produced by flies of different genotypes leads to the conclusion that the products of the otu ⁺ gene are utilized during at least six different periods in Drosophila oogenesis.     

Differential expression within a family of novel wound-induced genes in potato

Summary Wounding in higher plants leads to an increased synthesis of specific messenger RNAs. A cDNA clone complementary to a wound-induced message from potato tubers was used to isolate a lambda clone from a genomic library of Salanum tuberosum var. Maris Piper. DNA sequence analysis has shown that this single genomic clone contains two novel wound-induced genes, called win1 and win2, organised in close tandem array. The coding sequences of these two genes are highly homologous and are interrupted by a single intron. However, the sequences of the introns and flanking regions have diverged widely. Win1 and win2 encode cysteine-rich proteins of 200 and 211 amino-acids, respectively, which show striking homologies to several chitin-binding proteins. Southern analysis of genomic DNA has shown that win1 and win2 are members of a small multi-gene family which is estimated to have a minimum of five members per haploid genome of Maris Piper and appears to be conserved within the Solanaceae. We have shown by Northern analysis and S1 mapping that the two genes exhibit differential organ-specific expression after the wounding of a potato plant.     

Mutational analysis of the Bradyrhizobium japonicum common nod genes and further nod box-linked genomic DNA regions

Summary By insertional and deletional marker replacement mutagenesis the common nod region of Bradyrhizobium japonicum was examined for the presence of additional, essential nodulation genes. An open reading frame located in the 800 bp large intergenic region between nodD1 and nodA did not appear to be essential for nodulation of soybean. Furthermore, a strain with a deletion of the nodI- and nodJ-like genes downstream of nodC had a Nod⁺ phenotype. A mutant with a 1.7 kb deletion immediately downstream of nodD1 considerably delayed the onset of nodulation. This region carried a second copy of nodD (nodD2). A nodD1-nodD2 double mutant had a similar phenotype to the nodD2 mutant. Using a 22-mer oligonucleotide probe partially identical to the nod box sequence, a total of six hybridizing regions were identified in B. japonicum genomic DNA and isolated from a cosmid library. Sequencing of the hybridizing regions revealed that at least three of them represented true nod box sequences whereas the others showed considerable deviations from the consensus sequence. One of the three nod box sequences was the one known to be associated with nodA, whereas the other two were located 60 to 70 kb away from nif cluster I. A deletion of one of these two sequences plus adjacent DNA material mmutant 308) led to a reduced nodulation on Vigna radiata but not on soybean. Thus, this region is probably involved in the determination of host specificity.Dedicated to Prof. Giorgio Semenza on the occasion of his 60th birthday