Major Sperm Protein Genes from Globodera rostochiensis.

Three genes in the major sperm protein (MSP) gene family from the potato cyst nematode Globodera rostochiensis were cloned and sequenced. In contrast to the absence of introns in Caenorhabditis elegans MSP genes, these genes in G. rostochiensis contained a 57 nucleotide intron, with normal exon-intron boundaries, in the same relative location as the intron in Onchocerca volvulus. The MSP genes of G. rostochiensis had putative CAAT, TATA, and polyadenylation signals. The predicted G. rostochiensis MSP gene product is 126 amino acids long, one residue shorter than the products in the other species. The comparison of MSP amino acid sequences from four diverse nematode species suggests that O. volvulus, Ascaris suum, and C. elegans may be more closely related to each other than they are to G. rostochiensis.

Interaction of microtubules and the mechanism of chromosome movement (zipper hypothesis). 3 Theoretical analysis of energy requirements and computer simulation of chromosome movement.

A theoretical analysis of the energy requirements and a computer analysis of a special case (symmetrical pull in one plane during anaphase), of the zipper model for chromosome movement (Bajer, 1973a,b) is presented. The conclusions are general, however, and can be applied to any stage of mitosis. It is assumed that the movement is due to a series of short lateral interactions (called 'zips') between microtubules, and it is shown that a particular zip could begin if the bending energy requirement for two microtubules is met, and could terminate when a strain restriction is exceeded. The series of zips ends when the energy requirements for initiation of a particular zip are not met. In such conditions, certain predictions concerning the behaviour of microtubules in the spindle can be made (see Conclusions). It is shown that hydrolysis of ATP or GTP can yield sufficient energy to bend microtubules as specified by the model, that insignificant quantities of the triphosphate would be utilized, and that the linearity of chromosome movement predicted by the model is consistent with the linearity observed in vitro.

DNA sequence analysis of the oli1 gene reveals amino acid changes in mitochondrial ATPase subunit 9 from oligomycin-resistant mutants of Saccharomyces cerevisiae.

The nucleotide sequence of the oli1 gene encoding mitochondrial ATPase subunit 9 (76 amino acids) has been determined for five oligomycin-resistant mutants of Saccharomyces cerevisiae. Three of the mutations affect amino acids in the vicinity of the glutamic acid residue 59 at which dicylohexyl carbodiimide binds. Two other mutations lead to substitution of amino acid 23, which would lie very close to residue 59 in the folded hairpin conformation that this protein is thought to adopt in the inner mitochondrial membrane. The apposition of residues 23 and those adjacent to residue 59, lying respectively in the two hydrophobic membrane-spanning arms of subunit 9, is considered to constitute an oligomycin-binding domain. By consideration of the amino acid substitutions in those mutants cross-resistant to venturicidin, a domain of resistance for venturicidin is defined to lie within the oligomycin-binding domain, also centered on residues 23 and 59. These data also clarify the genetic recombination behaviour of alleles previously defined to form part of the oli3 locus (mutants characterized by resistance to both oligomycin and venturicidin) together with alleles defined to form part of the oli1 locus (mutants not cross-resistant to venturicidin). The oli1 and oli3 loci can now be seen to form two overlapping extended groups within the oli1 gene, with sequenced oli3 mutations being as far apart as 125 nucleotides within the subunit 9 coding region of 231 nucleotides.

Biogenesis of mitochondria: DNA sequence analysis of mit- mutations in the mitochondrial oli1 gene coding for mitochondrial ATPase subunit 9 in Saccharomyces cerevisiae.

The nucleotide sequence of the yeast mitochondrial olil gene has been obtained in a series of mit- mutants with mutations in this gene, which codes for subunit 9 of of the mitochondrial ATPase complex. Subunit 9 is the proteolipid, 76 amino acids in length, necessary for the proton translocation function of the membrane Fo-sector. These mutants were classified on the basis of their rescue by a petite strain shown here to retain the entire wild-type olil gene. The mutation in one mit- strain removes a positively charged residue (Arg39----Met) which is likely to be located in a segment of subunit 9 that protrudes from the inner mitochondrial membrane. In a second mit- mutant, a negatively charged residue replaces a conserved glycine residue (Gly18----Asp) in a glycine-rich segment of the protein that is most likely embedded within the membrane. Other mit- mutations result in frameshifts with predicted products 7, 65 and 68 amino acid residues long. In each mit- mutant, there is the loss of one or more of the amino acid residues that are highly conserved among diverse species. The location and nature of specific changes pinpoint amino acid residues in subunit 9 essential to the activity of the mitochondrial ATPase complex.

Biogenesis of Mitochondria: Genetic and molecular analysis of the oli2 region of mitochondrial DNA in Saccharomyces cerevisiae.

A genetic and molecular analysis of the oli2 region in mitochondrial DNA of Saccharomyces cerevisiae has been carried out. The oli2 gene codes for subunit 6 of the mitochondrial ATPase complex. We have isolated a series of 41 mit (-), temperature sensitive and oligomycin resistant mutants containing mutations located in the oli2 region. In addition the sequence in wild type mtDNA of 2820 nucleotides covering the oli2 gene and its flanking regions has been determined. A library of petite mutants which have various deletion end points in the oli2 region has been used to generate marker rescue data that allow the assignment of each mit (-), temperature sensitivity and oligomycin resistance mutation to one of seven uniquely ordered genetic groups. The following physical reference points allow the limits of the physical map positions of a number of the genetic groups to be established: DNA sequence data have been obtained for three of the petites so as to determine unambiguously their relevant deletion end points and the base substitution constituting the mtDNA sequence change in one oligomycin resistant mutant [oli2-23r] has been determined to lie at nucleotide +523 of the oli2 coding region. One group of mit (-) mutations (group N) are known to map within the aap1 gene that is found some 700 bases upstream of the oli2 gene, and which codes for subunit 8 of the mitochondrial ATPase complex. Mutations in the remaining six groups (A-F) map within (or very close to) the coding region of the oli2 gene. Various features of the DNA sequence in and around the oli2 gene are considered, including protein coding regions, sequence divergence in the A,T-rich spacer regions, properties of G,C-rich clusters, and potential ribosome binding sites. Consideration of the transcriptional map of this region of mtDNA suggests that an abundant 4,500 nucleotide transcript may represent the first defined example of a dicistronic messenger RNA in yeast mitochondria carrying distinct coding regions (aap1 and oli2) for two different proteins.

Biogenesis of mitochondria: the mitochondrial gene (aap1) coding for mitochondrial ATPase subunit 8 in Saccharomyces cerevisiae.

A mitochondrial gene (denoted aap1) in Saccharomyces cerevisiae has been characterized by nucleotide sequence analysis of a region of mtDNA between the oxi3 and oli2 genes. The reading frame of the aap1 gene specifies a hydrophobic polypeptide containing 48 amino acids. The functional nature of this reading frame was established by sequence analysis of a series of mit- mutants and revertants. Evidence is presented that the aap1 gene codes for a mitochondrially synthesized polypeptide associated with the mitochondrial ATPase complex. This polypeptide (denoted subunit 8) is a proteolipid whose size has been previously assumed to be 10 kilodaltons based on its mobility on SDS-polyacrylamide gels, but the sequence of the aap1 gene predicts a molecular weight of 5,815 for this protein.

Nucleotide sequence and expression in E. coli of a human interferon-alpha gene selected from a genomic library using synthetic oligonucleotides.

The complete nucleotide sequence of a human interferon-alpha gene is reported. The gene, designated IFN-alpha M1, was isolated from a human genomic library in phage lambda Charon 4A using synthetic oligonucleotides as hybridization probes. Based on a comparison of nucleotide sequence data obtained from this recombinant phage with published interferon-alpha gene sequences, a region of DNA capable of coding for a pre-interferon of 189 amino acids was identified. An AluI fragment containing the coding region for the mature interferon was inserted into the HincII site of the phage M13mp11, resulting in a fusion of portions of the IFN-alpha M1 and the beta-galactosidase genes. Antiviral activity was detected in extracts from E. coli infected with the recombinant M13 phage carrying the fused gene. The antiviral activity was completely neutralized by antibodies to human interferon-alpha.