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PubMed1988

SPA1: a gene important for chromosome segregation and other mitotic functions in S. cerevisiae.

Human autoantibodies that recognize the spindle poles of mammals, plants, and insects were found to recognize two antigens in yeast. One of these proteins, called SPA1 (for Spindle Pole Antigen), is antigenically related to the spindle poles of a diverse set of organisms. The gene encoding SPA1 was cloned by immunoscreening a lambda gt11 yeast genomic DNA expression library with autoantibody probes. Mutational analysis of the SPA1 gene demonstrates that it is important for cell growth, chromosome segregation, and other cellular processes; spa1 mutants are viable but grow poorly at 30 degrees C, missegregate chromosomes at an increased frequency, and often contain deformed spindles. A significant fraction of spa1 mutant cells contain two or more nuclei, and others contain none; these abnormal cells may arise through a nuclear migration defect. Thus SPA1 represents a new fidelity gene that is important for chromosome segregation and other mitotic functions.

Autoantibodies↗
PubMed2000

Orbit, a novel microtubule-associated protein essential for mitosis in Drosophila melanogaster.

We describe a Drosophila gene, orbit, that encodes a conserved 165-kD microtubule-associated protein (MAP) with GTP binding motifs. Hypomorphic mutations in orbit lead to a maternal effect resulting in branched and bent mitotic spindles in the syncytial embryo. In the larval central nervous system, such mutants have an elevated mitotic index with some mitotic cells showing an increase in ploidy. Amorphic alleles show late lethality and greater frequencies of hyperploid mitotic cells. The presence of cells in the hypomorphic mutant in which the chromosomes can be arranged, either in a circular metaphase or an anaphase-like configuration on monopolar spindles, suggests that polyploidy arises through spindle and chromosome segregation defects rather than defects in cytokinesis. A role for the Orbit protein in regulating microtubule behavior in mitosis is suggested by its association with microtubules throughout the spindle at all mitotic stages, by its copurification with microtubules from embryonic extracts, and by the finding that the Orbit protein directly binds to MAP-free microtubules in a GTP-dependent manner.

Alleles↗
PubMed1987

Transcriptional activity of constitutive heterochromatin in the mammal Microtus agrestis (Rodentia, Cricetidae).

Two cytological approaches were applied to the analysis of genetic activity in euchromatin and heterochromatin in the European field vole Microtus agrestis. The first is based on the transcriptional activity of prematurely condensed chromosomes, the second on in situ hybridization of labeled cellular RNA to metaphase chromosomes. The results show that in cultivated fibroblasts, the transcriptional activity of constitutive heterochromatin is in the same order of magnitude as that of euchromatin; its chromosomal pattern is nonrandom. It is concluded that--as has already been shown for insects and amphibians--also in mammals, constitutive heterochromatin is transcribed in its decondensed state but is genetically inactive when forming chromocenters at interphase.

Animals↗
PubMed2002

Mass rearing of temperature sensitive genetic sexing strains in the Mediterranean fruit fly (Ceratitis capitata).

Genetic sexing strains (GSS) based on the temperature sensitive lethal (tsl) mutation are being used to produce sterile male medflies for large scale sterile insect technique (SIT) programmes for this pest. The use of male-only strains increases the overall efficiency of the technique. Currently more than 1.4 billion sterile male-only pupae are produced per week in different facilities around the world. Due to the mutations used to construct these strains, that is, translocations and selectable markers, they require different and more careful mass rearing procedures than do bisexual strains (BSS). The basic rearing technology has been developed and can be used to produce only males on a predictable basis to a level of 99.9% accuracy. If specific rearing procedures are followed, then tsl-based GSS has a rearing efficiency that is equal to that of a BSS and it is already know that males produced by the tsl-based GSS are of equal quality to males produced by BSS. Based on current rearing technology the cost of production of male pupae is about the same for both types of strain. This is due to the large colony that is required for the tsl-based GSS. This paper discusses the considerations that need to be taken into account during mass rearing of GSS and identifies the most efficient production processes that are currently available.

Animal Feed↗
PubMed1999

Translation initiation at the CUU codon is mediated by the internal ribosome entry site of an insect picorna-like virus in vitro.

AUG-unrelated translation initiation was found in an insect picorna-like virus, Plautia stali intestine virus (PSIV). The positive-strand RNA genome of the virus contains two nonoverlapping open reading frames (ORFs). The capsid protein gene is located in the 3'-proximal ORF and lacks an AUG initiation codon. We examined the translation mechanism and the initiation codon of the capsid protein gene by using various dicistronic and monocistronic RNAs in vitro. The capsid protein gene was translated cap independently in the presence of the upstream cistron, indicating that the gene is translated by internal ribosome entry. Deletion analysis showed that the internal ribosome entry site (IRES) consisted of approximately 250 bases and that its 3' boundary extended slightly into the capsid-coding region. The initiation codon for the IRES-mediated translation was identified as the CUU codon, which is located just upstream of the 5' terminus of the capsid-coding region by site-directed mutagenesis. In vitro translation assays of monocistronic RNAs lacking the 5' part of the IRES showed that this CUU codon was not recognized by scanning ribosomes. This suggests that the PSIV IRES can effectively direct translation initiation without stable codon-anticodon pairing between the initiation codon and the initiator methionyl-tRNA.

Amino Acid Sequence↗
PubMed2006

Living with genome instability: the adaptation of phytoplasmas to diverse environments of their insect and plant hosts.

Phytoplasmas ("Candidatus Phytoplasma," class Mollicutes) cause disease in hundreds of economically important plants and are obligately transmitted by sap-feeding insects of the order Hemiptera, mainly leafhoppers and psyllids. The 706,569-bp chromosome and four plasmids of aster yellows phytoplasma strain witches' broom (AY-WB) were sequenced and compared to the onion yellows phytoplasma strain M (OY-M) genome. The phytoplasmas have small repeat-rich genomes. This comparative analysis revealed that the repeated DNAs are organized into large clusters of potential mobile units (PMUs), which contain tra5 insertion sequences (ISs) and genes for specialized sigma factors and membrane proteins. So far, these PMUs appear to be unique to phytoplasmas. Compared to mycoplasmas, phytoplasmas lack several recombination and DNA modification functions, and therefore, phytoplasmas may use different mechanisms of recombination, likely involving PMUs, for the creation of variability, allowing phytoplasmas to adjust to the diverse environments of plants and insects. The irregular GC skews and the presence of ISs and large repeated sequences in the AY-WB and OY-M genomes are indicative of high genomic plasticity. Nevertheless, segments of approximately 250 kb located between the lplA and glnQ genes are syntenic between the two phytoplasmas and contain the majority of the metabolic genes and no ISs. AY-WB appears to be further along in the reductive evolution process than OY-M. The AY-WB genome is approximately 154 kb smaller than the OY-M genome, primarily as a result of fewer multicopy sequences, including PMUs. Furthermore, AY-WB lacks genes that are truncated and are part of incomplete pathways in OY-M.

Animals↗
PubMed2001

Molecular biology of the chromo domain: an ancient chromatin module comes of age.

The chromo domain motif is found in proteins from fungi, protists, plants, fish, insects, amphibians, birds, and mammals. The chromo domain peptide fold may have its origins as a chromosomal protein in a common ancestor of archea and eukaryota, making it a particularly ancient protein structural module. Chromo domains have been found in single or multiple copies in proteins with diverse structures and activities, most or all of which are connected with chromosome structure/function. In this review, our current knowledge of chromo domain properties is summarized and a variety of contexts in which chromo domains participate in aspects of chromatin metabolism are discussed.

Amino Acid Sequence↗
PubMed2003

[Variation and evolution of meiosis].

Meiosis arose in the evolution of primitive unicellular organisms as a part of sexual process. One type of meiosis, the so-called classical type, predominates in all kingdoms of eukaryotes. Meiosis is controlled by hundreds of genes, both shared with mitosis and specifically meiotic ones. In a wide range of taxa, which in some cases include kingdoms, meiotic genes and features obey Vavilov's law of homologous variation series. Synaptonemal complexes (SCs) temporarily binding homologous chromosomes at prophase I, ensure precise and equal crossing over and interference. SC proteins have 60-80% homology within the class of mammals but differ from the corresponding proteins in fungi and plants. Thus, nonhomologous SC proteins perform similar functions in different taxa. Some recombination enzymes in fungi and insects have common epitopes. The molecular mechanism of recombination is inherited by eukaryotes from prokaryotes and operates in special compartments: SC recombination nodules. Chiasmata, i.e., physical crossovers of nonsister chromatids, are preserved in bivalents until metaphase I due to local cohesion of sister chromatids in the remaining SC fragments. Owing to chiasmata, homologous chromosomes participate in meiosis I in pairs rather than individually, which, along with unipolarity of kinetochores (only in meiosis 1), ensures segregation of homologous chromosomes. The appearance of SC and chiasmata played a key role in the evolution of unicellular organisms since it promoted the development of a progressive type of meiosis. Some lower eukaryotes retain primitive meiosis types. These primitive modes of meiosis also occur in the sex of some insects that is heterozygous for sex chromosomes. I suggest an explanation for these cases. Mutations at meiotic genes impair meiosis; however, due to the preservation of archaic meiotic genes in the genotype, bypass metabolic pathways arise, which provide partial rescue of the traits damaged by mutations. Individual blocks of genetic program of meiotic regulation have probably evolved independently.

Animals↗
PubMed1997

Organization of DNA sequences near the centromere of the Drosophila melanogaster Y chromosome.

The structural analysis of a yeast artificial chromosome clone from Drosophila melanogaster enriched in dodecasatellite sequences has led us to find a new retrotransposon that we have called Circe. Moreover, this retrotransposon has allowed the isolation of a contig encompassing approximately 200 kb near the centromere of the Y chromosome, providing an entry point into a region from which very little sequence information has been obtained to date. The molecular characterization of the contig has shown the presence of HeT-A telomeric retrotransposons close to the centromere of the Y chromosome, suggesting a telocentric origin for this submetacentric chromosome.

Animals↗
PubMed1999

[Chromosome rearrangement: two ways of influencing crossing over in Drosophila].

Insertion of the Y-material into the 34A Is(Y;2L)419 region diminished recombinational length of the left arm of chromosome 2 (2L) from 49.1 to 15.0 cM. This decrease was compensated by the increase of recombinational length in the other chromosomal arms due to interchromosomal effect. The increase in the X chromosome was 11.4 cM; it was 2.0 cM in chromosome 2R; and 17.3 cM in chromosome 3. The insertion-induced decrease of the 2L recombinational length could be eliminated by evoking interchromosomal effects from other chromosomes. The presence of the inversion in the X chromosome increased the 2L recombinational length from 15.0 to 30.2 cM, while its association with the In(3LR)D inversion increased this length to 45.6 cM. The interchromosomal effects in the inductor chromosome were induced by distortion of pairing rather than by the low recombinational length of this chromosome. For example, the interchromosomal effect of the insertion on the X chromosome was higher in the Is(Y;2L)/+; In(3LR)/+ females than in the Is(Y;2L)/+; +/+ females (15.4 versus 11.5 cM), though the 2L recombinational length in the females with the former genotype (30.2 cM) was twofold higher than in females with the latter genotype (15.0 cM). It is suggested that chromosomal rearrangement hampers the development of local contacts in the homologues. This delay affects crossing over in the given pair of homologues in two ways: directly via diminishing the number of exchange sites, and indirectly through regulatory delay of crossing over determination in the meiocyte. The effects of the insertion on crossing over in nonhomologous chromosomes are implemented by through the second way.

Animals↗
PubMed2001

From first base: the sequence of the tip of the X chromosome of Drosophila melanogaster, a comparison of two sequencing strategies.

We present the sequence of a contiguous 2.63 Mb of DNA extending from the tip of the X chromosome of Drosophila melanogaster. Within this sequence, we predict 277 protein coding genes, of which 94 had been sequenced already in the course of studying the biology of their gene products, and examples of 12 different transposable elements. We show that an interval between bands 3A2 and 3C2, believed in the 1970s to show a correlation between the number of bands on the polytene chromosomes and the 20 genes identified by conventional genetics, is predicted to contain 45 genes from its DNA sequence. We have determined the insertion sites of P-elements from 111 mutant lines, about half of which are in a position likely to affect the expression of novel predicted genes, thus representing a resource for subsequent functional genomic analysis. We compare the European Drosophila Genome Project sequence with the corresponding part of the independently assembled and annotated Joint Sequence determined through "shotgun" sequencing. Discounting differences in the distribution of known transposable elements between the strains sequenced in the two projects, we detected three major sequence differences, two of which are probably explained by errors in assembly; the origin of the third major difference is unclear. In addition there are eight sequence gaps within the Joint Sequence. At least six of these eight gaps are likely to be sites of transposable elements; the other two are complex. Of the 275 genes in common to both projects, 60% are identical within 1% of their predicted amino-acid sequence and 31% show minor differences such as in choice of translation initiation or termination codons; the remaining 9% show major differences in interpretation.

Animals↗
PubMed1993

A fertility region on the Y chromosome of Drosophila melanogaster encodes a dynein microtubule motor.

A clone encoding a portion of the highly conserved ATP-binding domain of a dynein heavy-chain polypeptide was mapped to a region of the Drosophila melanogaster Y chromosome. Dyneins are large multisubunit enzymes that utilize the hydrolysis of ATP to move along microtubules. They were first identified as the motors that provide the force for flagellar and ciliary bending. Seven different dynein heavy-chain genes have been identified in D. melanogaster by PCR. In the present study, we demonstrate that one of the dynein genes, Dhc-Yh3, is located in Y chromosome region h3, which is contained within kl-5, a locus required for male fertility. The PCR clone derived from Dhc-Yh3 is 85% identical to the corresponding region of the beta heavy chain of sea urchin flagellar dynein but only 53% identical to a cytoplasmic dynein heavy chain from Drosophila. In situ hybridization to Drosophila testes shows Dhc-Yh3 is expressed in wild-type males but not in males missing the kl-5 region. These results are consistent with the hypothesis that the Y chromosome is needed for male fertility because it contains conventional genes that function during spermiogenesis.

Amino Acid Sequence↗
PubMed1995

The Sex-lethal gene homologue in Chrysomya rufifacies is highly conserved in sequence and exon-intron organization.

A great variety of sex determination mechanisms exists in insect species. In Drosophila melanogaster sex is determined by the ratio between X chromosomes and autosomes, while in the blowfly Chrysomya rufifacies it is maternally determined. A cascade of genes which are involved in sex determination has been identified in D. melanogaster with the Sex-lethal gene (Sxl) as the key gene. We screened genomic libraries of C. rufifacies with a probe of the Sxl gene from D. melanogaster and isolated a genomic region that included most of the homologous gene. DNA- and protein-sequence comparison showed a high percent identity between the Chrysomya and the Drosophila gene. Up to 90% identity of the amino acid sequences was found in the region that contained the RNA-binding domains. The degree of identity is much lower outside of this functionally important region (18% identity). cDNA analysis showed a highly conserved exon-intron structure between the two species, although sex-specific splicing as used in D. melanogaster for the regulation of Sxl activity, could not be detected in C. rufifacies.

Amino Acid Sequence↗
PubMed2004

Isolation and characterization of Y chromosome sequences from the African malaria mosquito Anopheles gambiae.

The karyotype of the African malaria mosquito Anopheles gambiae contains two pairs of autosomes and a pair of sex chromosomes. The Y chromosome, constituting approximately 10% of the genome, remains virtually unexplored, despite the recent completion of the A. gambiae genome project. Here we report the identification and characterization of Y chromosome sequences of total length approaching 150 kb. We developed 11 Y-specific PCR markers that consistently yielded male-specific products in specimens from both laboratory colony and natural populations. The markers are characterized by low sequence polymorphism in samples collected across Africa and by presence in more than one copy on the Y. Screening of the A. gambiae BAC library using these markers allowed detection of 90 Y-linked BAC clones. Analysis of the BAC sequences and other Y-derived fragments showed massive accumulation of a few transposable elements. Nevertheless, more complex sequences are apparently present on the Y; these include portions of an approximately 48-kb-long unmapped AAAB01008227 scaffold from the whole genome shotgun assembly. Anopheles Y appears not to harbor any of the genes identified in Drosophila Y. However, experiments suggest that one of the ORFs from the AAAB01008227 scaffold represents a fragment of a gene with male-specific expression.

Animals↗
PubMed1996

Mutations in the gene stand still disrupt germ cell differentiation in Drosophila ovaries.

During oogenesis in Drosophila, germ cells appear in sequential clusters of 16 interconnected cells. The events surrounding the differentiation of these cells are not fully understood. Here we present genetic and morphological analysis of mutations in the gene stand still (stil). Through complementation analyses we have refined the location of this gene to cyological region 49B-C. Our analyses of ovaries from ethylmethane sulfonate (EMS)-induced mutant alleles of this gene suggest that mutations in the stil gene produce a wide range of phenotypic abnormalities, from the absence of germ cells in the most severe alleles, to egg chambers with cytoskeletal defects in the less severe alleles. Our results suggest a role for this gene in specifying or maintaining a cytoskeletal component, with consequences during oogenesis and possibly during germ line sex determination.

Actins↗
PubMed1999

The hydroxamic acid pathway.

An important component of general defence mechanisms of plants are toxic secondary metabolites that function as natural pesticides. The cyclic hydroxamic acids DIBOA (2,4-dihydroxy-1,4-benzoxazin-3-one) and DIMBOA (2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one) play an important role in the chemical defence of cereals against pests such as insects and pathogenic fungi and bacteria. Five genes that are clustered on chromosome four are sufficient to encode the enzymes to synthesize DIBOA. The first gene in the pathway, Bx1, encodes an enzyme resembling a tryptophan synthase alpha subunit that catalyses the formation of indole and thereby establishes the branchpoint that leads to the secondary metabolites. Four cytochrome P450-dependent monooxygenases encoded by Bx2-Bx5 catalyse consecutive hydroxylations to form DIBOA. This pathway can be generalized for grasses, since identical enzyme activities have been found in rye. The pathway is relatively short and begins with a metabolite ubiquitous to plants. Therefore DIBOA biosynthesis could be introduced into other plant species to confer improved disease resistance.

Hydroxamic Acids↗
PubMed2004

The dice of fate: the csd gene and how its allelic composition regulates sexual development in the honey bee, Apis mellifera.

Perhaps 20% of known animal species are haplodiploid: unfertilized haploid eggs develop into males and fertilized diploid eggs into females. Sex determination in such haplodiploid species does not rely on a difference in heteromorphic sex chromosome composition but the genetic basis has been elucidated in some hymenopteran insects (wasps, sawflies, ants, bees). In these species, the development into one sex or the others depends on an initial signal whether there is only one allele or two different alleles of a single gene, the complementary sex determiner (csd), in the zygotic genome. The gene has been most-recently identified in the honey bee and has been found to encode an arginine serine-rich (SR) type protein. Heterozygosity generates an active protein that initiates female development while hemizygosity/homozygosity results in a non-active CSD protein and default male development. I will discuss plausible models of how the molecular decision of male and female is made and implemented. Comparison to hierarchies of dipteran insects suggests that SR-type protein has facilitated the differentiation of sex-determining systems and hierarchies.

Alleles↗