Proceedings of the National Academy of Sciences of the United States of America
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Behavioral and sensory basis of courtship success in Drosophila melanogaster. In Drosophila some individuals are more successful at mating than others. Reproductive fitness is strongly dependent upon the ability to recognize and compete for members of the opposite sex. Experiments were designed to answer two questions. (i) What behavioral components are characteristic or predictive of successful courtship? and (ii) How important is the information transmitted in the different sensory channels for courtship success in each sex? These questions were approached by two experimental procedures. Flies having a sensory deficiency (olfactory, auditory, or visual) competed with wild-type flies of the same sex for mates. Males were found to rely upon sensory channels different from those used by females in order to court successfully. In addition, the courtships of pairs of various genotypes were recorded and subjected to multivariate analysis. The multivariate courtship profiles deviated most widely from those of successful wild-type pairs when the male or female was unable to receive information in the sensory channel most important for successful mating by that sex. Both sequential and quantitative courtship properties were altered when one participant was deficient in ability to receive certain sensory information.
Proceedings of the National Academy of Sciences of the United States of America - Tập 84 Số 17 - Trang 6200-6204 - 1987
Sexual orientation in Drosophila is altered by the satori mutation in the sex-determination gene fruitless that encodes a zinc finger protein with a BTB domain. We have isolated a new Drosophila mutant, satori (sat), the males of which do not court or copulate with female flies. The sat mutation comaps with fruitless (fru) at 91B and does not rescue the bisexual phenotype of fru, indicating that sat is allelic to fru (fru(sat)). The fru(sat) adult males lack a male-specific muscle, the muscle of Lawrence, as do adult males with other fru alleles. Molecular cloning and analyses of the genomic and complementary DNAs indicated that transcription of the fru locus yields several different transcripts. The sequence of fru cDNA clones revealed a long open reading frame that potentially encodes a putative transcription regulator with a BTB domain and two zinc finger motifs. In the 5' noncoding region, three putative transformer binding sites were identified in the female transcript but not in male transcripts. The fru gene is expressed in a population of brain cells, including those in the antennal lobe, that have been suggested to be involved in determination of male sexual orientation. We suggest that fru functions downstream of tra in the sex-determination cascade in some neural cells and that inappropriate sexual development of these cells in the fru mutants results in altered sexual orientation of the fly.
Proceedings of the National Academy of Sciences of the United States of America - Tập 93 Số 18 - Trang 9687-9692 - 1996
Gender-selective patterns of aggressive behavior in <i>Drosophila melanogaster</i>
Complex behaviors, such as aggression, are comprised of distinct stereospecific behavioral patterns (modules). How such patterns get wired into nervous systems remains unknown. Recently, we reported on a quantitative analysis of fighting behavior in male flies of the common Canton-S strain of
Drosophila melanogaster
. Here, we report a similar analysis of fighting behavior in females of the same species. Fights were carried out between pairs of virgin and pairs of mated females in competition for a yeast resource. Each fight was videotaped and analyzed by using transition matrices and Markov chain analyses. We observe only small difference in fighting intensity between virgin and mated females. In contrast to what is seen in male fights, however, no clear hierarchical relationship is formed in the female fights. A further comparison of the behavioral patterns making up male and female fights reveals that some modules are shared by both sexes, whereas others are highly selective. Within the shared components, transitions between the modules also show gender-selective differences. By using the powerful genetic methods available for examining behavior in fruit flies, it should be possible to use the gender-selective differences in fighting behavior to address the question of how these behavioral patterns get established in the brains of fruit flies.
Proceedings of the National Academy of Sciences of the United States of America - Tập 101 Số 33 - Trang 12342-12347 - 2004
A central neural circuit for experience-independent olfactory and courtship behavior in <i>Drosophila melanogaster</i>
We have studied the function of the major central olfactory
pathway in fruit flies. Key elements of this pathway, the projection
neurons (PNs), connect the antennal lobes with the lateral
protocerebrum both directly and indirectly, the latter via the mushroom
bodies (MBs). Transgenic expression of tetanus toxin in the majority of
PNs and few MB neurons leads to defects in odor detection and male
courtship. Considering behavioral data from flies lacking MBs, our
results argue that the direct PN-to-lateral protocerebrum pathway is
necessary and sufficient to process these experience-independent
behaviors. Moreover, the involvement of an olfactory pathway in male
courtship suggests a role of volatile attractive female pheromones in
Drosophila
.
Proceedings of the National Academy of Sciences of the United States of America - Tập 98 Số 26 - Trang 15336-15341 - 2001
A family of genes encoding neurotransmitter transporters. The genomic and cDNA clones of the mouse gamma-aminobutyric acid transporter were sequenced and analyzed. The genomic clone contains 12 introns including 1 intron prior to the initiator methionine. The second intron comes immediately after the stretch of amino acids that is most conserved among the neurotransmitter transporters sequenced so far. By using a probe constructed according to this conserved region, several partial genomic clones were isolated. Sequence analysis of those clones reveals not only homology to the family of neurotransmitter transporters within the reading frame but also an identical location of an exon-intron junction after the conserved region. A search of the GenBank data base (April 1991) revealed that two invertebrate genes exhibit homology to the conserved sequence of the above family. One, a Drosophila melanogaster gene, encoded the N-terminal part of a protein homologous to neurotransmitter transporters and the second was in Caenorhabditis elegans. The Drosophila gene contains an intron that starts at a position identical to the corresponding positions of all the mammalian genes of the family.
Proceedings of the National Academy of Sciences of the United States of America - Tập 89 Số 14 - Trang 6639-6643 - 1992
Tracking the establishment of local endemic populations of an emergent enteric pathogen Significance
Shigella sonnei
is a globally emerging agent of bacterial dysentery. Here, we use genomics to examine the microevolution of
S. sonnei
in Vietnam. We show that
S. sonnei
was introduced into Vietnam in the early 1980s, where it continued to evolve, spreading geographically to establish localized founder populations. The population in Ho Chi Minh City has undergone several localized clonal replacement events, during which a small number of microevolutionary changes have risen to dominance. These changes, induced by horizontal gene transfer and substitution mutations, confer high-level antimicrobial resistance and the ability to kill other gut bacteria. This work provides a unique, high-resolution insight into the microevolution of a pioneering human pathogen during its establishment in a new host population.
Proceedings of the National Academy of Sciences of the United States of America - Tập 110 Số 43 - Trang 17522-17527 - 2013
Biosynthesis of a protein containing a nonprotein amino acid by Escherichia coli: L-2-aminohexanoic acid at position 21 in human epidermal growth factor. Endeavoring to develop a method to biosynthesize proteins substituted with nonprotein amino acids, we attempted the incorporation of L-2-aminohexanoic acid (Ahx) into human epidermal growth factor (hEGF). Escherichia coli YK537 strain harboring plasmid pTA1522, which has the phoA promoter-phoA signal peptide-hEGF gene, was used. Cells were cultured first in high-phosphate medium and then, for induction of the hEGF-encoding gene, transferred to low-phosphate medium containing Ahx (0.25 mg/ml). hEGF and Ahx-substituted hEGF, [Ahx21]hEGF, secreted into the periplasm were recovered. After treatment with H2O2, [Ahx21]-hEGF was clearly separated from methionine-oxidized hEGF by one-step reverse-phase HPLC. Substitution of the methionine residue of hEGF with Ahx was confirmed by the amino acid analysis of [Ahx21]hEGF. The three biological activities of [Ahx21]hEGF were the same as those of hEGF. From the successful production of [Ahx21]hEGF, a basic strategy was established for preparing proteins substituted with nonprotein amino acid (alloprotein). Induction of the phoA promoter of pho regulon and secretion of the product to the periplasm may depress heat shock-like responses and subsequent hydrolysis of the product by cytoplasmic protease.
Proceedings of the National Academy of Sciences of the United States of America - Tập 85 Số 17 - Trang 6237-6241 - 1988
Pyrrolysine is not hardwired for cotranslational insertion at UAG codons
Pyrrolysine (Pyl), the 22nd naturally encoded amino acid, gets acylated to its distinctive UAG suppressor tRNA
Pyl
by the cognate pyrrolysyl-tRNA synthetase (PylRS). Here we determine the RNA elements required for recognition and aminoacylation of tRNA
Pyl
in vivo
by using the Pyl analog
N
-ε-cyclopentyloxycarbonyl-
l
-lysine. Forty-two
Methanosarcina barkeri
tRNA
Pyl
variants were tested in
Escherichia coli
for suppression of the
lac
amber A24 mutation; then relevant tRNA
Pyl
mutants were selected to determine
in vivo
binding to
M. barkeri
PylRS in a yeast three-hybrid system and to measure
in vitro
tRNA
Pyl
aminoacylation. tRNA
Pyl
identity elements include the discriminator base, the first base pair of the acceptor stem, the T-stem base pair G51:C63, and the anticodon flanking nucleotides U33 and A37. Transplantation of the tRNA
Pyl
identity elements into the mitochondrial bovine tRNA
Ser
scaffold yielded chimeric tRNAs active both
in vitro
and
in vivo
. Because the anticodon is not important for PylRS recognition, a tRNA
Pyl
variant could be constructed that efficiently suppressed the
lac
opal U4 mutation in
E. coli
. These data suggest that tRNA
Pyl
variants may decode numerous codons and that tRNA
Pyl
:PylRS is a fine orthogonal tRNA:synthetase pair that facilitated the late addition of Pyl to the genetic code.
Proceedings of the National Academy of Sciences of the United States of America - Tập 104 Số 9 - Trang 3141-3146 - 2007
Engineering a tRNA and aminoacyl-tRNA synthetase for the site-specific incorporation of unnatural amino acids into proteins <i>in vivo</i>
In an effort to expand the scope of protein mutagenesis, we have completed the first steps toward a general method to allow the site-specific incorporation of unnatural amino acids into proteins
in vivo
. Our approach involves the generation of an “orthogonal” suppressor tRNA that is uniquely acylated in
Escherichia coli
by an engineered aminoacyl-tRNA synthetase with the desired unnatural amino acid. To this end, eight mutations were introduced into tRNA
2
Gln
based on an analysis of the x-ray crystal structure of the glutaminyl-tRNA aminoacyl synthetase (GlnRS)–tRNA
2
Gln
complex and on previous biochemical data. The resulting tRNA satisfies the minimal requirements for the delivery of an unnatural amino acid: it is not acylated by any endogenous
E. coli
aminoacyl-tRNA synthetase including GlnRS, and it functions efficiently in protein translation. Repeated rounds of DNA shuffling and oligonucleotide-directed mutagenesis followed by genetic selection resulted in mutant GlnRS enzymes that efficiently acylate the engineered tRNA with glutamine
in vitro
. The mutant GlnRS and engineered tRNA also constitute a functional synthetase–tRNA pair
in vivo
. The nature of the GlnRS mutations, which occur both at the protein–tRNA interface and at sites further away, is discussed.
Proceedings of the National Academy of Sciences of the United States of America - Tập 94 Số 19 - Trang 10092-10097 - 1997
Structure of pyrrolysyl-tRNA synthetase, an archaeal enzyme for genetic code innovation
Pyrrolysine (Pyl), the 22nd natural amino acid and genetically encoded by UAG, becomes attached to its cognate tRNA by pyrrolysyl-tRNA synthetase (PylRS). We have determined three crystal structures of the
Methanosarcina mazei
PylRS complexed with either AMP–PNP, Pyl–AMP plus pyrophosphate, or the Pyl analogue
N
-ε-[(cylopentyloxy)carbonyl]-
l
-lysine plus ATP. The structures reveal that PylRS utilizes a deep hydrophobic pocket for recognition of the Pyl side chain. A comparison of these structures with previously determined class II tRNA synthetase complexes illustrates that different substrate specificities derive from changes in a small number of residues that form the substrate side-chain-binding pocket. The knowledge of these structures allowed the placement of PylRS in the aminoacyl-tRNA synthetase (aaRS) tree as the last known synthetase that evolved for genetic code expansion, as well as the finding that Pyl arose before the last universal common ancestral state. The PylRS structure provides an excellent framework for designing new aaRSs with altered amino acid specificity.
Proceedings of the National Academy of Sciences of the United States of America - Tập 104 Số 27 - Trang 11268-11273 - 2007
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