Drosophila female precopulatory behavior is modulated by ecdysteroids.

The effect of ecdysteroid signaling on Drosophila female precopulatory behavior was investigated using two types of mutants with either globally reduced ecdysteroid availability or reduced expression of ecdysone receptors in fruitless neurons, known to control sexual behavior. While being courted by males, mutant females performed significantly less full ovipositor extrusion behavior to reject male copulation attempts. Ecdysteroid depleted females (ecdysoneless(1)) performed male-like courtship behaviors, including unilateral wing extension and song production with patterns very similar to male courtship song. These results support the hypothesis that ecdysteroids modulate female sexual behavior, perhaps acting as a regulator of sexual motivation, and as a component affecting the performance of sex specific behavior patterns.

Molecular methods for differentiating grey seal (Halichoerus grypus) and harbour seal (Phoca vitulina) scat.

Pinniped dietary information is crucial for understanding marine ecosystems; however, in the North Atlantic, grey and harbour seals haulout concomitantly and their faeces are visually indistinguishable. Therefore, we developed molecular methods to differentiate the species' scat. PCR primers were created that amplify a portion of mitochondrial 16S ribosomal DNA in grey and harbour seals. The samples were amplified and the resulting products were digested using the restriction enzyme Tsp509I, producing diagnostic banding patterns. These tools provide a mechanism by which separate dietary analysis can be achieved for grey and harbour seals at mixed haulouts in New England.

pkl1(+)and klp2(+): Two kinesins of the Kar3 subfamily in fission yeast perform different functions in both mitosis and meiosis.

We have identified Klp2p, a new kinesin-like protein (KLP) of the KAR3 subfamily in fission yeast. The motor domain of this protein is 61% identical and 71% similar to Pkl1p, another fission yeast KAR3 protein, yet the two enzymes are different in behavior and function. Pkl1p is nuclear throughout the cell cycle, whereas Klp2p is cytoplasmic during interphase. During mitosis Klp2p enters the nucleus where it forms about six chromatin-associated dots. In metaphase-arrested cells these migrate back and forth across the nucleus. During early anaphase they segregate with the chromosomes into two sets of about three, fade, and are replaced by other dots that form on the spindle interzone. Neither klp2(+) nor pkl1(+) is essential, and the double deletion is also wild type for both vegetative and sexual reproduction. Each deletion rescues different alleles of cut7(ts), a KLP that contributes to spindle formation and elongation. When either or both deletions are combined with a dynein deletion, vegetative growth is normal, but sexual reproduction fails: klp2 Delta,dhc1-d1 in karyogamy, pkl1 Delta,dhc1-d1 in multiple phases of meiosis, and the triple deletion in both. Deletion of Klp2p elongates a metaphase-arrested spindle, but pkl1 Delta shortens it. The anaphase spindle of klp2 Delta becomes longer than the cell, leading it to curl around the cell's ends. Apparently, Klp2p promotes spindle disassembly and contributes to the behavior of mitotic chromosomes.

Biochemical interactions within a ternary complex of the bacteriophage T4 recombination proteins uvsY and gp32 bound to single-stranded DNA.

The presynaptic phase of homologous recombination requires the formation of a filament of single-stranded DNA (ssDNA) coated with a recombinase enzyme. In bacteriophage T4, at least three proteins are required for the assembly of this presynaptic filament. In addition to the T4 recombinase, uvsX protein, the T4 ssDNA binding protein (gp32), and the uvsY recombination accessory protein are also required. Here we report on a detailed analysis of a tripartite filament containing ssDNA bound by stoichiometric quantities of both uvsY and gp32, which appears to be an important intermediate in the assembly of the T4 presynaptic filament. We demonstrate that uvsY and gp32 simultaneously co-occupy the ssDNA in a noncompetitive fashion. In addition, we show that protein-protein interactions between uvsY and gp32 are not required for the assembly of this ternary complex and do not affect the affinity of uvsY for the ssDNA lattice. Finally, we demonstrate that the interaction of gp32 with the ssDNA is destabilized within this complex, in a manner which is independent of gp32-uvsY interactions. The data suggest that the uvsY protein acts to remodel the gp32-ssDNA complex via uvsY-ssDNA interactions. The implications of these findings for the mechanism of presynapsis in the T4 recombination system are discussed.

Single-stranded DNA binding properties of the uvsY recombination protein of bacteriophage T4.

The uvsY protein is an essential component of the bacteriophage T4 general recombination machinery. The properties of this 16 kDa protein include selective binding to ssDNA, as well as specific protein-protein interactions with other T4 recombination proteins including uvsX (general recombinase) and gp32 (ssDNA-binding protein). uvsY promotes the assembly of uvsX-ssDNA filaments, the active species in uvsX-catalyzed DNA rearrangements, apparently by helping uvsX displace gp32 from the ssDNA. To better understand the role of uvsY in the T4 recombination system, here we characterize the thermodynamic and molecular properties of the interaction of the uvsY protein with a model single-stranded polynucleotide, epsilonDNA, which is a fluorescent, etheno-modified form of random-sequence ssDNA. We have found that the binding of uvsY protein enhances the fluorescence of the epsilonDNA lattice and that the maximal amount of fluorescence enhancement observed is dependent on salt concentration. In addition, we have used the epsilonDNA fluorescence enhancement assay to establish thermodynamic parameters of binding and to define some of the molecular details of uvsY-epsilonDNA interactions. We show that uvsY binds to epsilonDNA in a non-cooperative manner, with a binding site size of four nucleotide residues per monomer of uvsY, and that this binding is salt-sensitive and involves the displacement of anions from the uvsY protein. We further show that uvsY protein binds preferentially to epsilonDNA over unmodified ssDNA. The significance of these results is discussed in light of current models of uvsY action in the T4 recombination system.