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Although sperm serine protease and proteasome have long been believed to play an important role in the fertilization process, the molecular mechanism is still controversial. In this study, we have produced double-knockout mice lacking two sperm serine proteases, ACR and PRSS21, to uncover the functional role of the trypsinlike activity in fertilization. The double-knockout male mice were subfertile, likely owing to the incompleteness of fertilization in the oviductal ampulla. Despite male subfertility, the mutant epididymal sperm exhibited the inability to undergo acrosomal exocytosis on the zona pellucida (ZP) surface and to traverse the ZP, thus resulting in the failure of fertilization in vitro. The double-knockout epididymal sperm were also defective in penetration through the cumulus matrix to reach the ZP. When epididymal sperm were artificially injected into the uterus of wild-type mice, the 2-cell embryos, which had previously been fertilized by double-knockout sperm, were recovered at a low but significant level. The mutant epididymal sperm were also capable of fertilizing the oocytes in the presence of uterine fluids in vitro. These data demonstrate that the trypsinlike protease activity of ACR and PRSS21 is essential for the process of sperm penetration through the cumulus matrix and ZP in vitro, and suggest that the female reproductive tract partially compensates for the loss of the sperm function. We therefore conclude that the sperm trypsinlike activity is still important but not essential for fertilization in vivo in the mouse.

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Cells must coordinate adjustments in genome expression to accommodate changes in their environment. We hypothesized that the amount of transcriptome change is proportional to the amount of environmental change. To capture the effects of environmental changes on the transcriptome, we compared transcriptome diversities (defined as the Shannon entropy of frequency distribution) of silkworm fat-body tissues cultured with several concentrations of phenobarbital. Although there was no proportional relationship, we did identify a drug concentration “tipping point” between 0.25 and 1.0 mM. Cells cultured in media containing lower drug concentrations than the tipping point showed uniformly high transcriptome diversities, while those cultured at higher drug concentrations than the tipping point showed uniformly low transcriptome diversities. The plasticity of transcriptome diversity was corroborated by cultivations of fat bodies in MGM-450 insect medium without phenobarbital and in 0.25 mM phenobarbital-supplemented MGM-450 insect medium after previous cultivation (cultivation for 80 hours in MGM-450 insect medium without phenobarbital, followed by cultivation for 10 hours in 1.0 mM phenobarbital-supplemented MGM-450 insect medium). Interestingly, the transcriptome diversities of cells cultured in media containing 0.25 mM phenobarbital after previous cultivation (cultivation for 80 hours in MGM-450 insect medium without phenobarbital, followed by cultivation for 10 hours in 1.0 mM phenobarbital-supplemented MGM-450 insect medium) were different from cells cultured in media containing 0.25 mM phenobarbital after previous cultivation (cultivation for 80 hours in MGM-450 insect medium without phenobarbital). This hysteretic phenomenon of transcriptome diversities indicates multi-stability of the genome expression system. Cellular memories were recorded in genome expression networks as in DNA/histone modifications.

The amounts of simian virus 40 structural polypeptides Vpl, Vp2, and Vp3 in different subcellular fractions at various times after lytic infection were determined by a quantitative immunoblotting procedure. Simian virus 40-infected cells were lysed with a buffer containing Nonidet P-40 to yield a soluble fraction. The Nonidet P-40-insoluble fraction was further fractionated in the presence of deoxycholate and Tween 40 to yield a soluble fraction (cytoskeletal) and an insoluble fraction (Nuc), which is primarily cell nuclei. At 33 h postinfection, the majority of viral structural proteins was found in the cell nucleus, whereas, at 48 to 65 h postinfection, Vpl was distributed evenly among all cell fractions and Vp2 and Vp3 were found predominantly in the cytoskeletal and Nuc fractions. Thus, not all of the viral polypeptides synthesized in the cytoplasm migrated into the cell nucleus. Throughout infection, the molar ratio (Vp3/Vp2) was rather constant in all subcellular fractions, indicating that the synthesis or processing or both of Vp2 and Vp3 are coordinately regulated. The molar ratio of Vpl/(Vp2 + Vp3) varied among the fractions. The Vpl/(Vp2 + Vp3) molar ratio in the soluble fraction varied during the course of infection; however, constant ratios were maintained in the cytoskeletal and Nuc fractions. Thus, the mechanism which controls the movement of Vpl to different compartments of the cell appears to be different from that of Vp2 and Vp3. The Vpl/(Vp2 + Vp3) value in the Nuc fraction was similar to the ratio found in virus particles. The constant molar distribution of Vpl, Vp2, and Vp3 in the Nuc fraction throughout infection suggests that there is a specific mechanism which regulates the transport of viral structural proteins. These results support the hypothesis that the structural proteins of simian virus 40 are transported into the cell nucleus in precise proportions.

Many butterflies acquire nutrients from non-nectar sources such as puddles. To better understand how male Papilio butterflies identify suitable sites for puddling, we used behavioral and electrophysiological methods to examine the responses of Japanese Papilio butterflies to Na+, K+, Ca2+, and Mg2+. Based on behavioral analyses, these butterflies preferred a 10-mM Na+ solution to K+, Ca2+, and Mg2+ solutions of the same concentration and among a tested range of 1 mM to 1 M NaCl. We also measured the ion concentrations of solutions sampled from puddling sites in the field. Na+ concentrations of the samples were up to 6 mM, slightly lower than that preferred by butterflies in the behavioral experiments. Butterflies that sipped the 10 mM Na+ solution from the experimental trays did not continue to puddle on the ground. Additionally, butterflies puddled at sites where the concentrations of K+, Ca2+, and/or Mg2+ were higher than that of Na+. This suggests that K+, Ca2+, and Mg2+ do not interfere with the detection of Na+ by the Papilio butterfly. Using an electrophysiological method, tip recordings, receptor neurons in contact chemosensilla inside the proboscis evoked regularly firing impulses to 1, 10, and 100 mM NaCl solutions but not to CaCl2 or MgCl2. The dose–response patterns to the NaCl solutions were different among the neurons, which were classified into three types. These results showed that Japanese Papilio butterflies puddle using Na+ detected by the contact chemosensilla in the proboscis, which measure its concentration.Electronic supplementary materialThe online version of this article (doi:10.1007/s00114-012-0976-3) contains supplementary material, which is available to authorized users.

We studied damage to wool fabrics dyed with different natural and chemical dyestuffs by the larvae of varied carpet beetle, Anthrenus verbasci, as part of a study on the functions of natural dyestuffs. Eight of ten natural dyestuffs showed an antifeeding effect against A. verbasci. Strength of the antifeeding effect of natural dyestuffs in a feeding preference test was in the order lac dye, gallnut, catechu, red cabbage, Cricula cocoon extract > cochineal, indigo, Amur cork tree extract > chemical dye. Lithospermi radix and turmeric were less effective against A. verbasci. The damage to dyed fabrics by the insect was not related to the extent of color depth or shade of the dyed fabric. Water-soluble substances having absorption peaks at around 280 nm, commonly were present in the natural dyestuffs except for turmeric. The polyphenols, tannic acid and catechin having absorption peaks at around 280 nm, seemed to be not related with the antifeeding effect. An alkali degummed Cricula cocoon sample that had almost all of the cocoon filament sericin removed showed almost the same level of damage by insect feeding as that of natural Cricula cocoons. Discipline: Insect pestAdditional key words: fabric insect pest, extracts of natural pigment, feeding test, feeding preference test, absorption spectra

Eggs of the migratory locust, Locusta migratoria (Orthoptera: Acrididae), hatch synchronously when in a pod, but only sporadically when kept separately. Here, we aimed to detect the vibrational stimuli emitted by eggs that initiate synchronous hatching. First, we recorded the vibrations emitted by an egg pod and single eggs. One bout of vibrations consisted of 2 to 46 vibrations. The total number and amplitude of vibrations in single eggs increased as the time to hatch decreased. Eggs kept separately were continuously subjected during the last 2 days before hatching to recordings of vibrations from a single egg. Recordings made during the last 2.5 h before hatching caused these eggs to hatch signifi cantly earlier than those not subjected to this treatment, the control. In contrast, eggs subjected to recordings made 8 to 10 h before hatching signifi cantly delayed their hatching relative to the controls, which indicates that synchronous hatching of eggs is induced by age-dependent changes in vibrations from neighbouring eggs. Exposure to one large bout of vibrations (consisting of 40 vibrations in 101 s) was suffi cient to induce synchronous hatching in eggs kept separately when applied 5 h before hatching, but not 36 or 11.5 h before hatching. Visual inspection of the spectra indicated that the vibrations had two peaks at about 100 Hz and 1.5 kHz. Only exposure to the latter altered the hatching time of eggs. The embryo moved the posterior part of its abdomen when emitting the vibrations. These results indicate that the synchronous hatching of locust eggs is induced when the embryos emit particular vibrations.