U2AF binding selects for the high conservation of the C. elegans 3' splice site

Caenorhabditis elegans is unusual among animals in having a highly conserved octamer sequence at the 3' splice site: UUUU CAG/R. This sequence can bind to the essential heterodimeric splicing factor U2AF, with U2AF65 contacting the U tract and U2AF35 contacting the splice site itself (AG/R). Here we demonstrate a strong correspondence between binding to U2AF of RNA oligonucleotides with variant octamer sequences and the frequency with which such variations occur in splice sites. C. elegans U2AF has a strong preference for the octamer sequence and exerts much of the pressure for 3' splice sites to have the precise UUUUCAG/R sequence. At two positions the splice site has a very strong preference for U even though alternative bases can also bind tightly to U2AF, suggesting that evolution can select against sequences that may have a relatively modest reduction in binding. Although pyrimidines are frequently present at the first base in the exon, U2AF has a very strong bias against them, arguing there is a mechanism to compensate for weakened U2AF binding at this position. Finally, the C in the consensus sequence must remain adjacent to the AG/R rather than to the stretch of U's, suggesting this C is recognized by U2AF35.     

Investigating the Effect of Chain Connectivity on the Folding of a Beta-Sheet Protein On and Off the Ribosome

Determining the relationship between protein folding pathways on and off the ribosome remains an important area of investigation in biology. Studies on isolated domains have shown that alteration of the separation of residues in a polypeptide chain, while maintaining their spatial contacts, may affect protein stability and folding pathway. Due to the vectorial emergence of the polypeptide chain from the ribosome, chain connectivity may have an important influence upon cotranslational folding. Using MATH, an all β-sandwich domain, we investigate whether the connectivity of residues and secondary structure elements is a key determinant of when cotranslational folding can occur on the ribosome. From Φ-value analysis, we show that the most structured region of the transition state for folding in MATH includes the N and C terminal strands, which are located adjacent to each other in the structure. However, arrest peptide force-profile assays show that wild-type MATH is able to fold cotranslationally, while some C-terminal residues remain sequestered in the ribosome, even when destabilized by 2–3?kcal?mol^?1. We show that, while this pattern of Φ-values is retained in two circular permutants in our studies of the isolated domains, one of these permutants can fold only when fully emerged from the ribosome. We propose that in the case of MATH, onset of cotranslational folding is determined by the ability to form a sufficiently stable folding nucleus involving both β-sheets, rather than by the location of the terminal strands in the ribosome tunnel.     

Preparation of soybean seed samples for FT-IR microspectroscopy

Typical preparation of seed samples for infrared (IR) microspectroscopy involves imbibition of the seed for varying time periods followed by cryosectioning. Imbibition, however, may initiate germination even at 4° C with associated changes in the chemistry of the sample. We have found that it is possible to section seeds that are sufficiently hard, such as soybeans, on a standard laboratory microtome without imbibition. The use of dry sectioning of unimbibed seeds is reported here, as well as a comparison of different mounting media and modes of analysis. Glycerol, Tissue-Tek, and ethanol were used as mounting media, and the quality of the resulting spectra was assessed. Ethanol was the preferred mountant, because it dried quickly with no residue and thus did not interfere with the spectrum of interest. Analysis in transmission mode using barium fluoride windows to hold the samples was compared with transmission-reflection analysis with sections mounted on special infrared-reflecting slides. The two modes of analysis performed well in different regions of the spectrum. The mode of analysis (transmission vs. transmission-reflection) should be based on the components of greatest interest in the sample.     

Shoal size as a key determinant of vulnerability to capture under a simulated fishery scenario

Group living is widespread among animals and has a range of positive effects on individual foraging and predator avoidance. For fishes, capture by humans constitutes a major source of mortality, and the ecological effects of group living could carry‐over to harvest scenarios if fish are more likely to interact with fishing gears when in social groups. Furthermore, individual metabolic rate can affect both foraging requirements and social behaviors, and could, therefore, have an additional influence on which fish are most vulnerable to capture by fishing. Here, we studied whether social environment (i.e., social group size) and metabolic rate exert independent or interactive effects on the vulnerability of wild zebrafish (Danio rerio) to capture by a baited passive trap gear. Using video analysis, we observed the tendency for individual fish to enter a deployed trap when in different shoal sizes. Fish in larger groups were more vulnerable to capture than fish tested individually or at smaller group sizes. Specifically, focal fish in larger groups entered traps sooner, spent more total time within the trap, and were more likely to re‐enter the trap after an escape. Contrary to expectations, there was evidence that fish with a higher SMR took longer to enter traps, possibly due to a reduced tendency to follow groupmates or attraction to conspecifics already within the trap. Overall, however, social influences appeared to largely overwhelm any link between vulnerability and metabolic rate. The results suggest that group behavior, which in a natural predation setting is beneficial for avoiding predators, could be maladaptive under a trap harvest scenario and be an important mediator of which traits are under harvest associated selection.     

Time course and action spectrum of vibrotactile adaptation

In a series of experiments designed to explore the processes underlying adaptation of the sense of flutter-vibration, vibrotactile threshold was measured on the pad of the index finger, using Békésy tracking. Unadapted thresholds were first measured, for a number of frequencies (4-90 Hz) and contactor sizes (1-8 mm diameter). As expected, these measurements indicated the presence of (1) a Pacinian system possessing spatial summation and increasing in sensitivity, as frequency was raised, at the rate of 12 dB/octave; and (2) a non-Pacinian system showing little spatial summation, and with a frequency characteristic matching that of the NP I mechanism of Bolanowski et al. (1988). These baseline data of Experiment 1 guided the selection of stimulus parameters for subsequent experiments, in which threshold for a test stimulus was measured before, during, and after periods of vibrotactile adaptation. In Experiment 2, test stimuli of 10 Hz and 50 Hz were combined factorially with 30-dB SL adapting stimuli of the same two frequencies. When the test stimulus was 10 Hz, the two adapting frequencies were equally effective in raising threshold; however, when the 50-Hz test stimulus was used, the 50-Hz adapting stimulus raised threshold by a greater amount than did the 10-Hz adapter. These results confirm on the finger the independence of adaptation in Pacinian and non-Pacinian channels, a result previously established on the thenar by other workers. For all four frequency combinations, threshold rose exponentially with a time constant of 1.5-2 min. In Experiment 3, an action spectrum was determined, showing the adapting amplitude needed at each of a series of frequencies to raise the threshold of a 10-Hz stimulus by 10 dB; this spectrum was essentially flat from 30 to 90 Hz. The results, taken in conjunction with what is known about rapidly adapting cutaneous mechanoreceptors, imply that the effectiveness of an adapting stimulus is not determined solely by the amount of activity it generates in first-order afferents.     

Ambulation in labour.

In a randomised prospective study of 68 women in spontaneous labour half were allocated to an ambulant group and half to a recumbent group. The duration of labour was significantly shorter, the need for analgesia significantly less, and the incidence of fetal heart abnormalities significantly smaller in the ambulant group than in the recumbent group. Apgar scores at one and five minutes were also significantly greater in the ambulant group. More patients in the recumbent group required augmentations with oxytocic drugs. There was no statistically significant difference in the third stage loss in the two groups. Ambulation in labour should be encouraged: it may bring human benefits while allowing the advantages of hospital supervision.     

Calls,colours, shape,and genes: a multi‐trait approach to the study of geographic variation in the Amazonian frog Allobates femoralis

Evolutionary divergence in behavioural traits related to mating may represent the initial stage of speciation. Direct selective forces are usually invoked to explain divergence in mate‐recognition traits, often neglecting a role for neutral processes or concomitant differentiation in ecological traits. We adopted a multi‐trait approach to obtain a deeper understanding of the mechanisms behind allopatric divergence in the Amazonian frog, Allobates femoralis. We tested the null hypothesis that geographic distance between populations correlates with genetic and phenotypic divergence, and compared divergence between mate‐recognition (acoustic) and ecological (coloration, body‐shape) traits. We quantified geographic variation in 39 phenotypic traits and a mitochondrial DNA marker among 125 individuals representing eight populations. Geographic variation in acoustic traits was pronounced and tracked the spatial genetic variation, which appeared to be neutral. Thus, the evolution of acoustic traits tracked the shared history of the populations, which is unexpected for pan‐Amazonian taxa or for mate‐recognition traits. Divergence in coloration appeared uncorrelated with genetic distance, and might be partly attributed to local selective pressures, and perhaps to Batesian mimicry. Divergence in body‐shape traits was low. The results obtained depict a complex evolutionary scenario and emphasize the importance of considering multiple traits when disentangling the forces behind allopatric divergence. ©2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 98 , 826–838.