Defect-Facilitated Buckling in Supercoiled Double-Helix DNA

We present a statistical-mechanical model for stretched twisted double-helix DNA, where thermal fluctuations are treated explicitly from a Hamiltonian without using any scaling hypotheses. Our model applied to defect-free supercoiled DNA describes coexistence of multiple plectoneme domains in long DNA molecules at physiological salt concentrations (≈ 0.1 M Na+) and stretching forces (≈ 1 pN). We find higher (lower) number of domains at lower (higher) ionic strengths and stretching forces, in accord with experimental observations. We use our model to study the effect of an immobile point defect on the DNA contour that allows a localized kink. The degree of the kink is controlled by the defect size, such that a larger defect further reduces the bending energy of the defect-facilitated kinked end loop. We find that a defect can spatially pin a plectoneme domain via nucleation of a kinked end loop, in accord with experiments and simulations. Our model explains previously-reported magnetic tweezer experiments [1] showing two buckling signatures: buckling and ‘rebuckling’ in supercoiled DNA with a base-unpaired region. Comparing with experiments, we find that under 1 pN force, a kinked end loop nucleated at a base-mismatched site reduces the bending energy by ≈ 0.7 kBT per unpaired base. Our model predicts coexistence of three states at the buckling and rebuckling transitions that warrants new experiments.

Molecular dissection of meiotic silencing by unpaired DNA

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT REQUEST OF AUTHOR.] Meiotic silencing by unpaired DNA (MSUD) is a genome surveillance mechanism that uses RNAi to protect against genetic parasites such as retrotransposons and viruses. This is accomplished by scanning the genome for DNA segments not paired with a homologous partner during meiosis and targeting the corresponding messenger RNAs for destruction. MSUD begins when a single-stranded RNA is made from the unpaired region and subsequently converted into small interfering RNA (siRNA), which are used as sequence specific guides for mRNA destruction. To date, nine meiotic silencing genes have been characterized and here we report the identification and characterization of two new candidates that are orthologous to the conserved proteins CBP80 and EPS15.

A Syntenic Region Conserved from Fish to Mammalian X Chromosome

Sex chromosomes bearing the sex-determining gene initiate development along the male or female pathway, no matter which sex is determined by XY male or ZW female heterogamety. Sex chromosomes originate from ancient autosomes but evolved rapidly after the acquisition of sex-determining factors which are highly divergent between species. In the heterogametic male system (XY system), the X chromosome is relatively evolutionary silent and maintains most of its ancestral genes, in contrast to its Y counterpart that has evolved rapidly and degenerated. Sex in a teleost fish, the Nile tilapia (Oreochromis niloticus), is determined genetically via an XY system, in which an unpaired region is present in the largest chromosome pair. We defined the differences in DNA contents present in this chromosome with a two-color comparative genomic hybridization (CGH) and the random amplified polymorphic DNA (RAPD) approach in XY males. We further identified a syntenic segment within this region that is well conserved in several teleosts. Through comparative genome analysis, this syntenic segment was also shown to be present in mammalian X chromosomes, suggesting a common ancestral origin of vertebrate sex chromosomes.

Triplet Analysis That Identifies Unpaired Regions of Functional RNAs

We developed a novel method for analyzing RNA sequences, deemed triplet analysis, and applied the method in anin vitroRNA selection experiment in which HIV-1 Tat was the target. Aptamers are nucleic acids that bind a desired target (bait), and to date, many aptamers have been identified byin vitroselection from enough concentrated libraries in which many RNAs had an obvious consensus primary sequence after sufficient cycles of the selection. Therefore, the higher-order structural features of the aptamers that are indispensable for interaction with the bait must be determined by additional investigation of the aptamers. In contrast, our triplet analysis enabled us to extract important information on functional primary and secondary structure from minimally concentrated RNA libraries. As a result, by using our method, an important unpaired region that is similar to the bulge of TAR was readily predicted from a partially concentrated library in which no consensus sequence was revealed by a conventional sequence analysis. Moreover, our analysis method may be used to assess a variety of structural motifs with desired function.

Suppressed Recombination and a Pairing Anomaly on the Mating-Type Chromosome of Neurospora tetrasperma

Neurospora crassa and related heterothallic ascomycetes produce eight homokaryotic self-sterile ascospores per ascus. In contrast, asci of N. tetrasperma contain four self-fertile ascospores each with nuclei of both mating types (matA and mata). The self-fertile ascospores of N. tetrasperma result from first-division segregation of mating type and nuclear spindle overlap at the second meiotic division and at a subsequent mitotic division. Recently, Merino et al. presented population-genetic evidence that crossing over is suppressed on the mating-type chromosome of N. tetrasperma, thereby preventing second-division segregation of mating type and the formation of self-sterile ascospores. The present study experimentally confirmed suppressed crossing over for a large segment of the mating-type chromosome by examining segregation of markers in crosses of wild strains. Surprisingly, our study also revealed a region on the far left arm where recombination is obligatory. In cytological studies, we demonstrated that suppressed recombination correlates with an extensive unpaired region at pachytene. Taken together, these results suggest an unpaired region adjacent to one or more paired regions, analogous to the nonpairing and pseudoautosomal regions of animal sex chromosomes. The observed pairing and obligate crossover likely reflect mechanisms to ensure chromosome disjunction.

Volume Perception with Binocular Viewing

We reported earlier that the binocularly unpaired region plays an essential role in the perception of the volume of a solid object (Idesawa, 1991 Japanese Journal of Applied Physics30-4B L751 - L754). We have investigated this volume perception using both illusory and physical objects. The illusory objects used in the tests were of two types, either with or without binocularly unpaired contours. The physical objects used as stimuli in the present experiments were half-moon-shaped textured cylindrical objects, one with and one without a binocularly unpaired region. A rectangular probe was placed beyond the object, and subjects were asked to adjust its position in depth until they felt that the probe just touched the object. We found that the chosen depth settings were systematically ‘behind’ the objects with unpaired parts, while they were almost ‘correct’ for the objects without unpaired parts. In the former case, subjects thus had the impression that the volume extended further back. We also found that the rim of an object is important for volume perception, especially for objects without textures. A typical example is an imaginary cone occluding two disks, one a little larger than the bottom surface and placed near the bottom, the other a small one located just beyond the top. A complete illusory cone with volume (just like a ‘Santa cap’) could be seen!

Volume Perception and a Processing Method of Unpaired Region in Stereo Vision

In the human visual system, binocular unpaired regions, where binocular images do not correspond to each other, play a very important role on stereo perception. In our recent experiments, we found that binocular unpaired regions give a special effect on the volume perception of solid objects with curved surfaces. In this paper, we shall introduce phenomena of volume perception, and then propose some strategies for realizing such function on a computer vision system.

Binocular Displacement of Unpaired Region

Binocular displacement of binocularly unpaired parts of the stimulus was examined by means of the Poggendorff figure. The Poggendorff figure can be used to investigate displacement since lateral displacement of the transversal may cause bias in judgments of its collinearity. In experiment 1, the transversal had a disparity, and thus binocularly unpaired parts, relative to the rectangle. The magnitude of the Poggendorff illusion should not have changed by addition of binocular disparity if displacement occurred. There was no or slight change when the transversal was seen behind the rectangle, but there was significant decrease when the transversal was seen in front of the rectangle, suggesting absence of displacement in this case. There were two possible explanations. One was that displacement depended on the positional relation between the unpaired stimuli and the binocularly presented rectangle, ie the occlusion constraint, which the case with the transversal in front did not satisfy. The alternative was that the decrease was due to the perceived front depth of the transversal, and not related to binocular displacement at all. In order to discriminate between these two possibilities, the transversal was reduced to only the unpaired parts, resulting in dichoptic stimulation in experiment 2. In this stimulus, the positional relation between the unpaired and the paired stimuli was the same as in the previous experiment, yet no front depth could be perceived. The results showed similar asymmetry as in experiment 1. Thus we conclude that binocular displacement depends on the positional relation between the unpaired and the paired stimuli, regardless of their perceived depth. This may imply that binocular displacement is not symmetric about the sign of disparity, hence that it is not just averaging but is a reconstruction of the spatial layout of objects in the outside world to keep the visual direction of the unsuppressed unpaired region veridical by using explicit cues to depth discontinuity.