DNA hybridization and evolutionary relationships in three osmunda species.

Molecular hybridization techniques have been used to estimate the degree of DNA base sequence homology between some members of the fern genus Osmunda. Under conditions permitting extensive reassociation, measurements of the extent of inter-specific reaction and the thermal stability of the hybrid molecules indicated that O. claytoniana L. (interrupted fern) shares more DNA homology with O. cinnamomea L. (cinnamon fern) than it does with O. regalis L. (royal fern). These findings are in conflict with predictions from a recent analysis of living and fossil specimens by numerical techniques. However, they are consistent with the earlier, more traditional, taxonomic assignments.

Different Red Light Requirements for Phytochrome-Induced Accumulation of cab RNA and rbcS RNA.

For several species of plants the abundance of those transcripts encoding the chlorophyll a/b binding protein (cab RNA) and the small subunit of ribulose-1,5-biphosphate carboxylase-oxygenase (rbcS RNA) has been established as being under the control of phytochrome. However, this conclusion does not take into account the various types of phytochrome control based on both the fluence of red light necessary to induce the response and the ability of far red light either to induce or to reverse the response. The fluence of red light necessary to induce the accumulation of rbcS RNA was found to be 10,000 times greater than that necessary to induce the accumulation of cab RNA. Furthermore, far red light alone was capable of inducing the accumulation of cab RNA. It is possible, therefore, that developing pea buds accumulate cab RNA before rbcS and that cab RNA is not subject to the normal end-of-day signals affecting many phytochrome responses.

Decoding the dynamic representation of musical pitch from human brain activity.

In music, the perception of pitch is governed largely by its tonal function given the preceding harmonic structure of the music. While behavioral research has advanced our understanding of the perceptual representation of musical pitch, relatively little is known about its representational structure in the brain. Using Magnetoencephalography (MEG), we recorded evoked neural responses to different tones presented within a tonal context. Multivariate Pattern Analysis (MVPA) was applied to "decode" the stimulus that listeners heard based on the underlying neural activity. We then characterized the structure of the brain's representation using decoding accuracy as a proxy for representational distance, and compared this structure to several well established perceptual and acoustic models. The observed neural representation was best accounted for by a model based on the Standard Tonal Hierarchy, whereby differences in the neural encoding of musical pitches correspond to their differences in perceived stability. By confirming that perceptual differences honor those in the underlying neuronal population coding, our results provide a crucial link in understanding the cognitive foundations of musical pitch across psychological and neural domains.

cis-Acting Elements for Light Regulation of Pea Ferredoxin I Gene Expression Are Located within Transcribed Sequences.

An intact pea gene encoding ferredoxin I (Fed-1) and several chimeric constructs containing portions of Fed-1 were introduced into tobacco plants by Agrobacterium-mediated transformation. The intact gene was correctly transcribed and translated to produce a protein that was imported into the chloroplast and processed to its mature size. Fed-1 mRNA accumulation in these plants was strongly light-dependent, as it is in pea leaves. In chimeric constructs, the Fed-1 promoter was active but no light responses were seen, even when as much as 2 kilobases of 5[prime] -flanking sequence were included. We also failed to observe clear light responses with a construct containing 3[prime] -flanking sequences from Fed-1 attached to a [beta]-glucuronidase gene driven by the cauliflower mosaic virus 35S promoter. However, the transcribed portion of Fed-1 conveyed normal light responsiveness when driven by the 35S promoter. The results are discussed in terms of the hypothesis that light determines Fed-1 mRNA abundance by affecting RNA stability rather than by affecting transcription.

Characterization of Post-Transcriptionally Suppressed Transgene Expression That Confers Resistance to Tobacco Etch Virus Infection in Tobacco.

Tobacco lines expressing transgenes that encode tobacco etch virus (TEV) coat protein (CP) mRNA with or without nonsense codons give rise to TEV-resistant tissues that have reduced levels of TEV CP mRNA while maintaining high levels of transgene transcriptional activity. Two phenotypes for virus resistance in the lines containing the transgene have been described: immune (no virus infection) and recovery (initial systemic symptoms followed by gradual recovery over several weeks). Here, we show that at early times in development, immune lines are susceptible to TEV infection and accumulate full-length CP mRNA. Therefore, immune lines also exhibit meiotic resetting, as is seen in the recovery lines, providing molecular evidence for a common mechanism of gene silencing and virus resistance in both cases. We also investigated the characteristics of two sets of low molecular weight RNAs that appear only in silenced tissue. One set has nearly intact 5[prime] ends, lacks poly(A) tails, and is associated with polyribosomes; the second set contains the 3[prime] end of the mRNA. Treating silenced leaf tissue with cycloheximide resulted in decreased levels of full-length mRNA and an increase in the levels of the low molecular weight RNAs, supporting a cytoplasmic decay mechanism that does not require ongoing translation. Surprisingly, mRNA from the transgene containing nonsense codons was associated with more ribosomes than expected, possibly resulting from translation from a start codon downstream of the introduced translational stop codons. We present a hypothesis for transgene/viral RNA degradation in which RNA degradation occurs in the cytoplasm while in association with polyribosomes.

Sequence of Lhcb3*1, a gene encoding a Photosystem II chlorophyll a/b-binding protein in Pisum.

We have cloned and sequenced a pea Lhcb3 gene, encoding a Photosystem II chlorophyll a/b-binding protein. Sequence analysis indicates that the gene contains two introns and predicts a polypeptide of 265 amino acids. The predicted polypeptide sequence is highly homologous to the polypeptide sequences deduced from Lhcb3 genes previously characterized in tomato and barley.

Secretion of intrinsic factor from dispersed mucosal cells isolated from guinea pig and rabbit stomach.

In dispersed mucosal cells prepared from rabbit and guinea pig stomach, the secretion of intrinsic factor was constant (0.3-0.4%/min) for at least 30 min incubation at 37 degrees C. Histamine or isobutyl methylxanthine increased cyclic AMP and intrinsic factor secretion in both cells preparations. Isobutyl methylxanthine potentiated and cimetidine competitively inhibited (ki = 5.10-7 M) both effects of histamine. Dibutyryl cyclic AMP (1.0 mM), also caused a 3-fold increase in intrinsic factor secretion. These results suggest that in rabbit and guinea pig histamine interacts with H2-receptors to increase cyclic AMP which mediates the rise in the rate of intrinsic factor secretion.

DNase I sensitivity of ribosomal RNA genes in chromatin and nucleolar dominance in wheat.

Ribosomal RNA genes at different nucleolar organizer (NOR) loci in hexaploid wheat are expressed at different levels. The degree of expression of a particular organizer depends on the genetic background, especially on the presence of other NOR loci. For example, when chromosome 1U of Aegilops umbellulata is introduced into the hexaploid wheat cultivar "Chinese Spring" the A. umbellulata NOR accounts for most of the nucleolar activity and seems to suppress the activity of the wheat NOR loci. Even in wild-type "Chinese Spring", the NOR on chromosome 1B is partially dominant to that on chromosome 6B, since the 1B locus is more active in spite of having fewer genes. We have previously shown that these and other examples of nucleolar dominance in wheat are associated with undermethylation of cytosine residues in certain regions of the dominant rDNA. Here, we show that rRNA genes at dominant loci are organized in a chromatin conformation that renders them more sensitive to DNase I digestion than other rRNA genes. In addition, we have mapped several DNase I-hypersensitive sites in the intergenic spacer region of the rDNA repeating unit. Some of these sites are located near the initiation region for the 45 S rRNA precursor, while others are associated with a series of short direct repeats 5' to the 45 S rRNA initiation site. The results are discussed in terms of a model in which repeated sequences in the wheat intergenic DNA are presumed to function as upstream promoters and transcriptional enhancers similar to those in Xenopus.

Regulation of cytosine methylation in ribosomal DNA and nucleolus organizer expression in wheat.

Cytosine methylation has been studied in wheat rRNA genes at nucleolar organizers displaying different activities. The methylation pattern within a specific multigene locus is influenced by the number and type of rRNA genes in other rDNA loci in the cell. One CCGG site 164 base-pairs upstream from the start of transcription is preferentially unmethylated in some genes. Dominant, very active loci have a higher proportion of rRNA genes with unmethylated cytosine residues in comparison with recessive and inactive loci. It is concluded that cytosine methylation in rDNA is regulated and that the methylation pattern correlates with the transcription potential of an rRNA gene.

Developmental regulation of cytosine methylation in the nuclear ribosomal RNA genes of Pisum sativum.

Prominent features of the cytosine methylation pattern of the Pisum sativum nuclear ribosomal RNA genes have been defined. Cytosine methylation within the C-C-G-G sequence was studied using the restriction enzymes HpaII and MspI and gel blot hybridizations of the restriction digests. The extent to which particular features of the methylation pattern change during seedling development has also been determined. Total cellular DNA, purified from defined sections of pea seedlings grown under different lighting conditions, was analyzed with DNA hybridization probes derived from different portions of a cloned member of the nuclear rRNA gene family. By use of an indirect end-labeling technique, a map of 23 cleavable HpaII and/or MspI sites in genomic rDNA was constructed. The map covers about 90% of the rDNA repeat including the entire non-transcribed spacer region and most of the rRNA coding sequences. One notable feature of the map is that the most prominent HpaII site, located about 800 base-pairs upstream from the 5' end of the mature 18 S rRNA, is cleaved only in one of the two most abundant rDNA length variants (the short variant). With a gel blot assay specific for cleavage at this site, we estimated the HpaII sensitivity of DNA preparations from several stages of pea seedling development. We find that, while methylation is generally low in young seedlings, DNA obtained from the apical buds of pea seedlings is highly methylated. Further, the methylation level of rDNA within the pea bud decreases as the buds are allowed to develop under continuous white light. Our data, taken together with published studies on pea seedling development, indicate that cytosine methylation levels may be related to the regulated expression of the nuclear rRNA genes in pea.

Chloroplast DNA variation and evolution in pisum: patterns of change and phylogenetic analysis.

Variation in 30 chloroplast DNAs, representing 22 wild and cultivated accessions in the genus Pisum, was analyzed by comparing fragment patterns produced by 16 restriction endonucleases. Three types of mutations were detected. First, an inversion of between 2.2 kilobase pairs (kb) and 5.2 kb distinguished a population of P. humile from all other Pisum accessions examined. Second, deletions and insertions of between 50 and 1200 base pairs produced small restriction fragment length variations in four regions of the 120-kb chloroplast genome. Two of these regions-one of which is located within the sequence that is inverted in P. humile-showed a high degree of size polymorphism, to the extent that size differences were detected between individuals from the same accession. Finally, a total of only 11 restriction site mutations were detected among the 165 restriction sites sampled in the 30 DNAs. Based on these results and previous data, we conclude that the chloroplast genome is evolving very slowly relative to nuclear and mitochondrial DNAs. The Pisum chloroplast DNA restriction site mutations define two major lineages: One includes all tested accessions of P. fulvum, which is known to be cytogenetically quite distinct from all other Pisum taxa. The second includes 12 of 13 cultivated lines of the garden pea (P. sativum) and a wild population of P. humile from northern Israel. These observations strongly reinforce an earlier conclusion that the cultivated pea was domesticated primarily from northern populations of P. humile. A 13th P. sativum cultivar has a chloroplast genome that is significantly different from those of the aforementioned lines and somewhat more similar to those of P. elatius and southern populations of P. humile. This observation indicates that secondary hybridization may have occurred during the domestication of the garden pea.

Transposon tagging of the sulfur gene of tobacco using engineered maize Ac/Ds elements.

The Sulfur gene of tobacco is nuclearly encoded. A Su allele at this locus acts as a dominant semilethal mutation and causes reduced accumulation of chlorophyll, resulting in a yellow color in the plant. An engineered transposon tagging system, based upon the maize element Ac/Ds, was used to mutate the gene. High frequency of transposon excision from the Su locus produced variegated sectors. Plants regenerated from the variegated sector exhibited a similar variegated phenotype. Genetic analyses showed that the variegation was always associated with the transposase construct and the transposon was linked to the Su locus. Sequences surrounding the transposon were isolated, and five revertant sectors possessed typical direct repeats following Ds excisions. These genetic and molecular data are consistent with the tagging of the Su allele by the transposon.

Individual Members of the Cab Gene Family Differ Widely in Fluence Response.

Chlorophyll a/b-binding protein genes (Cab genes) can be extremely sensitive to light. Transcript accumulation following a red light pulse increases with fluence over 8 orders of magnitude (L.S. Kaufman, W.F. Thompson, W.R. Briggs [1984] Science 226: 1447-1449). We have constructed fluence-response curves for individual Cab genes. At least two Cab genes (Cab-8 and AB96) show a very low fluence response to a single red light pulse. In contrast, two other Cab genes (AB80 and AB66) fail to produce detectable transcript following a single pulse of either red or blue light but are expressed in continuous red light. Thus, very low fluence responses and high irradiance responses occur in the same gene family.

Clone banks of the mung bean, pea and spinach chloroplast genomes.

All but one of the PstI restriction fragments from mung bean, pea, and spinach chloroplast DNAs have been stably cloned into pBR322. Large fragments (15-54 kb) were cloned at low efficiencies which decreased with increasing fragment length. However, plasmids containing fragments above 25-30 kb were too unstable to be useful. In particular, pBR322 derivatives containing the largest mung bean and spinach fragments (34 kb and 54 kb, respectively) are extremely unstable and rapidly delete parts of the plasmid sequence. The PstI fragments of mung bean chloroplast DNA which cover the 34-kb PstI fragment have been cloned into pACYC177. After a search of several thousand recombinants we were unable to recover a clone containing a 12.2-kb pea chloroplast PstI fragment and suggest that some property of its sequence may be inimical to the cloning process. The identity of the cloned fragments to native chloroplast DNA restriction fragments is demonstrated by restriction analysis and the ability to construct detailed restriction maps of the mung bean and pea chloroplast genomes.

Cell Survival in the Uncrossed Projection of the Mammalian Retina is Independent of Birthdate.

Naturally occurring cell loss in the retinal ganglion cell population of one eye can be interrupted by removal of the other eye in newborn rodents. Many of the rescued retinal ganglion cells which project ipsilaterally reside in the nasal retina, that part of the retina normally giving rise to primarily crossed optic axons. Their naturally occurring elimination has been attributed to their hypothesized late neurogenesis and the consequent delayed time of arrival of their axons in the target visual nuclei, thereby placing them at a competitive disadvantage with other, early arriving, optic axons. By combining the technique of tritiated thymidine autoradiography with the retrograde axonal transport of horseradish peroxidase in rats that had been enucleated on the day of birth, we report here that rescued cells in the nasal retina which project ipsilaterally are generated at the same time as their neighbours in the temporal retina. Time of genesis does not distinguish them; consequently, their axons should not differ in their arrival times within the target visual nuclei. Since their only obvious anomaly is one of pathway choice at the optic chiasm, their place of arrival, rather than their time, may ultimately determine their naturally occurring elimination.

Birthdates of neurons in the retinal ganglion cell layer of the ferret.

The present study determined the temporal and spatial patterns of genesis for neurons of different sizes in the retinal ganglion cell layer of the ferret. Fetal ferrets were exposed to tritiated thymidine on embryonic days E-22 through E-36. One to 3 months after birth, they were perfused and their retinae dissected, and autoradiographs were prepared from resin-embedded sections throughout the entire flattened retinal ganglion cell layer. Soma size differences in conjunction with separate retrograde labeling and calbindin immunocytochemical studies were used as criteria for identifying different retinal ganglion cell subtypes in juvenile and adult ferrets. Neurons of different sizes in the ganglion cell layer were generated at different stages during development. Medium sized cells were generated primarily between E-22 and E-26; the largest cells were generated between E-24 and E-29; small cells were generated between E-26 and E-32; and very small cells were generated between E-29 and E-36. The former three groups were interpreted to be three subtypes of retinal ganglion cells, while the latter group was interpreted to be displaced amacrine cells. This temporal order of the genesis of ganglion cell classes is consistent with the spatial ordering of their fibers in the mature optic chiasm and tract, and it is consistent with the developmental change in decussation pattern recently shown in the optic pathway of embryonic ferrets. The spatial pattern of genesis suggests that ganglion cells of a particular class are added to the ganglion cell layer in a centroperipheral fashion initiated in the dorsocentral retina nasal to the area centralis. No evidence was found for a wave of ganglion cell addition that proceeded in a spiralling pattern around the area centralis, as has been reported in the cat.

Music performance and the perception of key.

The effect of music performance on perceived key movement was examined. Listeners judged key movement in sequences presented without performance expression (mechanical) in Experiment 1 and with performance expression in Experiment 2. Modulation distance varied. Judgments corresponded to predictions based on the cycle of fifths and toroidal models of key relatedness, with the highest correspondence for performed versions with the toroidal model. In Experiment 3, listeners compared mechanical sequences with either performed sequences or modifications of performed sequences. Modifications preserved expressive differences between chords, but not between voices. Predictions from Experiments 1 and 2 held only for performed sequences, suggesting that differences between voices are informative of key movement. Experiment 4 confirmed that modifications did not disrupt musicality. Analyses of performances further suggested a link between performance expression and key.

Illusory conjunctions of pitch and duration in unfamiliar tone sequences.

In 3 experiments, the authors examined short-term memory for pitch and duration in unfamiliar tone sequences. Participants were presented a target sequence consisting of 2 tones (Experiment 1) or 7 tones (Experiments 2 and 3) and then a probe tone. Participants indicated whether the probe tone matched 1 of the target tones in both pitch and duration. Error rates were relatively low if the probe tone matched 1 of the target tones or if it differed from target tones in pitch, duration, or both. Error rates were remarkably high, however, if the probe tone combined the pitch of 1 target tone with the duration of a different target tone. The results suggest that illusory conjunctions of these dimensions frequently occur. A mathematical model is presented that accounts for the relative contribution of pitch errors, duration errors, and illusory conjunctions of pitch and duration.

Post-transcriptional light regulation of nuclear-encoded genes.

A significant number of studies have detected a post-transcriptional component in the light responses of nuclear genes. As yet there are few in-depth studies of the mechanism(s) involved, and it seems likely some additional examples have been missed. For instance, transcriptional responses have sometimes been inferred on the basis of experiments with translational fusions containing both the promoter and 5' UTR of the test gene, but we now know that elements within the 5' UTR can mediate post-transcriptional light responses. Similarly, because of possible changes in translation rates and protein turnover, the common assumption that mRNA levels directly dictate protein levels is tenuous at best. It is no longer permissible to assume that the biological effect of a gene is a simple function of its transcription. Thus it is likely that with careful experimental design, reports of nuclear-encoded post-transcriptional gene regulation will become increasingly prevalent.

Occult spinal dysraphism. Case report and review of the literature.

This case report and review of the literature is presented to create a greater diagnostic awareness of occult spinal dysraphism. Early recognition is based upon an understanding of this congenital anomaly and its variable presentations. These most commonly include abnormal gait, various cutaneous manifestations, particularly subcutaneous lipomata, and less frequently urological complaints. Surgical intervention, to arrest disease progression, is the primary mode of treatment, and functional improvement is variable. Long-term prognosis is dependent upon severity of neurologic deficits prior to surgery and the type of lesion found intraoperatively. Familial occurrence has been reported and genetic counseling may be an important preventive measure. Recent radiologic investigations have been concerned with the use of ultrasonography in screening infants at risk.