Enhancement and phototransduction in the ventral eye of limulus

Limulus ventral photoreceptors were voltage clamped to the resting (dark) potential and stimulated by a 20-ms test flash and a 1-s conditioning flash. At a constant level of adaptation, we measured the response to the test flash given in the dark (control) and the incremental response produced when the test flash occurred within the duration of the conditioning flash. The incremental response is defined as the response to the conditioning and test flashes minus the response to the conditioning flash given alone. When the test flash was presented within 100 ms after the onset of the conditioning flash we observed that: (a) for dim conditioning flashes the incremental response equaled the control response; (b) for intermediate intensity conditioning flashes the incremental response was greater than the control response (we refer to this as enhancement); (c) for high intensity conditioning flashes the incremental response nearly equaled the control response. Using 10-μm diam spots of illumnination, we stimulated two spatially separate regions of one photoreceptor. When the test flash and the conditioning flash were presented to the same region, enhancement was present; but when the flashes were applied to separate regions, enhancement was nearly absent. This result indicates that enhancement is localized to the region of illumination. We discuss mechanisms that may account for enhancement.     

Diversity of a major repetitive DNA sequence in diploid and polyploid Triticeae

About 90 members of a major tandemly repeated DNA sequence family originally described in rye as pSc119.2 have been isolated from 11 diploid and polyploid Triticeae species using primers from along the length of the sequence for PCR amplification. Alignment and similarity analysis showed that the 120-bp repeat unit family is diverse with single nucleotide changes and few insertions and deletions occurring throughout the sequence, with no characteristic genome or species-specific variants having developed during evolution of the extant genomes. Fluorescent in situ hybridization showed that each of the large blocks of the repeat at chromosomal sites harboured many variants of the 120-bp repeat. There were substantial copy number differences between genomes, with abundant sub-terminal sites in rye, interstitial sites in the B genome of wheat, and relatively few sites in the A and D genome. We conclude that sequence homogenization events have not been operative in this repeat and that the common ancestor of the Triticeae tribe had multiple sequences of the 120-bp repeat with a range of variation not unlike that seen within and between species today. This diversity has been maintained when sites are moved within the genome and in all species since their divergence within the Triticeae.     

The genomic organization and evolutionary distribution of a tandemly repeated DNA sequence family in the genus Crocus (Iridaceae)

From a recombinant DNA-library from Crocus vernus, two closely related clones of highly repetitive DNA, pCvKB7 and pCvKB8, were sequenced and their genomic distribution and organization were investigated by Southern and in situ hybridization. The lengths of the clones were 181 and 178 bp respectively; the sequences were approximately 85% identical, and thus belonged to a sequence family, named the pCvKB8-family. No homologous sequences were found in the databases (BLAST made may 2004). The presence of pCvKB8 in 52 Crocus species and six species from other genera were analyzed by Southern hybridization. The sequence family was essentially Crocus-specific. However, the distribution of hybridization signal across the genus showed poor agreement with the taxonomic structure of the Crocus genus, suggesting that the subdivision does not follow the phylogeny of this sequence family. The chromosomal distribution on three Crocus species was essentially identical: tandem organization close to all telomeres and most centromeres, with a few additional intercalary sites.     

The cDNA cloning and molecular evolution of reptile and pigeon lactate dehydrogenase isozymes

The cDNAs encoding lactate dehydrogenase isozymes LDH-A (muscle) and LDH-B (heart) from alligator and turtle and LDH-A, LDH-B, and LDH-C (testis) from pigeon were cloned and sequenced. The evolutionary relationships among vertebrate LDH isozymes were analyzed. Contrary to the traditional belief that the turtle lineage branched off before the divergence between the lizard/alligator and bird lineages, the turtle lineage was found to be clustered with either the alligator lineage or the alligator-bird clade, while the lizard lineage was found to have branched off before the divergence between the alligator/turtle and bird lineages. The pigeon testicular LDH-C isozyme was evidently duplicated from LDH-B (heart), so it is not orthologous to the mammalian testicular LDH-C isozymes.      

High-resolution mapping of repetitive DNA by in situ hybridization: molecular and chromosomal features of prominent dispersed and discretely localized DNA families from the wild beet species Beta procumbens

Members of three prominent DNA families of Beta procumbens have been isolated as Sau3A repeats. Two families consisting of repeats of about 158 bp and 312 bp are organized as satellite DNAs (Sau3A satellites I and II), whereas the third family with a repeat length of 202 bp is interspersed throughout the genome. Multi-colour fluorescence in situ hybridization was used for physical mapping of the DNA families, and has shown that these tandemly organized families occur in large heterochromatic and DAPI positive blocks. The Sau3A satellite I hybridized exclusively around or near the centromeres of 10, 11 or 12 chromosomes. The Sau3A satellite family I showed high intraspecific variability and high-resolution physical mapping was performed on pachytene chromosomes using differentially labelled repeats. The physical order of satellite subfamily arrays along a chromosome was visualized and provided evidence that large arrays of plant satellite repeats are not contiguous and consist of distinct subfamily domains. Re-hybridization of a heterologous rRNA probe to mitotic metaphase chromosomes revealed that the 18S-5.8S-25S rRNA genes are located at subterminal position on one chromosome pair missing repeat clusters of the Sau3A satellite family I. It is known that arrays of Sau3A satellite I repeats are tightly linked to a nematode (Heterodera schachtii) resistance gene and our results show that the gene might be located close to the centromere. Large arrays of the Sau3A satellite II were found in centromeric regions of 16 chromosomes and, in addition, a considerable interspersion of repeats over all chromosomes was observed. The family of interspersed 202 bp repeats is uniformly distributed over all chromosomes and largely excluded from the rRNA gene cluster but shows local amplification in some regions. Southern hybridization has shown that all three families are specific for genomes of the section Procumbentes of the genus Beta.     

Relationships between species ofLeymus,Psathyrostachys, andHordeum (Poaceae,Triticeae) inferred from Southern hybridization of genomic and cloned DNA probes

We have used total genomic DNA as a probe to size-fractionated restriction enzyme digests of genomic DNA from a range ofTriticeae species from the generaLeymus Hochst.,Psathyrostachys Nevski, andHordeum L., and hybrids betweenHordeum andLeymus to investigate their taxonomic relationships. Genomic Southern hybridization was found to be an effective and simple way to assess the distribution and diversity of essentially species-specific and common, repetitive DNA sequences, and is hence especially useful in evolutionary studies. The DNA sequences ofH. vulgare seem to diverge substantially from those ofH. brachyantherum, H. lechleri, H. procerum, andH. depressum. The genome ofThinopyron bessarabicum shows little homology to those of theLeymus species investigated, confirming thatT. bessarabicum is not an ancestral genome inLeymus. Although the genomes ofLeymus andPsathyrostachys share substantial proportions of DNA sequences, they include divergent repeated sequences as well. Hybridization with a ribosomal DNA probe (pTa 71) showed that the coding regions containing structural genes encoding the 18 S, 5.8 S, and 26 S ribosomal RNA were conserved among the species investigated, whereas the intergenic spacer region was more variable, presenting different sizes of restriction fragments and enabling a classification of the species. The rye heterochromatin probe pSc 119.2 hybridized to DNA fromH. lechleri andT. bessarabicum, but not to DNA from the other species investigated.     

Investigations of Genome Relationships Between Leymus, Psathyrostachys and Hordeum Inferred by Genomic DNA:DNA in situ Hybridization

Genome relationships between the genera Leymus Hochst., PsathyrostachysNevski and Hordeum L. (Poaceae, Triticeae) were investigatedby fluorescent in situ hybridization using both total genomicDNA and cloned DNA sequences as probes. In hybrids between speciesof Hordeum and Leymus there was a clear differentiation betweenthe H genomes of Hordeum species and the genomes of Leymus speciesafter probing with genomic Hordeum or Leymus DNA. Chromosomesof species of Leymus and Psathyrostachys were also differentiatedby subtelomeric heterochromatic segments or by negative bandsalong their length. The number and location of 18S-5·8S-26SrRNA genes varied between the investigated genera. Unusually,L. angustus and P. stoloniformis rDNA sites were localized onboth ends of some chromosomes. Interphase nuclei of the Hordeumx Leymus hybrids had groups of chromosomes from both parentalgenomes in discrete, non-intermixed domains.Copyright 1994,1999 Academic Press Taxonomy, evolution, molecular evolution, repetitive DNA, rDNA sites, in situ hybridization, Triticeae, Leymus, Hordeum, Psathyrostachys     

Cytochalasin effects on structure and movement in the pollen tube of Iris

Summary Cytochalasins B and D in the concentration range 0.5–10 g ml^–1 produced similar effects on growth, movement and cytoplasmic structure in the pollen tubes of Iris spp. cultured in vitro. Continuous video recording showed that at 5 g ml_–1, CB was capable of stopping organelle circulation in as short a period as 20 s. The usually elongated vegetative nuclei were also arrested, and subsequently contracted irregularly. Generative cells were not radically changed in shape, but occasionally moved erratically before being halted. Detailed examination of CD- and CB-treated tubes regarded as being capable of recovering growth upon transfer to normal medium revealed several characteristic effects on cytoplasmic structure. Fibrils presumed to consist of, or contain, microfilament bundles are readily visible in the older parts of the living tube where they form the pathways of organelle movement; these were either condensed into amorphous columns or fragmented by treatment. In the distal parts of the tube, the cytoplasm had contracted into amorphous masses which continued to show very slow shape changes. With the arrest of extension growth, pectin accumulated over the tube tip and in patches along the flanks. In a medium containing 1 mM ATP, recovery from treatment was achieved in some instances within l min. Organelle circulation in the younger tubes was resumed, and fresh adventive tube tips were formed. The fibrillar system of the older tubes was not restored, however; instead, the cytoplasm in these zones formed aggregates which underwent continuous amoeboid movement, the organelles within moving rapidly in irregular trajectories with no indication of the resumption of the original long-range cyclotic flux. Some possible implications of the results are briefly discussed.