Robotic excision and ureteroureterostomy of blind-ending ureteral 'Duplication'.

INTRODUCTION: Duplicated ureteral anatomy can be a reconstructive challenge. Blind-ending ureteral duplication has been reported with recommendations for surgical excision. OBJECTIVE: This video reviews the importance of exposure of anatomic landmarks in surgical excision of a blind-ending ureteral duplication. MATERIALS AND METHODS: This is a retrospective case report of a patient who presented with a blind-ending ureteral duplication. DISCUSSION: A 13-year-old female presented with a right-sided abdominal mass. Abdominal and pelvic imaging revealed a tubular structure adjacent to and below the right kidney, possibly connecting to the right lower pole. While initially observed, the patient re-presented with urosepsis. A retrograde pyelogram showed no connection between the right ureter and this structure. The patient underwent robotic-assisted excision of this structure. Intra-operatively, it was connected to the right lower pole calyces. A ureteroureterostomy to the orthotopic ureter was performed. Although the structure was adjacent to the bladder dome, there was no communication distally. Postoperatively, the patient did well; follow-up imaging showed a non-dilated lower pole. The pathology of this structure was benign urothelium. CONCLUSIONS: Surgical management of aberrant ureteral duplications should focus on identifying known landmarks and should be considered to prevent symptomatic infections and renal scarring.

A plant-transformation-competent BIBAC library from the Arabidopsis thaliana Landsberg ecotype for functional and comparative genomics.

The genome of the model plant Arabidopsis thaliana has been sequenced to near completion. To facilitate experimental determination of the function of every gene in the species, we constructed a large-insert library from the Landsberg ecotype using a plant-transformation-competent binary BAC vector, BIBAC2. The library contains 11,520 clones with an estimated average insert size of 162 kb. Of a sample of 102 clones, 17.6% had no inserts; further, in the library as a whole, 287 clones contained chloroplast DNA, and 25 contained mitochondrial DNA. Thus it is estimated that 9,295 clones originated from the nuclear genome, representing a 11.5 x coverage. The library was further characterized by screening with probes corresponding to 180-bp repeats, 5S rDNA, 18S-25S rDNA and 23 single-copy RFLP markers. The results showed that 92 clones contained 180-bp centromeric repeats, 78 contained 5S rDNA and 95 contained 18S-25S rDNA, approximately 1%, 0.8% and 1%, respectively, of the nuclear clones in the library. Screening the library with the 23 RFLP markers showed that each one hybridized to an average of seven clones. This library is the first large-insert DNA library for the widely studied Landsberg erecta strain. It will greatly facilitate gene identification by complementation screening, and will enhance analysis of the structure, organization and evolution of the A. thaliana genome.

Gene families from the Arabidopsis thaliana pollen coat proteome.

The pollen extracellular matrix contains proteins mediating species specificity and components needed for efficient pollination. We identified all proteins >10 kilodaltons in the Arabidopsis pollen coating and showed that most of the corresponding genes reside in two genomic clusters. One cluster encodes six lipases, whereas the other contains six lipid-binding oleosin genes, including GRP17, a gene that promotes efficient pollination. Individual oleosins exhibit extensive divergence between ecotypes, but the entire cluster remains intact. Analysis of the syntenic region in Brassica oleracea revealed even greater divergence, but a similar clustering of the genes. Such allelic flexibility may promote speciation in plants.

Sequence and analysis of the Arabidopsis genome.

The comprehensive analysis of the genome sequence of the plant Arabidopsis thaliana has been completed recently. The genome sequence and associated analyses provide the foundations for rapid progress in many fields of plant research, such as the exploitation of genetic variation in Arabidopsis ecotypes, the assessment of the transcriptome and proteome, and the association of genome changes at the sequence level with evolutionary processes. Nevertheless, genome sequencing and analysis are only the first steps towards a new plant biology. Much remains to be done to refine the analysis of encoded genes, to define the functions of encoded proteins systematically, and to establish new generations of databases to capture and relate diverse data sets generated in widely distributed laboratories.

Systematic search and molecular characterization of the antigenic targets of myeloma immunoglobulins: a monoclonal IgA from a female patient targeting sperm-specific cylicin II.

The identification of the antigenic stimuli of B-cell neoplasms might be of considerable importance since a causal relationship between these neoplasms and antigenic stimulation has been suggested. To date the identification of such antigens has been erratic and accidental. For a systematic search and molecular characterization of human proteins that are antigenic target structures of myeloma-associated immunoglobulins, we applied SEREX (serological analysis of antigens by recombinant cDNA expression cloning) using a testis cDNA expression library and myeloma proteins from 42 patients. A monoclonal IgA from a female patient was shown to target sperm-specific cylicin II. The specificity of the reaction was confirmed by the characteristic staining of the equatorial belt of human sperm heads by the patient's myeloma protein. Serological analysis of recombinantly expressed cDNAs is a straightforward and high throughput approach for the molecular characterization of the targets of myeloma-associated immunoglobulins. The analysis of the antigenic spectrum of immunoglobulins associated with B-cell neoplasms will provide valuable information for the understanding of the pathogenesis of these diseases.

Pollen tube targeting and axon guidance: parallels in tip growth mechanisms.

The growth of pollen tubes to plant egg cells and the guidance of axons to neural synapses are classic examples of targeted cell growth. Despite the evolutionary time that separates animals and plants, axon and pollen tube guidance share remarkable mechanistic similarities. In both instances, extracellular cues are transduced by intracellular signal-transduction pathways that culminate in directed tip growth. Do the mechanistic similarities extend to the molecular level? Here, we address this question by a comprehensive review of the molecules and pathways involved in pollen tube targeting and axon guidance. The emerging scenario is that similar intracellular molecules are recruited to control tip growth, while different extracellular molecules mediate guidance through the distinct plant and animal extracellular matrices.

Alterations in CER6, a gene identical to CUT1, differentially affect long-chain lipid content on the surface of pollen and stems.

Very long chain lipids contribute to the hydrophobic cuticle on the surface of all land plants and are an essential component of the extracellular pollen coat in the Brassicaceae. Mutations in Arabidopsis CER genes eliminate very long chain lipids from the cuticle surface and, in some cases, from the pollen coat. In Arabidopsis, the loss of pollen coat lipids can disrupt interactions with the stigma, inhibiting pollen hydration and causing sterility. We have positionally cloned CER6 and demonstrate that a wild-type copy complements the cer6-2 defect. In addition, we have identified a fertile, intragenic suppressor, cer6-2R, that partially restores pollen coat lipids but does not rescue the stem wax defect, suggesting an intriguing difference in the requirements for CER6 activity on stems and the pollen coat. Importantly, analysis of this suppressor demonstrates that low amounts of very long chain lipids are sufficient for pollen hydration and germination. The predicted CER6 amino acid sequence resembles that of fatty acid-condensing enzymes, consistent with its role in the production of epicuticular and pollen coat lipids >28 carbons long. DNA sequence analysis revealed the nature of the cer6-1, cer6-2, and cer6-2R mutations, and segregation analysis showed that CER6 is identical to CUT1, a cDNA previously mapped to a different chromosome arm. Instead, we have determined that a new gene, CER60, with a high degree of nucleotide and amino acid similarity to CER6, resides at the original CUT1 locus.

Tetrad analysis in higher plants. A budding technology.

Tetrad analysis, the ability to manipulate and individually study the four products of a single meiotic event, has been critical to understanding the mechanisms of heredity. The Arabidopsis quartet (qrt) mutation, which causes the four products of male meiosis to remain attached, enables plant biologists to apply this powerful tool to investigations of gamete development, cell division, chromosome dynamics, and recombination. Here we highlight several examples of how qrt has been used to perform tetrad analysis and suggest additional applications including a genetic screen for gametophytic mutants and methods for investigating gene interactions by synthetic lethal analysis.

Pollen-stigma adhesion in Arabidopsis: a species-specific interaction mediated by lipophilic molecules in the pollen exine.

To investigate the nature and role of cell adhesion in plants, we analyzed the initial step of pollination in Arabidopsis: the binding of pollen grains to female stigma cells. Here we show this interaction occurs within seconds of pollination. Because it takes place prior to pollen hydration, it also requires adhesion molecules that can act in a virtually dry environment. We developed assays that monitored adhesion of populations of pollen grains and individual cells. Adhesion between pollen and stigma cells is highly selective - Arabidopsis pollen binds with high affinity to Arabidopsis stigmas, while pollen from other species fails to adhere. Initial binding is independent of the extracellular pollen coat (tryphine), indicating that adhesion molecules reside elsewhere on the pollen surface, most likely within the exine walls. Immediately after pollination, the stigma surface becomes altered at the interface, acquiring a pattern that interlocks with the exine; this pattern is evident only with pollen from Arabidopsis and its close relatives. Purified exine fragments bind to stigma cells, and biochemical analyses indicate that this specific, rapid and anhydrous adhesion event is mediated by lipophilic interactions.

Centromeres in the genomic era: unraveling paradoxes.

The centromeres of higher plants and animals share many common features, though current models fail to account for all aspects of centromere composition and function. This dilemma is likely to be resolved in the next few years in Arabidopsis where robust assays for centromere function are available and the sequence of the entire genome will be determined.

The mating game: pollination and fertilization in flowering plants.

Recent work has revealed signaling molecules that control pollination, including small peptides that mediate pollen recognition and glycoproteins that support pollen tube growth. The polarized growth of pollen tubes requires a calcium-mediated signal cascade, and cues derived from the haploid and diploid ovule cells guide pollen tubes to the eggs.

Sequence and analysis of chromosome 2 of the plant Arabidopsis thaliana.

Arabidopsis thaliana (Arabidopsis) is unique among plant model organisms in having a small genome (130-140 Mb), excellent physical and genetic maps, and little repetitive DNA. Here we report the sequence of chromosome 2 from the Columbia ecotype in two gap-free assemblies (contigs) of 3.6 and 16 megabases (Mb). The latter represents the longest published stretch of uninterrupted DNA sequence assembled from any organism to date. Chromosome 2 represents 15% of the genome and encodes 4,037 genes, 49% of which have no predicted function. Roughly 250 tandem gene duplications were found in addition to large-scale duplications of about 0.5 and 4.5 Mb between chromosomes 2 and 1 and between chromosomes 2 and 4, respectively. Sequencing of nearly 2 Mb within the genetically defined centromere revealed a low density of recognizable genes, and a high density and diverse range of vestigial and presumably inactive mobile elements. More unexpected is what appears to be a recent insertion of a continuous stretch of 75% of the mitochondrial genome into chromosome 2.

Genetic definition and sequence analysis of Arabidopsis centromeres.

High-precision genetic mapping was used to define the regions that contain centromere functions on each natural chromosome in Arabidopsis thaliana. These regions exhibited dramatic recombinational repression and contained complex DNA surrounding large arrays of 180-base pair repeats. Unexpectedly, the DNA within the centromeres was not merely structural but also encoded several expressed genes. The regions flanking the centromeres were densely populated by repetitive elements yet experienced normal levels of recombination. The genetically defined centromeres were well conserved among Arabidopsis ecotypes but displayed limited sequence homology between different chromosomes, excluding repetitive DNA. This investigation provides a platform for dissecting the role of individual sequences in centromeres in higher eukaryotes.

The Arabidopsis transposable element Tag1 is widely distributed among Arabidopsis ecotypes.

Tag1 is an autonomous transposable element (3.3 kb in length) first identified as an insertion in the CHL1 (NRT1) gene of Arabidopsis thaliana. Tag1 has been found in the Landsberg erecta ecotype of A. thaliana but not in Columbia or WS. In this paper, 41 additional ecotypes were examined for the presence of Tag1. Using an internal Tag1 fragment as probe, we found that DNA form 19 of the 41 ecotypes strongly hybridized to Tag1. Almost all of the Tag1-containing ecotypes had only one or two copies of Tag1 per haploid genome, as determined by Southern blot analysis. The only exception, Bf-1 from Bretagny-sur-Orge, France, had four copies. Two ecotypes, Di-G and S96, gave identical Southern blot patterns to that of Landsberg erecta and were subsequently shown to contain Tag1 at the same two positions found in Landsberg erecta (loci designated as Tag1-2 and Tag1-3). Two other ecotypes, Ag-0 and Lo-1, had a Tag1 element located at Tag1-2 but not at Tag1-3. The distance between these two loci was determined to be 0.37 cM. Analysis of DNA from two related species, A. griffithiana and A. pumila, showed that both species contain sequences that hybridize to Tag1 and that could be amplified with an oligonucleotide specific to the terminal inverted repeats of Tag1. These results show that Tag1 and related elements are present, and may be useful for insertional mutagenesis, in many A. thaliana ecotypes and several Arabidopsis species.

Assaying genome-wide recombination and centromere functions with Arabidopsis tetrads.

During meiosis, crossover events generate new allelic combinations, yet the abundance of these genetic exchanges in individual cells has not been measured previously on a genomic level. To perform a genome-wide analysis of recombination, we monitored the assortment of genetic markers in meiotic tetrads from Arabidopsis. By determining the number and distribution of crossovers in individual meiotic cells, we demonstrated (i) surprisingly precise regulation of crossover number in each meiosis, (ii) considerably reduced recombination along chromosomes carrying ribosomal DNA arrays, and (iii) an inversely proportional relationship between recombination frequencies and chromosome size. This use of tetrad analysis also achieved precise mapping of all five Arabidopsis centromeres, localizing centromere functions in the intact chromosomes of a higher eukaryote.

TETRASPORE is required for male meiotic cytokinesis in Arabidopsis thaliana.

In flowering plants, male meiosis occurs in the microsporocyte to produce four microspores, each of which develops into a pollen grain. Here we describe four mutant alleles of TETRASPORE (TES), a gene essential for microsporocyte cytokinesis in Arabidopsis thaliana. Following failure of male meiotic cytokinesis in tes mutants, all four microspore nuclei remain within the same cytoplasm, with some completing their developmental programmes to form functional pollen nuclei. Both of the mitotic divisions seen in normal pollen development take place in tes mutants, including the asymmetric division required for the differentiation of gametes; some tes grains perform multiple asymmetric divisions in the same cytoplasm. tes pollen shows a variety of abnormalities subsequent to the cytokinetic defect, including fusion of nuclei, formation of ectopic internal walls, and disruptions to external wall patterning. In addition, ovules fertilized by tes pollen often abort, possibly because of excess paternal genomes in the endosperm. Thus tes mutants not only reveal a gene specific to male meiosis, but aid investigation of a wide range of processes in pollen development and function.