Biomedical subjects

Jonathan C Lamb

Publications and source records attributed to Jonathan C Lamb.

11 recordsLinked to original sources

PubMed2007

Recent proliferation and translocation of pollen group 1 allergen genes in the maize genome.

The dominant allergenic components of grass pollen are known by immunologists as group 1 allergens. These constitute a set of closely related proteins from the beta-expansin family and have been shown to have cell wall-loosening activity. Group 1 allergens may facilitate the penetration of pollen tubes through the grass stigma and style. In maize (Zea mays), group 1 allergens are divided into two classes, A and B. We have identified 15 genes encoding group 1 allergens in maize, 11 genes in class A and four genes in class B, as well as seven pseudogenes. The genes in class A can be divided by sequence relatedness into two complexes, whereas the genes in class B constitute a single complex. Most of the genes identified are represented in pollen-specific expressed sequence tag libraries and are under purifying selection, despite the presence of multiple copies that are nearly identical. Group 1 allergen genes are clustered in at least six different genomic locations. The single class B location and one of the class A locations show synteny with the rice (Oryza sativa) regions where orthologous genes are found. Both classes are expressed at high levels in mature pollen but at low levels in immature flowers. The set of genes encoding maize group 1 allergens is more complex than originally anticipated. If this situation is common in grasses, it may account for the large number of protein variants, or group 1 isoallergens, identified previously in turf grass pollen by immunologists.

Antigens, Plant↗

PubMed2006

Telomere-mediated chromosomal truncation in maize.

Direct repeats of Arabidopsis telomeric sequence were constructed to test telomere-mediated chromosomal truncation in maize. Two constructs with 2.6 kb of telomeric sequence were used to transform maize immature embryos by Agrobacterium-mediated transformation. One hundred seventy-six transgenic lines were recovered in which 231 transgene loci were revealed by a FISH analysis. To analyze chromosomal truncations that result in transgenes located near chromosomal termini, Southern hybridization analyses were performed. A pattern of smear in truncated lines was seen as compared with discrete bands for internal integrations, because telomeres in different cells are elongated differently by telomerase. When multiple restriction enzymes were used to map the transgene positions, the size of the smears shifted in accordance with the locations of restriction sites on the construct. This result demonstrated that the transgene was present at the end of the chromosome immediately before the integrated telomere sequence. Direct evidence for chromosomal truncation came from the results of FISH karyotyping, which revealed broken chromosomes with transgene signals at the ends. These results demonstrate that telomere-mediated chromosomal truncation operates in plant species. This technology will be useful for chromosomal engineering in maize as well as other plant species.

Chromosomes, Plant↗

PubMed2006

Cytological visualization of DNA transposons and their transposition pattern in somatic cells of maize.

Global genomic analysis of transposable element distributions of both natural (En/Spm, Ac-Ds, and MuDR/Mu) and modified (RescueMu) types was performed by fluorescence in situ hybridization (FISH) on somatic chromosomes coupled with karyotyping of each chromosome. In lines without an active transposable element, the locations of silent En/Spm, Ac-Ds, and MuDR/Mu elements were visualized, revealing variation in copy number and position among lines but no apparent locational bias. The ability to detect single elements was validated by using previously mapped active Ac elements. Somatic transpositions were documented in plants containing an engineered Mutator element, RescueMu, via use of the karyotyping system. By analyzing the RescueMu lines, we found that transposition of RescueMu in root-tip cells follows the cut-and-paste type of transposition. This work demonstrates the utility of FISH and karyotyping in the study of transposon activity and its consequences.

DNA Transposable Elements↗

PubMed2006

Retroelement genome painting: cytological visualization of retroelement expansions in the genera Zea and Tripsacum.

Divergence of abundant genomic elements among the Zea and Tripsacum genera was examined cytologically and a tool kit established for subsequent studies. The LTR regions from the CRM, Huck, Grande, Prem1, Prem2/Ji, Opie, Cinful-1, and Tekay retroelement families were used as FISH probes on mitotic chromosome spreads from a "trispecies" hybrid containing chromosomes from each of three species: Zea mays (2n = 20), Z. diploperennis (2n = 20), and Tripsacum dactyloides (2n = 36). Except for Tekay, which painted both Zea and Tripsacum chromosomes with nearly equal intensity, the retroelement probes hybridized strongly to the Zea chromosomes, allowing them to be distinguished from those of Tripsacum. Huck and Grande hybridized more intensely to maize than to Z. diploperennis chromosomes. Tripsacum genomic clones containing retroelement sequences were isolated that specifically paint Tripsacum chromosomes. The retroelement paints proved effective for distinguishing different genomes in interspecific hybrids and visualizing alien chromatin from T. dactyloides introgressed into maize lines. Other FISH probes (180-bp knob, TR-1, 5S, NOR, Cent4, CentC, rp1, rp3, and alpha-ZeinA) could be simultaneously visualized with the retroelement probes, emphasizing the value of the retroelement probes for cytogenetic studies of Zea and Tripsacum.

Chromosome Painting↗

PubMed2006

High frequency of centromere inactivation resulting in stable dicentric chromosomes of maize.

Somatic chromosome spreads from maize (Zea mays L.) plants containing B-A translocation chromosomes undergoing the chromosome type breakage-fusion-bridge cycle were examined by FISH. The size and type of extra chromosomes varied among cells of the same individual. A collection of minichromosomes derived from the chromosome type breakage-fusion-bridge cycle was examined for the presence of stable dicentric chromosomes. Six of 23 chromosomes in the collection contained two regions with DNA sequences typical of centromeres. Functional analysis and immunolabeling of CENH3, the centromere-specific histone H3 variant, revealed only one functional centromere per chromosome, despite the duplicate centromere sequences. One plant was found with an inactive B centromere that had been translocated to the short arm of chromosome 9. The translocated centromere region appeared identical to that of a normal B chromosome. The inactivation of the centromeres was stable for at least four generations. By using dicentrics from dispensable chromosomes, centromere inactivation was found to be quite common under these circumstances.

Biological Transport↗

PubMed2005

Molecular and functional dissection of the maize B chromosome centromere.

The centromere of the maize (Zea mays) B chromosome contains several megabases of a B-specific repeat (ZmBs), a 156-bp satellite repeat (CentC), and centromere-specific retrotransposons (CRM elements). Here, we demonstrate that only a small fraction of the ZmBs repeats interacts with CENH3, the histone H3 variant specific to centromeres. CentC, which marks the CENH3-associated chromatin in maize A centromeres, is restricted to an approximately 700-kb domain within the larger context of the ZmBs repeats. The breakpoints of five B centromere misdivision derivatives are mapped within this domain. In addition, the fraction of this domain remaining after misdivision correlates well with the quantity of CENH3 on the centromere. Thus, the functional boundaries of the B centromere are mapped to a relatively small CentC- and CRM-rich region that is embedded within multimegabase arrays of the ZmBs repeat. Our results demonstrate that the amount of CENH3 at the B centromere can be varied, but with decreasing amounts, the function of the centromere becomes impaired.

Centromere↗

PubMed2005

Advances in plant chromosome identification and cytogenetic techniques.

Recent developments that improve our ability to distinguish slightly diverged genomes from each other, as well as to distinguish each of the nonhomologous chromosomes within a genome, add a new dimension to the study of plant genomics. Differences in repetitive sequences among different species have been used to develop multicolor fluorescent in situ hybridization techniques that can define the components of allopolyploids in detail and reveal introgression between species. Bacterial artificial chromosome probes and repetitive sequence arrays have been used to distinguish each of the nonhomologous somatic chromosomes within a species. Such karyotype analysis opens new avenues for the study of chromosomal variation and behavior, as well as for the localization of individual genes and transgenes to genomic position.

Chromosomes, Plant↗

PubMed2004

Sequences associated with A chromosome centromeres are present throughout the maize B chromosome.

Maize chromosome spreads containing the supernumerary B chromosome were hybridized with probes from various repetitive elements including CentC, CRM, and CentA, which have been localized to centromeric regions on the A chromosomes. Repetitive elements that are enriched or found exclusively near the centromeres of A chromosomes hybridized to many sites distinct from the centromere on the B chromosome. To examine whether these elements recruit kinetochore proteins at locations other than the canonical B centromere, cells were labeled with antibodies against CENH3, a key kinetochore protein. No labeling was detected outside the normal centromere and no evidence of B chromosome holocentromeric activity was observed. This finding suggests that, as in other higher eukaryotes, DNA sequence alone is insufficient to dictate kinetochore location in plants. Additionally, examination of the B centromere region in pachytene chromosomes revealed that the B-specific element ZmBs hybridizes to a much larger region than the site of hybridization of CentC, CRM, and CentA and the labeling by anti-CENH3 antibodies.

Centromere↗

PubMed2004

Chromosome painting using repetitive DNA sequences as probes for somatic chromosome identification in maize.

Study of the maize (Zea mays L.) somatic chromosomes (2n = 20) has been difficult because of a lack of distinguishing characteristics. To identify all maize chromosomes, a multicolor fluorescence in situ hybridization procedure was developed. The procedure uses tandemly repeated DNA sequences to generate a distinctive banding pattern for each of the 10 chromosomes. Fluorescence in situ hybridization screening trials of nonsubtracted or subtracted PCR libraries resulted in the isolation of microsatellite 1-26-2, subtelomeric 4-12-1, and 5S rRNA 2-3-3 clones. These three probes, plus centromeric satellite 4 (Cent4), centromeric satellite C (CentC), knob, nucleolus-organizing region (NOR), pMTY9ER telomere-associated sequence, and tandemly repeated DNA sequence 1 (TR-1) were used as a mixture for hybridization to root-tip chromosomes. All 10 chromosomes were identified by the banding and color patterns in the 14 examined lines. There was significant quantitative variation among lines for the knob, microsatellite, TR-1, and CentC signals. The same probe mixture identifies meiotic pachytene, late prophase I, and metaphase I chromosomes. The procedure could facilitate the study of chromosomal structure and behavior and be adapted for other plant species.

Chromosome Painting↗

PubMed2004

What's in a centromere?

The complete sequence of rice centromere 8 reveals a small amount of centromere-specific satellite sequence in blocks interrupted by retrotransposons and other repetitive DNA, in an arrangement that is strikingly similar in overall size and content to other centromeres of multicellular eukaryotes.

Animals↗

PubMed2003

The role of DNA sequence in centromere formation.

Centromeres are key to the correct segregation and inheritance of genetic information. Eukaryotic centromeres, which are located in large blocks of highly repetitive DNA, have been notoriously difficult to sequence. Several groups have recently succeeded in analyzing centromeric sequences in human, Drosophila and Arabidopsis, providing new insights into the importance of DNA sequence for centromere function.

Animals↗