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James A Birchler

Publications and source records attributed to James A Birchler.

At least 19 recordsLinked to original sources
PubMed2025

Genome imbalance modulates the expression of long non-coding RNAs in maize.

Genome imbalance, resulting from varying the dosage of individual chromosomes (aneuploidy), has a more detrimental effect than changes in complete sets of chromosomes (haploidy/polyploidy). This imbalance is likely due to disruptions in stoichiometry and interactions among macromolecular assemblies. Previous research has shown that aneuploidy causes global modulation of protein-coding genes (PCGs), microRNAs, and transposable elements (TEs), affecting both the varied chromosome (cis-located) and unvaried genome regions (trans-located) across various taxa. While long non-coding RNAs (lncRNAs) are important gene expression regulators, their roles in the context of genomic imbalance remain largely unexplored. In this study, we analyzed and compared the impact of aneuploidy and haploidy/polyploidy on lncRNA expression using RNA-seq data from maize mature leaf tissue. Our results indicate that cis-located lncRNAs are modulated from dosage compensation to a gene dosage effect, while trans-located lncRNAs exhibit trends ranging from an inverse effect to a positive correlation with chromosomal dosage. Remarkably, the ploidy series showed a lesser degree of lncRNA modulation. LncRNAs and TEs display a similar trend of inverse modulation but exhibit greater sensitivity to dosage changes compared to PCGs. The construction of cis-acting and trans-acting lncRNA co-expression networks indicates that lncRNAs likely function as dosage-sensitive regulators of gene expression under conditions of genomic imbalance. Overall, this study not only elucidates the dosage effect of plant lncRNAs but also serves as a valuable resource for exploring potential regulators of PCGs that play significant biological functions.

Zea mays↗
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

Genetic variation for the response to ploidy change in Zea mays L.

Polyploidization is an important process in the evolutionary history of most eukaryotic species. It oftentimes causes large-scale genomic reorganizations and is accompanied by a wide variety of phenotypic alterations in morphology, niche preference and fitness characteristics. Despite their importance, the morphological effects of alterations in ploidy are not well understood. We investigated these changes in four diverse maize inbred lines, using monoploid, diploid, triploid and tetraploid derivatives, measuring 13 characters in a randomized field study. Employing several analysis of variance approaches, we find that all characters investigated strongly respond to alterations in ploidy. This response appears to have two sources: one source is shared by all inbred lines and constitutes a common response to ploidy change. The other source is genotype specific and results in a response to ploidy change that varies among inbred lines. This finding demonstrates the existence of genetic variation for the morphological response to ploidy change in Zea mays.

Analysis of Variance↗
PubMed2006

Misregulation of sex-lethal and disruption of male-specific lethal complex localization in Drosophila species hybrids.

A major model system for the study of evolutionary divergence between closely related species has been the unisexual lethality resulting from reciprocal crosses of Drosophila melanogaster and D. simulans. Sex-lethal (Sxl), a critical gene for sex determination, is misregulated in these hybrids. In hybrid males from D. melanogaster mothers, there is an abnormal expression of Sxl and a failure of localization of the male-specific lethal (MSL) complex to the X chromosome, which causes changes in gene expression. Introduction of a Sxl mutation into this hybrid genotype will allow expression of the MSL complex but there is no sequestration to the X chromosome. Lethal hybrid rescue (Lhr), which allows hybrid males from this cross to survive, corrects the SXL and MSL defects. The reciprocal cross of D. simulans mothers by D. melanogaster males exhibits underexpression of Sxl in embryos.

Animals↗
PubMed2006

Polycomb, pairing and PIWI--RNA silencing and nuclear interactions.

In Drosophila, the RNA interference (RNAi) genes participate in Polycomb (Pc)-mediated transgene silencing. Recently, the involvement of the RNAi genes in Pc silencing, pairing-sensitive silencing and long-range contacts among Pc-associated sequences has been explored. These Pc-associated sequences are involved with the control of the proper expression of developmental HOX genes.

Animals↗
PubMed2006

Organization of endoreduplicated chromosomes in the endosperm of Zea mays L.

The chromosomes of the maize endosperm proceed through an endoreduplication phase in later stages of development. Endoreduplication is a process in which the cell cycle continues DNA synthesis but does not proceed through cytokinesis. When this occurs, the normally triploid endosperm cell can reach ploidy levels greater than 200x in some lines of maize. In this work, we examined the structure of the endoreduplicated chromosomes. Previous cytological work has indicated that, although the DNA content per cell increases, the number of nucleoli and knobs remains the same. Using fluorescence in situ hybridization and slot blot techniques, we show that the highly repetitive heterochromatic areas both on the A and B chromosomes, as well as several actively transcribed genes, are endoreduplicated. This result suggests that the entire genome follows that same trend. Further evidence shows that the various chromatin strands stay associated throughout the length of the chromosomes after they have been replicated, and that the DNA at the centromeric and knob regions is more tightly associated than the other regions of the chromosomes. Interploidy crosses between diploid and tetraploid derivatives of the same inbred exhibit changes in the chromatin organization of centromeres and heterochromatic knobs.

Base Sequence↗
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↗
PubMed2006

Induction of tetraploid derivatives of maize inbred lines by nitrous oxide gas treatment.

Maize (Zea mays L.) is a model organism for various genetic and physiological studies. Induction of autotetraploid lines from elite inbred lines is valuable for investigating gene dosage effects on the molecular level. We applied nitrous oxide gas at the time of fertilization (30-36 h after pollination) for 20 h on maize inbred line Oh43. The nitrous oxide gas treatment between pressures of 600-1000 kPa proved to be effective in inducing tetraploids. The treatment also significantly increased the rates of germless and shriveled kernels. Twelve inbred lines were treated with nitrous oxide gas for 20 h at either 800 or 900 kPa pressures, 30 or 36 h after pollination. Although tetraploid or tetraploid class aneuploid plants from 9 of 12 inbreds tested were successfully generated, only six genotypes produced progenies. The successful tetraploid inbred lines were from the A188, B73, H99, Oh43, Stock 6, and W22 genetic backgrounds. Aneuploids, plants with broken chromosomes and chimeras, were also found among the treated materials.

Chromosomes, Plant↗
PubMed2006

Commonalities in compensation.

The sex chromosomes of many species differ in dosage but the total gene expression output is similar, a phenomenon referred to as dosage compensation. Previously, diverse mechanisms were postulated to account for compensation in distantly related taxa. However, two recent papers present evidence that dosage compensation in Drosophila, mammals and nematodes share the property that there is an approximately two-fold upregulation of the single active X chromosome in each case.(1,2) The results suggest that a common mechanism might operate in these different cases.

Animals↗
PubMed2006

Characterization of a maize isochromosome 8S*8S.

An isochromosome was found in the maize HiII Parent B line during somatic karyotyping with a multiprobe fluorescence in situ hybridization (FISH) system. Cytological analyses showed that it pairs with the short arm of chromosome 8 during the pachytene stage of meiosis. The chromosome 8 short arm origin of this isochromosome was also confirmed by FISH at mitotic metaphase. Knob heterochromatin signals were present at the short arms of chromosome 8 when subjected to prolonged exposure and also observed at both ends of the isochromosome. This isochromosome can be a univalent or a trivalent by pairing with the normal chromosome 8 short arms during meiosis. At anaphase and telophase, the isochromosome lagged behind other chromosomes. It had a transmission rate of 17%-20% from both male and female gametes. One plant homozygous for the isochromosome contained 2 isochromosomes that differed in the quantity of their CentC centromere repeat sequence. Both variations of the isochromosome were transmitted to the next generation. Because the 2 isochromosomes should be identical by descent, these observations document a radical change in copy number of the centromere repeat array within 1 generation. Plants with 1 isochromosome were not normal as compared with the original HiII Parent B plants. Those that contained a pair of this isochromosome (6 total copies of 8S) were even more abnormal and had reduced fertility. The results indicate the ability of the somatic karyotyping system to recognize and characterize chromosomal aberrations.

Centromere↗
PubMed2005

RNA silencing in Drosophila.

Knowledge of the role of RNA in affecting gene expression has expanded in the past several years. Small RNAs serve as homology guides to target messenger RNAs for destruction at the post-transcriptional level in the experimental technique known as RNA interference and in the silencing of some transgenes. These small RNAs are also involved in sequence-specific targeting of chromatin modifications for transcriptional silencing of transgenes, transposable elements, heterochromatin and some cases of Polycomb-mediated gene silencing. RNA silencing processes in Drosophila are described.

Animals↗
PubMed2005

The dominant inhibitory chalcone synthase allele C2-Idf (inhibitor diffuse) from Zea mays (L.) acts via an endogenous RNA silencing mechanism.

The flavonoid pigment pathway in plants has been used as a model system for studying gene regulatory mechanisms. C2-Idf is a stable dominant mutation of the chalcone synthase gene, c2, which encodes the first dedicated enzyme in this biosynthetic pathway of maize. Homozygous C2-Idf plants show no pigmentation. This allele also inhibits expression of functional C2 alleles in heterozygotes, producing a less pigmented condition instead of the normal deeply pigmented phenotype. To explore the nature of this effect, the C2-Idf allele was cloned. The gene structure of the C2-Idf haplotype differs substantially from that of the normal c2 gene in that three copies are present. Two of these are located in close proximity to each other in a head-to-head orientation and the third is closely linked. Previous experiments showed that the lower level of pigmentation in heterozygotes is correlated with reduced enzyme activity and low steady-state mRNA levels. We found that c2 transcription occurs in nuclei of C2-Idf/C2 heterozygotes, but mRNA does not accumulate, suggesting that the inhibition is mediated by RNA silencing. Infection of C2-Idf/C2 heterozygotes with viruses that carry suppressors of RNA silencing relieved the phenotypic inhibition, restoring pigment production and mRNA levels. Finally, we detected small interfering RNAs (siRNAs) in plants carrying C2-Idf, but not in plants homozygous for the wild-type C2 allele. Together, our results indicate that the inhibitory effect of C2-Idf occurs through RNA silencing.

Acyltransferases↗
PubMed2005

A test for ectopic exchange catalyzed by Cre recombinase in maize.

A maize line expressing Cre recombinase as well as the recipient line without the transgene were assayed for evidence of ectopic recombination within the maize genome. Such a test is valuable for understanding the action of Cre as well as for its use to recombine two target lox sites present in the chromosomes. Pollen examination and seed set tests of material expressing Cre provided no evidence of ectopic recombination, which would be manifested in the production of translocations or inversions and result in pollen abortion and reduced seed set. Root-tip chromosome karyotype analysis was also performed on material with and without Cre expression. Chromosomal aberrations in Cre+ material were not observed above the background level.

Chromosome Aberrations↗
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

Gene expression analysis of the function of the male-specific lethal complex in Drosophila.

Dosage compensation refers to the equal expression of X-linked genes despite the difference in copy number between the two sexes. The male-specific lethal (MSL) complex is concentrated on the X chromosome in males. A gene expression assay for embryos was developed to examine the function of this complex. In mutant male embryos without either the MSL complex or MOF histone acetylase, dosage compensation is retained but autosomal expression is increased. Dosage compensation is lost in the double-mutant embryos. In embryos in which the MSL complex and MOF are targeted to the X chromosomes in females, the results are consistent with previous surveys showing that in general the X expression remains unchanged, but autosomal expression is reduced. Mutations in the ISWI chromatin-remodeling component cause increases specifically of X-linked genes in males. Thus, the function of the MSL complex in conjunction with ISWI is postulated to override the effect on gene expression of high histone acetylation on the male X. The basic determinant of dosage compensation is suggested to be an inverse dosage effect produced by an imbalance of transcription factors on the X vs. the autosomes. The sequestration of the MSL complex to the male X may have evolved to counteract a similar effect on the autosomes and to prevent an overexpression of the X chromosome in males that would otherwise occur due to the high levels of histone acetylation.

Acetyltransferases↗