Biomedical subjects

Ronald R Sederoff

Publications and source records attributed to Ronald R Sederoff.

13 recordsLinked to original sources

PubMed2005

Genetic architecture of transcript-level variation in differentiating xylem of a eucalyptus hybrid.

Species diversity may have evolved by differential regulation of a similar set of genes. To analyze and compare the genetic architecture of transcript regulation in different genetic backgrounds of Eucalyptus, microarrays were used to examine variation in mRNA abundance in the differentiating xylem of a E. grandis pseudobackcross population [E. grandis x F(1) hybrid (E. grandis x E. globulus)]. Least-squares mean estimates of transcript levels were generated for 2608 genes in 91 interspecific backcross progeny. The quantitative measurements of variation in transcript abundance for specific genes were mapped as expression QTL (eQTL) in two single-tree genetic linkage maps (F(1) hybrid paternal and E. grandis maternal). EQTL were identified for 1067 genes in the two maps, of which 811 were located in the F(1) hybrid paternal map, and 451 in the E. grandis maternal map. EQTL for 195 genes mapped to both parental maps, the majority of which localized to nonhomologous linkage groups, suggesting trans-regulation by different loci in the two genetic backgrounds. For 821 genes, a single eQTL that explained up to 70% of the transcript-level variation was identified. Hotspots with colocalized eQTL were identified in both maps and typically contained genes associated with specific metabolic and regulatory pathways, suggesting coordinated genetic regulation.

Cell Differentiation↗

PubMed2004

Microarray analyses of gene expression during adventitious root development in Pinus contorta.

In order to investigate the gene expression pattern during adventitious root development, RNA of Pinus contorta hypocotyls, pulse-treated with the auxin indole-3-butyric acid and harvested at distinct developmental time points of root development, was hybridized to microarrays containing 2,178 cDNAs from Pinus taeda. Over the period of observation of root development, the transcript levels of 220 genes changed significantly. During the root initiation phase, genes involved in cell replication and cell wall weakening and a transcript encoding a PINHEAD/ZWILLE-like protein were up-regulated, while genes related to auxin transport, photosynthesis, and cell wall synthesis were down-regulated. In addition, there were changes in transcript abundance of genes related to water stress. During the root meristem formation phase the transcript abundances of genes involved in auxin transport, auxin responsive transcription, and cell wall synthesis, and of a gene encoding a B-box zinc finger-like protein, increased, while those encoding proteins involved in cell wall weakening decreased. Changes of transcript abundance of genes related to water stress during the root meristem formation and root formation phase indicate that the plant roots had become functional in water transport. Simultaneously, genes involved in auxin transport were up-regulated, while genes related to cell wall modification were down-regulated. Finally, during the root elongation phase down-regulation of transcripts encoding proteins involved in cell replication and stress occurred. Based on the observed changes in transcript abundances, we suggest hypotheses about the relative importance of various physiological processes during the auxin-induced development of roots in P. contorta.

Cell Division↗

PubMed2004

Metabolic profiling: a new tool in the study of wood formation.

In the realm of plant genomics, metabolic profiling has become a valuable tool with which to assess the effect of genetic and/or environmental factors on plant development. This paper reports the first application of metabolic profiling on differentiating xylem tissue of loblolly pine. A protocol is presented for the analysis of loblolly pine xylem tissue. The effects of sample preparation, extraction, and derivatization on the corresponding metabolite profiles and yields have been investigated and are reported. Gas chromatography-mass spectroscopy has been used to quantify >60 polar and lipophilic metabolites from wood-forming tissue. It was possible to assign chemical structures to approximately half of these compounds. Comparison of six loblolly pine genotypes, three high cellulose (50-52%) and three medium (45-48%) cellulose, showed distinct metabolic profiles. Principal component analysis enabled the assignment of metabolic phenotypes using these large data sets. Metabolic phenotype clustering occurred in which the three high-cellulose genotypes were segregated from the medium-cellulose genotypes. These results demonstrate the use of metabolic profiling for the study of wood-forming tissue and as a tool in functional genomics.

Cellulose↗

PubMed2004

The effect of reduced glutathione on morphology and gene expression of white spruce (Picea glauca) somatic embryos.

Inclusions of reduced glutathione (GSH) in the maturation medium increased the conversion frequency of white spruce somatic embryos without the need of a partial drying treatment (PDT). This beneficial effect was the result of major alterations in morphology and gene expression during the maturation period. Compared with control embryos, GSH-treated embryos showed a differential accumulation of storage products, i.e. preferential deposition of starch, the reduced formation of protein bodies, and increased vacuolation of cells. These morphological changes correlated with extensive alterations of gene expression occurring throughout the maturation period. The transcript profiles of stage-specific embryos matured with or without GSH were analysed using a DNA microarray containing 2 178 cDNAs from loblolly pine (Pinus taeda). The efficiency of heterologous hybridization between spruce and pine species on microarrays has previously been documented. The results indicate that several genes involved in a variety of signal regulatory pathways were differentially expressed in developing GSH- treated embryos. The transcript levels of many genes involved in carbohydrate metabolism and protein synthesis were altered by the presence of GSH and denoted differences in physiology between treatments. Extensive changes in the expression of genes participating in hormone synthesis, nucleotide metabolism, and meristem formation were also observed and related to the post-embryonic performance of the embryos.

Desiccation↗

PubMed2004

An auxin-inducible gene from loblolly pine (Pinus taeda L.) is differentially expressed in mature and juvenile-phase shoots and encodes a putative transmembrane protein.

We have isolated a gene from loblolly pine, 5NG4, that is highly and specifically induced by auxin in juvenile loblolly pine shoots prior to adventitious root formation, but substantially down-regulated in physiologically mature shoots that are adventitious rooting incompetent. 5NG4 was highly auxin-induced in roots, stems and hypocotyls, organs that can form either lateral or adventitious roots following an auxin treatment, but was not induced to the same level in needles and cotyledons, organs that do not form roots. The deduced amino acid sequence shows homology to the MtN21 nodulin gene from Medicago truncatula. The expression pattern of 5NG4 and its homology to a protein from Medicago involved in a root-related process suggest a possible role for this gene in adventitious root formation. Homology searches also identified similar proteins in Arabidopsis thaliana and Oryza sativa. High conservation across these evolutionarily distant species suggests essential functions in plant growth and development. A 38-member family of genes homologous to 5NG4 was identified in the A. thaliana genome. The physiological significance of this redundancy is most likely associated with functional divergence and/or expression specificity of the different family members. The exact biochemical function of the gene is still unknown, but sequence and structure predictions and 5NG4::GFP fusion protein localizations indicate it is a transmembrane protein with a possible transport function.

Amino Acid Sequence↗

PubMed2004

Variation in transcript abundance during somatic embryogenesis in gymnosperms.

Somatic embryogenesis of Norway spruce (Picea abies L.) is a versatile model system to study molecular mechanisms regulating embryo development because it proceeds through defined developmental stages corresponding to specific culture treatments. Normal embryonic development involves early differentiation of proembryogenic masses (PEMs) into somatic embryos, followed by early and late embryogeny leading to the formation of mature cotyledonary embryos. In some cell lines there is a developmental arrest at the PEM-somatic embryo transition. To learn more about the molecular mechanisms regulating embryogenesis, we compared the transcript profiles of two normal lines and one developmentally arrested line. Ribonucleic acid, extracted from these cell lines at successive developmental stages, was analyzed on DNA microarrays containing 2178 expressed sequence tags (ESTs) (corresponding to 2110 unique cDNAs) from loblolly pine (Pinus taeda L.). Hybridization between spruce and pine species on microarrays has been shown to be effective (van Zyl et al. 2002, Stasolla et al. 2003). In contrast to the developmentally arrested line, the early phases of normal embryo development are characterized by a precise pattern of gene expression, i.e., repression followed by induction. Comparison of transcript levels between successive stages of embryogenesis allowed us to identify several genes that showed unique expression responses during normal development. Several of these genes encode proteins involved in detoxification processes, methionine synthesis and utilization, and carbohydrate metabolism. The potential role of these genes in embryo development is discussed.

Cycadopsida↗

PubMed2004

Genetics of postzygotic isolation in Eucalyptus: whole-genome analysis of barriers to introgression in a wide interspecific cross of Eucalyptus grandis and E. globulus.

The genetic architecture of hybrid fitness characters can provide valuable insights into the nature and evolution of postzygotic reproductive barriers in diverged species. We determined the genome-wide distribution of barriers to introgression in an F(1) hybrid of two Eucalyptus tree species, Eucalyptus grandis (W. Hill ex Maiden.) and E. globulus (Labill.). Two interspecific backcross families (N = 186) were used to construct comparative, single-tree, genetic linkage maps of an F(1) hybrid individual and two backcross parents. A total of 1354 testcross AFLP marker loci were evaluated in the three parental maps and a substantial proportion (27.7% average) exhibited transmission ratio distortion (alpha = 0.05). The distorted markers were located in distinct regions of the parental maps and marker alleles within each region were all biased toward either of the two parental species. We used a Bayesian approach to estimate the position and effect of transmission ratio distorting loci (TRDLs) in the distorted regions of each parental linkage map. The relative viability of TRDL alleles ranged from 0.20 to 0.72. Contrary to expectation, heterospecific (donor) alleles of TRDLs were favored as often as recurrent alleles in both backcrosses, suggesting that positive and negative heterospecific interactions affect introgression rates in this wide interspecific pedigree.

Alleles↗

PubMed2003

NMR analysis of lignins in CAD-deficient plants. Part 1. Incorporation of hydroxycinnamaldehydes and hydroxybenzaldehydes into lignins.

Peroxidase/H2O2-mediated radical coupling of 4-hydroxycinnamaldehydes produces 8-O-4-, 8-5-, and 8-8-coupled dehydrodimers as has been documented earlier, as well as the 5-5-coupled dehydrodimer. The 8-5-dehydrodimer is however produced kinetically in its cyclic phenylcoumaran form at neutral pH. Synthetic polymers produced from mixtures of hydroxycinnamaldehydes and normal monolignols provide the next level of complexity. Spectral data from dimers, oligomers, and synthetic polymers have allowed a more substantive assignment of aldehyde components in lignins isolated from a CAD-deficient pine mutant and an antisense-CAD-downregulated transgenic tobacco. CAD-deficient pine lignin shows enhanced levels of the typical benzaldehyde and cinnamaldehyde end-groups, along with evidence for two types of 8-O-4-coupled coniferaldehyde units. The CAD-downregulated tobacco also has higher levels of hydroxycinnamaldehyde and hydroxybenzaldehyde (mainly syringaldehyde) incorporation, but the analogous two types of 8-O-4-coupled products are the dominant features. 8-8-Coupled units are also clearly evident. There is clear evidence for coupling of hydroxycinnamaldehydes to each other and then incorporation into the lignin, as well as for the incorporation of hydroxycinnamaldehyde monomers into the growing lignin polymer. Coniferaldehyde and sinapaldehyde (as well as vanillin and syringaldehyde) co-polymerize with the traditional monolignols into lignins and do so at enhanced levels when CAD-deficiency has an impact on the normal monolignol production. The implication is that, particularly in angiosperms, the aldehydes behave like the traditional monolignols and should probably be regarded as authentic lignin monomers in normal and CAD-deficient plants.

Alcohol Oxidoreductases↗

PubMed2003

Up, down and up again is a signature global gene expression pattern at the beginning of gymnosperm embryogenesis.

Somatic embryogenesis of a gymnosperm, Picea abies, represents a sequence of specifically regulated developmental stages including proembryogenic mass (PEM), PEM-to-embryo transition, and early and late embryogeny. Here, we report cDNA array analysis of expression patterns of 373 genes in the beginning of P. abies embryo development. The analysis revealed a group of 107 genes (29% of arrayed cDNAs) which were upregulated upon PEM-to-embryo transition, then downregulated during early embryogeny and finally upregulated again at the beginning of late embryogeny. This major gene expression pattern was abrogated in a developmentally arrested cell line that is unable to pass through the PEM-to-embryo transition. Thirty-five genes (9.4% of arrayed cDNAs) were found to be differentially expressed during normal embryonic pattern formation. Among them, 22 genes (5.9% of arrayed cDNAs) were directly associated with embryo pattern formation and can be considered as marker genes for early stages of P. abies embryogenesis. The majority of the marker genes encode for proteins involved in translation and posttranslational modification. Among them, 18 genes displayed the major expression pattern.

Gene Expression↗

PubMed2003

An arabinogalactan protein associated with secondary cell wall formation in differentiating xylem of loblolly pine.

Arabinogalactan proteins (AGPs) are abundant plant proteoglycans implicated in plant growth and development. Here, we report the genetic characterization, partial purification and immunolocalization of a classical AGP (PtaAGP6, accession number AF101785) in loblolly pine (Pinus taeda L.). A PtaAGP6 full-length cDNA clone was expressed in bacteria. PtaAGP6 resembles tomato LeAGP-1 and Arabidopsis AtAGP17-19 in that they all possess a subdomain composed of basic amino acids. The accessibility of this domain in the glycoprotein makes it possible to label the PtaAGP6 epitopes on the cell surface or in the cell wall with polyclonal antibodies raised against this subdomain. The antibodies recognize the peptide of the basic subdomain and bind to the intact protein molecule. A soluble protein-containing fraction was purified from the differentiating xylem of pine trees by using beta-glucosyl Yariv reagent (beta-glcY) and was recognized by antibodies against the basic subdomain. Immunolocalization studies showed that the PtaAGP6 epitopes are restricted to a file of cells that just precede secondary cell wall thickening, suggesting roles in xylem differentiation and wood formation. The location of apparent labeling of the PtaAGP6 epitopes is separated from the location of lignin deposition. Multiple single nucleotide polymorphisms (SNPs) were detected in EST variants. Denaturing HPLC analysis of PCR products suggests that PtaAGP6 is encoded by a single gene. Mobility variation in denaturing gel electrophoresis was used to map PtaAGP6 SNPs to a site on linkage group 5.

Amino Acid Sequence↗

PubMed2003

Characterisation of Pt MYB1, an R2R3-MYB from pine xylem.

A cDNA encoding a member of the R2R3-MYB family of transcription factors was cloned from a library constructed from differentiating Pinus taeda (loblolly pine) xylem RNA. This MYB family member, Pinus taeda MYB1 (PtMYB1), was most abundantly expressed in differentiating xylem, as assessed by both ribonuclease protection assays, and by northern blot analysis with poly(A)-enriched RNA. Similar to other plant R2R3-MYB family members, recombinant Pt MYB1 protein was able to bind to AC elements in electrophoretic mobility shift assays (EMSAs). AC elements are DNA motifs rich in adenosine and cytosine that have been implicated in the xylem-localised regulation of genes encoding lignin biosynthetic enzymes. Pt MYB1 not only bound to AC elements, but was also able to induce AC-element-dependent shifts in the electrophoretic mobility of a plant promoter that contains three AC elements, the minimal PHENYLALANINE AMMONIA-LYASE 2 (PAL2) promoter from Phaseolus vulgaris. Transcriptional activation assays conducted using yeast showed that Pt MYB1 also activated transcription, and that it did so in an AC-element-dependent fashion. Pt MYB1 also activated transcription from the minimal PAL2 promoter in plant cells in an AC-element-dependent fashion, as revealed by transient transcriptional activation assays with microprojectile-bombarded tobacco NT-1 cells. Taken together, these finding are consistent with the hypothesis that Pt MYB1 may regulate transcription from cis -acting AC elements in pine xylem.

Adenosine Triphosphate↗

PubMed2003

Characterisation of a pine MYB that regulates lignification.

A member of the R2R3-MYB family of transcription factors was cloned from a cDNA library constructed from RNA isolated from differentiating pine xylem. This MYB, Pinus taeda MYB4 (PtMYB4), is expressed in cells undergoing lignification, as revealed by in situ RT-PCR. Electrophoretic mobility shift assays (EMSAs) showed that recombinant PtMYB4 protein is able to bind to DNA motifs known as AC elements. AC elements are ubiquitous in the promoters of genes encoding lignin biosynthetic enzymes. Transcriptional activation assays using yeast showed that PtMYB4 could activate transcription in an AC-element-dependent fashion. Overexpression of PtMYB4 in transgenic tobacco plants altered the accumulation of transcripts corresponding to genes encoding lignin biosynthetic enzymes. Lignin deposition increased in transgenic tobacco plants that overexpressed PtMYB4, and extended to cell types that do not normally lignify. Taken together, these findings are consistent with the hypothesis that PtMYB4 is sufficient to induce lignification, and that it may play this role during wood formation in pine.

Amino Acid Sequence↗

PubMed2003

Photosynthetic acclimation is reflected in specific patterns of gene expression in drought-stressed loblolly pine.

Because the product of a single gene can influence many aspects of plant growth and development, it is necessary to understand how gene products act in concert and upon each other to effect adaptive changes to stressful conditions. We conducted experiments to improve our understanding of the responses of loblolly pine (Pinus taeda) to drought stress. Water was withheld from rooted plantlets of to a measured water potential of -1 MPa for mild stress and -1.5 MPa for severe stress. Net photosynthesis was measured for each level of stress. RNA was isolated from needles and used in hybridizations against a microarray consisting of 2173 cDNA clones from five pine expressed sequence tag libraries. Gene expression was estimated using a two-stage mixed linear model. Subsequently, data mining via inductive logic programming identified rules (relationships) among gene expression, treatments, and functional categories. Changes in RNA transcript profiles of loblolly pine due to drought stress were correlated with physiological data reflecting photosynthetic acclimation to mild stress or photosynthetic failure during severe stress. Analysis of transcript profiles indicated that there are distinct patterns of expression related to the two levels of stress. Genes encoding heat shock proteins, late embryogenic-abundant proteins, enzymes from the aromatic acid and flavonoid biosynthetic pathways, and from carbon metabolism showed distinctive responses associated with acclimation. Five genes shown to have different transcript levels in response to either mild or severe stress were chosen for further analysis using real-time polymerase chain reaction. The real-time polymerase chain reaction results were in good agreement with those obtained on microarrays.

Acclimatization↗