Biomedical subjects

Robert B Meeley

Publications and source records attributed to Robert B Meeley.

15 recordsLinked to original sources

PubMed2005

Characterization of two GL8 paralogs reveals that the 3-ketoacyl reductase component of fatty acid elongase is essential for maize (Zea mays L.) development.

Prior analyses established that the maize (Zea mays L.) gl8a gene encodes 3-ketoacyl reductase, a component of the fatty acid elongase required for the biosynthesis of very long chain fatty acids (VLCFAs). A paralogous gene, gl8b, has been identified that is 96% identical to gl8a. The gl8a and gl8b genes map to syntenic chromosomal regions, have similar, but not identical, expression patterns, and encode proteins that are 97% identical. Both of these genes are required for the normal accumulation of cuticular waxes on seedling leaves. The chemical composition of the cuticular waxes from gl8a and gl8b mutants indicates that these genes have at least overlapping, if not redundant, functions in cuticular wax biosynthesis. Although gl8a and gl8b double mutant kernels have endosperms that cannot be distinguished from wild-type siblings, these kernels are non-viable because their embryos fail to undergo normal development. Double mutant kernels accumulate substantially reduced levels of VLCFAs. VLCFAs are components of a variety of compounds, for example, cuticular waxes, suberin, and sphingolipids. Consistent with their essential nature in yeast, the accumulation of the ceramide moiety of sphingolipids is substantially reduced and their fatty acid composition altered in gl8a and gl8b double mutant kernels relative to wild-type kernels. Hence, we hypothesize that sphingolipids or other VLCFA-containing compounds are essential for normal embryo development.

3-Oxoacyl-(Acyl-Carrier-Protein) Reductase↗

PubMed2004

The role of barren stalk1 in the architecture of maize.

The architecture of higher plants is established through the activity of lateral meristems--small groups of stem cells formed during vegetative and reproductive development. Lateral meristems generate branches and inflorescence structures, which define the overall form of a plant, and are largely responsible for the evolution of different plant architectures. Here, we report the isolation of the barren stalk1 gene, which encodes a non-canonical basic helix-loop-helix protein required for the initiation of all aerial lateral meristems in maize. barren stalk1 represents one of the earliest genes involved in the patterning of maize inflorescences, and, together with the teosinte branched1 gene, it regulates vegetative lateral meristem development. The architecture of maize has been a major target of selection for early agriculturalists and modern farmers, because it influences harvesting, breeding strategies and mechanization. By sampling nucleotide diversity in the barren stalk1 region, we show that two haplotypes entered the maize gene pool from its wild progenitor, teosinte, and that only one was incorporated throughout modern inbreds, suggesting that barren stalk1 was selected for agronomic purposes.

Amino Acid Sequence↗

PubMed2004

Regulation and a conserved intron sequence of liguleless3/4 knox class-I homeobox genes in grasses.

The nine class-I maize (Zea mays L.) knox genes are putative transcription factors normally expressed in shoot apices, but not in leaves. knotted1 (kn1) seems to function in shoot apical meristem maintenance, and rough sheath1 (rs1)-like genes may act in internode elongation. The function of liguleless3 (lg3)-type genes is still unknown. Here, we characterized lg3 as well as the two most closely related genes liguleless4a (lg4a, formerly knox11) and liguleless4b (lg4b, formerly knox5). We termed this subclass of knox genes lg3/4 genes. We studied the expression patterns of lg3/4 genes and compared their sequences. We obtained knockout mutants of lg3 by finding Mu transposon insertions into exons. Our results show that lg3 was not essential for plant development, and that lg4a and lg4b were likely to encode the redundant function. In addition, lg4a but not lg4b was ectopically expressed in the Lg4-O mutant, suggesting that this mutant was affected at the lg4a locus. We found that the lg3 gene was unique among knox genes as it was co-induced in the leaves of leaf mutants that ectopically expressed knox genes in the leaves. The leaf phenotype expressed in the dominant Rs1-O mutant was not altered when lg3 function was removed using the knockout. Genomic sequence comparisons of lg3, lg4a and lg4b from maize and the two homologous genes, osh6 and osh71, from rice revealed a 14-bp phylogenetic footprint in intron II. This sequence was conserved in nucleotide composition, position and polarity in the lg3/4 genes of divergent grasses representing six Gramineae subfamilies. In an independent experiment, this same conserved sequence was found in a yeast reverse one-hybrid screen for putative binding sites of the LG3 homeodomain protein. Distribution of this 14-bp sequence was examined within the public rice database. The possible function of this sequence in regulation of lg3/4 genes is discussed.

Amino Acid Sequence↗

PubMed2004

Coordination of meiotic recombination, pairing, and synapsis by PHS1.

Pairing, synapsis, and recombination are prerequisites for accurate chromosome segregation in meiosis. The phs1 gene in maize is required for pairing to occur between homologous chromosomes. In the phs1 mutant, homologous chromosome synapsis is completely replaced by synapsis between nonhomologous partners. The phs1 gene is also required for installation of the meiotic recombination machinery on chromosomes, as the mutant almost completely lacks chromosomal foci of the recombination protein RAD51. Thus, in the phs1 mutant, synapsis is uncoupled from recombination and pairing. The protein encoded by the phs1 gene likely acts in a multistep process to coordinate pairing, recombination, and synapsis.

Amino Acid Sequence↗

PubMed2004

ACC synthase expression regulates leaf performance and drought tolerance in maize.

Ethylene regulates entry into several types of plant developmental cell death and senescence programs besides mediating plant responses to biotic and abiotic stress. The response of cereals to conditions of drought includes loss of leaf function and premature onset of senescence in older leaves. In this study, ACC synthase (ACS) mutants, affecting the first step in ethylene biosynthesis, were isolated in maize and their effect on leaf function examined. Loss of ZmACS6 expression resulted in delayed leaf senescence under normal growth conditions and inhibited drought-induced senescence. Zmacs6 leaves continued to be photosynthetically active under both conditions indicating that leaf function was maintained. The delayed senescence phenotype associated with loss of ZmACS6 expression was complemented by exogenous ACC. Surprisingly, elevated levels of foliar chlorophyll, Rubisco, and soluble protein as well as improved leaf performance was observed for all Zmasc6 leaves, including young and fully expanded leaves which were far from initiating senescence. These observations suggest that ethylene may serve to regulate leaf performance throughout its lifespan as well as to determine the onset of natural senescence and mediate drought-induced senescence.

Chlorophyll↗

PubMed2003

Conserved subgroups and developmental regulation in the monocot rop gene family.

Rop small GTPases are plant-specific signaling proteins with roles in pollen and vegetative cell growth, abscisic acid signal transduction, stress responses, and pathogen resistance. We have characterized the rop family in the monocots maize (Zea mays) and rice (Oryza sativa). The maize genome contains at least nine expressed rops, and the fully sequenced rice genome has seven. Based on phylogenetic analyses of all available Rops, the family can be subdivided into four groups that predate the divergence of monocots and dicots; at least three have been maintained in both lineages. However, the Rop family has evolved differently in the two lineages, with each exhibiting apparent expansion in different groups. These analyses, together with genetic mapping and identification of conserved non-coding sequences, predict orthology for specific rice and maize rops. We also identified consensus protein sequence elements specific to each Rop group. A survey of ROP-mRNA expression in maize, based on multiplex reverse transcriptase-polymerase chain reaction and a massively parallel signature sequencing database, showed significant spatial and temporal overlap of the nine transcripts, with high levels of all nine in tissues in which cells are actively dividing and expanding. However, only a subset of rops was highly expressed in mature leaves and pollen. Intriguingly, the grouping of maize rops based on hierarchical clustering of expression profiles was remarkably similar to that obtained by phylogenetic analysis. We hypothesize that the Rop groups represent classes with distinct functions, which are specified by the unique protein sequence elements in each group and by their distinct expression patterns.

Amino Acid Sequence↗

PubMed2003

The maize disorganized aleurone layer 1 and 2 ( dil1, dil2) mutants lack control of the mitotic division plane in the aleurone layer of developing endosperm.

The maize ( Zea mays L.) endosperm consists of an epidermal like layer of isodiametric aleurone cells surrounding a central body of starchy endosperm cells. In disorgal1 ( dil1) and disorgal2 ( dil2) mutants the control of the mitotic division plane is relaxed or missing, resulting in mature grains with disorganized aleurone layers. In addition to orientation of the division plane, both the shape and size of the aleurone cells are affected, and often more than one layer of aleurone cells is present. Homozygous dil1 and dil2 grains are shrunken due to reduced accumulation of starchy endosperm and premature developmental arrest of the embryo, and mature mutant grains germinate at a very low rate and fail to develop into plants. However, homozygous mutant plants can be obtained through embryo rescue, revealing that both mutants have an irregular leaf epidermis as well as roots with a strongly reduced number of root hairs and aberrant root hair morphology. Our results suggest the presence of common regulatory mechanisms for the control of cell division orientation in the aleurone and plant epidermis.

Alleles↗

PubMed2003

The Nitrilase ZmNIT2 converts indole-3-acetonitrile to indole-3-acetic acid.

We isolated two nitrilase genes, ZmNIT1 and ZmNIT2, from maize (Zea mays) that share 75% sequence identity on the amino acid level. Despite the relatively high homology to Arabidopsis NIT4, ZmNIT2 shows no activity toward beta-cyano-alanine, the substrate of Arabidopsis NIT4, but instead hydrolyzes indole-3-acetonitrile (IAN) to indole-3-acetic acid (IAA). ZmNIT2 converts IAN to IAA at least seven to 20 times more efficiently than AtNIT1/2/3. Quantitative real-time polymerase chain reaction revealed the gene expression of both nitrilases in maize kernels where high concentrations of IAA are synthesized tryptophan dependently. Nitrilase protein and endogenous nitrilase activity are present in maize kernels together with the substrate IAN. These results suggest a role for ZmNIT2 in auxin biosynthesis.

Aminohydrolases↗

PubMed2003

sal1 determines the number of aleurone cell layers in maize endosperm and encodes a class E vacuolar sorting protein.

A microscopy-based screen of a large collection of maize Mutator (Mu) transposon lines identified the supernumerary aleurone layers 1-1 (sal1-1) mutant line carrying up to seven layers of aleurone cells in defective kernel endosperm compared with only a single layer in wild-type grains. Normal, well filled endosperm that is homozygous for the sal1-1 mutant allele contains two to three layers of aleurone cells. Cloning of the sal1 gene was accomplished by using Mu tagging, and the identity of the cloned gene was confirmed by isolating an independent sal1-2 allele by reverse genetics. Homozygous sal1-2 endosperm has two to three layers of aleurone cells in normal, well filled grains. In situ hybridization experiments reveal that the sal1 gene is ubiquitously expressed in vegetative as well as zygotic grain tissues, with no difference being detected between aleurone cells and starchy endosperm cells. Northern blot analysis failed to detect the sal1-2 transcript in leaves of homozygous plants, suggesting that the allele is a true sal1 knockout allele. The sal1 gene encodes a homologue of the human Chmp1 gene, a member of the conserved family of the class E vacuolar protein sorting genes implicated in membrane vesicle trafficking. In mammals, CHMP1 functions in the pathway targeting plasma membrane receptors and ligands to lysosomes for proteolytic degradation. Possible roles for the function of the sal1 gene in aleurone signaling, including a defect in endosome trafficking, are discussed.

Amino Acid Sequence↗

PubMed2003

A 2-oxoglutarate-dependent dioxygenase is integrated in DIMBOA-biosynthesis.

Benzoxazinoids are secondary metabolites of grasses that function as natural pesticides. While many steps of DIMBOA biosynthesis have been elucidated, the mechanism of the introduction of OCH(3)-group at the C-7 position was unknown. Inhibitor experiments in Triticum aestivum and Zea mays suggest that a 2-oxoglutarate-dependent dioxygenase catalyses the hydroxylation reaction at C-7. Cloning and reverse genetics analysis have identified the Bx6 gene that encodes this enzyme. Bx6 is located in the Bx-gene cluster of maize.

Amino Acid Sequence↗

PubMed2003

Zea mays ZmMybst1 cDNA, encodes a single Myb-repeat protein with the VASHAQKYF motif.

A cDNA clone from a 4 DAP dissected maize embryo sac encoding a novel Zea mays single-repeat Myb protein is reported here. This full-length cDNA contains an ORF of 948 bp. The gene ZmMybst1 contains two introns (1166 and 706 bp) and is a single copy gene. The ZmMybst1 protein shares high sequence identity with the potato Mybst1 protein (58%). Northern blot, RT-PCR and electronic northern analysis shows that ZmMybst1 is expressed in endosperm between 4 and 30 DAP, coinciding with the period of aleurone cell differentiation and development.

Amino Acid Sequence↗

PubMed2003

The maize low-phytic acid mutant lpa2 is caused by mutation in an inositol phosphate kinase gene.

Reduced phytic acid content in seeds is a desired goal for genetic improvement in several crops. Low-phytic acid mutants have been used in genetic breeding, but it is not known what genes are responsible for the low-phytic acid phenotype. Using a reverse genetics approach, we found that the maize (Zea mays) low-phytic acid lpa2 mutant is caused by mutation in an inositol phosphate kinase gene. The maize inositol phosphate kinase (ZmIpk) gene was identified through sequence comparison with human and Arabidopsis Ins(1,3,4)P(3) 5/6-kinase genes. The purified recombinant ZmIpk protein has kinase activity on several inositol polyphosphates, including Ins(1,3,4)P(3), Ins(3,5,6)P(3), Ins(3,4,5,6)P(4), and Ins(1,2,5,6)P(4). The ZmIpk mRNA is expressed in the embryo, the organ where phytic acid accumulates in maize seeds. The ZmIpk Mutator insertion mutants were identified from a Mutator F(2) family. In the ZmIpk Mu insertion mutants, seed phytic acid content is reduced approximately 30%, and inorganic phosphate is increased about 3-fold. The mutants also accumulate myo-inositol and inositol phosphates as in the lpa2 mutant. Allelic tests showed that the ZmIpk Mu insertion mutants are allelic to the lpa2. Southern-blot analysis, cloning, and sequencing of the ZmIpk gene from lpa2 revealed that the lpa2-1 allele is caused by the genomic sequence rearrangement in the ZmIpk locus and the lpa2-2 allele has a nucleotide mutation that generated a stop codon in the N-terminal region of the ZmIpk open reading frame. These results provide evidence that ZmIpk is one of the kinases responsible for phytic acid biosynthesis in developing maize seeds.

Alleles↗

PubMed2003

Insertional inactivation of the methionine s-methyltransferase gene eliminates the s-methylmethionine cycle and increases the methylation ratio.

Methionine (Met) S-methyltransferase (MMT) catalyzes the synthesis of S-methyl-Met (SMM) from Met and S-adenosyl-Met (Ado-Met). SMM can be reconverted to Met by donating a methyl group to homocysteine (homo-Cys), and concurrent operation of this reaction and that mediated by MMT sets up the SMM cycle. SMM has been hypothesized to be essential as a methyl donor or as a transport form of sulfur, and the SMM cycle has been hypothesized to guard against depletion of the free Met pool by excess Ado-Met synthesis or to regulate Ado-Met level and hence the Ado-Met to S-adenosylhomo-Cys ratio (the methylation ratio). To test these hypotheses, we isolated insertional mmt mutants of Arabidopsis and maize (Zea mays). Both mutants lacked the capacity to produce SMM and thus had no SMM cycle. They nevertheless grew and reproduced normally, and the seeds of the Arabidopsis mutant had normal sulfur contents. These findings rule out an indispensable role for SMM as a methyl donor or in sulfur transport. The Arabidopsis mutant had significantly higher Ado-Met and lower S-adenosylhomo-Cys levels than the wild type and consequently had a higher methylation ratio (13.8 versus 9.5). Free Met and thiol pools were unaltered in this mutant, although there were moderate decreases (of 30%-60%) in free serine, threonine, proline, and other amino acids. These data indicate that the SMM cycle contributes to regulation of Ado-Met levels rather than preventing depletion of free Met.

Amino Acids↗

PubMed2003

Duplicate FLORICAULA/LEAFY homologs zfl1 and zfl2 control inflorescence architecture and flower patterning in maize.

The homologous transcription factors FLORICAULA of Antirrhinum and LEAFY of Arabidopsis share conserved roles in flower meristem identity and floral patterning. While roles for FLORICAULA/LEAFY homologs in flower development have been demonstrated in numerous dicots, little is known about the function of these meristem identity genes in the more distantly related flowering plants, the monocots. We used reverse genetics to investigate the role of two duplicate FLORICAULA/LEAFY homologs in maize (Zea mays L. ssp. mays) - a monocot species with dramatically different flower and inflorescence morphology from that of dicot species. Transposon insertions into the maize genes, zfl1 and zfl2, led to a disruption of floral organ identity and patterning, as well as to defects in inflorescence architecture and in the vegetative to reproductive phase transition. Our results demonstrate that these genes share conserved roles with their dicot counterparts in flower and inflorescence patterning. The phenotype of zfl1; zfl2 double mutants suggests that these maize FLORICAULA/LEAFY homologs act as upstream regulators of the ABC floral organ identity genes, and this along with previously published work, indicates that the transcriptional network regulating flower development is at least partially conserved between monocots and dicots. Our data also suggest that the zfl genes may play a novel role in controlling quantitative aspects of inflorescence phyllotaxy in maize, consistent with their candidacy for quantitative trait loci that control differences in inflorescence structure between maize and its progenitor, teosinte.

Amino Acid Sequence↗

PubMed2002

Maize HSP101 plays important roles in both induced and basal thermotolerance and primary root growth.

HSP101 belongs to the ClpB protein subfamily whose members promote the renaturation of protein aggregates and are essential for the induction of thermotolerance. We found that maize HSP101 accumulated in mature kernels in the absence of heat stress. At optimal temperatures, HSP101 disappeared within the first 3 days after imbibition, although its levels increased in response to heat shock. In embryonic cells, HSP101 concentrated in the nucleus and in some nucleoli. Hsp101 maps near the umc132 and npi280 markers on chromosome 6. Five maize hsp101-m-::Mu1 alleles were isolated. Mutants were null for HSP101 and defective in both induced and basal thermotolerance. Moreover, during the first 3 days after imbibition, primary roots grew faster in the mutants at optimal temperature. Thus, HSP101 is a nucleus-localized protein that, in addition to its role in thermotolerance, negatively influences the growth rate of the primary root. HSP101 is dispensable for proper embryo and whole plant development in the absence of heat stress.

Acclimatization↗