Specific interactions with TBP and TFIIB in vitro suggest that 14-3-3 proteins may participate in the regulation of transcription when part of a DNA binding complex.
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The 14-3-3 family of multifunctional proteins is highly conserved among animals, plants, and yeast. Several studies have shown that these proteins are associated with a G-box DNA binding complex and are present in the nucleus in several plant and animal species. In this study, 14-3-3 proteins are shown to bind the TATA box binding protein (TBP), transcription factor IIB (TFIIB), and the human TBP-associated factor hTAF(II)32 in vitro but not hTAF(II)55. The interactions with TBP and TFIIB were highly specific, requiring amino acid residues in the box 1 domain of the 14-3-3 protein. These interactions do not require formation of the 14-3-3 dimer and are not dependent on known 14-3-3 recognition motifs containing phosphoserine. The 14-3-3-TFIIB interaction appears to occur within the same domain of TFIIB that binds the human herpes simplex virus transcriptional activator VP16, because VP16 and 14-3-3 were able to compete for interaction with TFIIB in vitro. In a plant transient expression system, 14-3-3 was able to activate GAL4-dependent beta-glucuronidase reporter gene expression at low levels when translationally fused with the GAL4 DNA binding domain. The in vitro binding with general transcription factors TBP and TFIIB together with its nuclear location provide evidence supporting a role for 14-3-3 proteins as transcriptional activators or coactivators when part of a DNA binding complex. (+info)
Effect of wheat and corn variety on fiber digestion in beef steers fed high-grain diets.
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Six Salers steers, fitted with ruminal and duodenal cannulas, were used in a double 3x3 Latin square design to assess the depressive effect of the nature of wheat, flint corn, and dent corn on fiber digestion in animals fed high-concentrate diets, and to determine the mechanisms involved in these negative digestive effects. Diets were balanced to be equal in starch content (47.7+/-2.3%). The three cereals were characterized by ruminal starch digestibilities of 86.6, 60.8, and 34.8% for the wheat, dent corn, and flint corn, respectively. Ruminal digestion of NDF was lower with wheat- than with corn-based diets (49.4 vs. 55.2%; P<.001), and with dent corn than with flint corn (53 vs. 57.3%; P<.01). Degradability of hay in nylon bags was not affected by the grain source in the diet (P>.1). The mean retention time of forage particles in the rumen was similar between wheat and corn diets (P>.1), but it was lower for steers fed dent corn than for those fed flint corn (P<.05). Most fibrolytic activities of the solid-associated microorganisms were lower (P<.05) in animals fed wheat than in those fed corn. Differences in fibrolytic activities of the solid-associated microorganisms between the two corn genotypes were not statistically significant (P>.1), but activities of all fibrolytic enzymes were lower (P<.05) with the dent than with the flint corn diet. Protozoal number in ruminal fluid was lower in animals receiving wheat than in those fed corn (177 vs. 789x10(3)/mL; P<.001) and was related to the high ruminal acidity (P<.01) of the wheat diet. Large modifications in the rumen microbial ecosystem between the two corn genotypes were not visible in protozoal numbers or pH. Total-tract digestion of NDF was the same for wheat and for corn diets, averaging 55% for the three diets. A postruminal compensation of NDF digestion (14% of the total tract NDF digestion) seemed to occur with the wheat diet. The lack of any postruminal NDF digestion (0%) with the two corn diets may suggest negative digestive interactions in the hindgut similar to those in the rumen. (+info)
Effects of laidlomycin propionate and monensin on the in vitro mixed ruminal microorganism fermentation.
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The objective of this study was to compare the effects of laidlomycin propionate and monensin on the in vitro fermentation of ground corn, Trypticase, or alfalfa hay by mixed ruminal microorganisms. Ruminal fluid was collected from two steers fed 9.27 kg DM of a high-concentrate (62.2% ground corn and 17.4% cottonseed hulls) diet per day and composited. In the first study, no ionophore was included in the diet; the diet in the second study contained 11.1 g of laidlomycin propionate per ton of feed. The animals were allowed an adjustment period of 14 d for each dietary treatment before samples were collected. When ruminal fluid from unadapted animals was used, both monensin and laidlomycin propionate decreased (P<.05) CH4 concentration and the acetate:propionate ratio with ground corn and alfalfa hay. Monensin reduced (P<.05) in vitro dry matter disappearance of alfalfa and increased (P<.05) final pH in the ground corn and alfalfa hay fermentations. Both laidlomycin propionate and monensin decreased (P<.05) concentrations of acetate, propionate, isobutyrate, isovalerate, CH4, and NH3 in Trypticase fermentations. When ruminal fluid from adapted animals was used, both ionophores still reduced the concentrations of most fermentation products. However, there was generally less inhibition compared with fermentations inoculated with unadapted mixed ruminal microorganisms. In the presence of 5 mM maltose, mixed ruminal bacteria produced high concentrations (10 to 11 mM) of lactate, and addition of both ionophores to these fermentations was effective in reducing (P<.05) lactate production. In conclusion, laidlomycin propionate alters the mixed ruminal microorganism fermentation in a manner similar to monensin, but, at the concentrations used in this study, monensin seemed to be a more potent inhibitor. (+info)
Sugars modulate an unusual mode of control of the cell-wall invertase gene (Incw1) through its 3' untranslated region in a cell suspension culture of maize.
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We show here that a cell-wall invertase encoded by the Incw1 gene is regulated at both the transcriptional and posttranscriptional levels by sugars in a heterotrophic cell suspension culture of maize. The Incw1 gene encoded two transcripts: Incw1-S (small) and Incw1-L (large); the size variation was attributable to different lengths in the 3' untranslated region. Both metabolizable and nonmetabolizable sugars induced Incw1-L RNA apparently by default. However, only the metabolizable sugars, sucrose and D-glucose, were associated with the increased steady-state abundance of Incw1-S RNA, the concomitant increased levels of INCW1 protein and enzyme activity, and the downstream metabolic repression of the sucrose synthase gene, Sh1. Conversely, nonmetabolizable sugars, including the two glucose analogs 3-O-methylglucose and 2-deoxyglucose, induced greater steady-state levels of the Incw1-L RNA, but this increase did not lead to either an increase in the levels of the INCW1 protein/enzyme activity or the repression of the Sh1 gene. We conclude that sugar sensing and the induction of the Incw1 gene is independent of the hexokinase pathway. More importantly, our results also suggest that the 3' untranslated region of the Incw1 gene acts as a regulatory sensor of carbon starvation and may constitute a link between sink metabolism and cellular translation in plants. (+info)
Meiotic drive of chromosomal knobs reshaped the maize genome.
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Meiotic drive is the subversion of meiosis so that particular genes are preferentially transmitted to the progeny. Meiotic drive generally causes the preferential segregation of small regions of the genome; however, in maize we propose that meiotic drive is responsible for the evolution of large repetitive DNA arrays on all chromosomes. A maize meiotic drive locus found on an uncommon form of chromosome 10 [abnormal 10 (Ab10)] may be largely responsible for the evolution of heterochromatic chromosomal knobs, which can confer meiotic drive potential to every maize chromosome. Simulations were used to illustrate the dynamics of this meiotic drive model and suggest knobs might be deleterious in the absence of Ab10. Chromosomal knob data from maize's wild relatives (Zea mays ssp. parviglumis and mexicana) and phylogenetic comparisons demonstrated that the evolution of knob size, frequency, and chromosomal position agreed with the meiotic drive hypothesis. Knob chromosomal position was incompatible with the hypothesis that knob repetitive DNA is neutral or slightly deleterious to the genome. We also show that environmental factors and transposition may play a role in the evolution of knobs. Because knobs occur at multiple locations on all maize chromosomes, the combined effects of meiotic drive and genetic linkage may have reshaped genetic diversity throughout the maize genome in response to the presence of Ab10. Meiotic drive may be a major force of genome evolution, allowing revolutionary changes in genome structure and diversity over short evolutionary periods. (+info)
Maize R2R3 Myb genes: Sequence analysis reveals amplification in the higher plants.
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Transcription factors containing the Myb-homologous DNA-binding domain are widely found in eukaryotes. In plants, R2R3 Myb-domain proteins are involved in the control of form and metabolism. The Arabidopsis genome harbors >100 R2R3 Myb genes, but few have been found in monocots, animals, and fungi. Using RT-PCR from different maize organs, we cloned 480 fragments corresponding to a 42-44 residue-long sequence spanning the region between the conserved DNA-recognition helices (Myb(BRH)) of R2R3 Myb domains. We determined that maize expresses >80 different R2R3 Myb genes, and evolutionary distances among maize Myb(BRH) sequences indicate that most of the amplification of the R2R3 Myb gene family occurred after the origin of land plants but prior to the separation of monocots and dicots. In addition, evidence is provided for the very recent duplication of particular classes of R2R3 Myb genes in the grasses. Together, these findings render a novel line of evidence for the amplification of the R2R3 Myb gene family in the early history of land plants and suggest that maize provides a possible model system to examine the hypothesis that the expansion of Myb genes is associated with the regulation of novel plant cellular functions. (+info)
Inferences on the genome structure of progenitor maize through comparative analysis of rice, maize and the domesticated panicoids.
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Corn and rice genetic linkage map alignments were extended and refined by the addition of 262 new, reciprocally mapped maize cDNA loci. Twenty chromosomal rearrangements were identified in maize relative to rice and these included telomeric fusions between rice linkage groups, nested insertion of rice linkage groups, intrachromosomal inversions, and a nonreciprocal translocation. Maize genome evolution was inferred relative to other species within the Panicoideae and a progenitor maize genome with eight linkage groups was proposed. Conservation of composite linkage groups indicates that the tetrasomic state arose during maize evolution either from duplication of one progenitor corn genome (autoploidy) or from a cross between species that shared the composite linkages observed in modern maize (alloploidy). New evidence of a quadruplicated homeologous segment on maize chromosomes 2 and 10, and 3 and 4, corresponded to the internally duplicated region on rice chromosomes 11 and 12 and suggested that this duplication in the rice genome predated the divergence of the Panicoideae and Oryzoideae subfamilies. Charting of the macroevolutionary steps leading to the modern maize genome clarifies the interpretation of intercladal comparative maps and facilitates alignments and genomic cross-referencing of genes and phenotypes among grass family members. (+info)
Essential arginine residues in maize starch synthase IIa are involved in both ADP-glucose and primer binding.
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The arginine-specific reagent phenylglyoxal inactivated the activity of maize starch synthase IIa (SSIIa), due to the modification of at least one arginine residue out of a possible 42. The addition of ADPGlc completely protected SSIIa from the inactivation, indicating that arginine may be involved in the interaction of this anionic substrate with SSIIa. However, site-directed mutagenesis of the conserved Arg-214 in SSIIa showed that this amino acid is important for apparent affinity of SSIIa for its primer (amylopectin and glycogen), as evidenced by a marked increase in the K(m) for primer upon substitution of this amino acid with no concomitant change in V(max), K(m) for ADPGlc, or secondary structure. Therefore, Arg-214 of SSIIa appears to play a role in its primer binding. (+info)