Unknown,Transcriptomics,Genomics,Proteomics

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Gene expression non-additivity in immature ears of a heterotic F1 maize hybrid

ABSTRACT: Non-additive gene regulation has been recently suggested as an important factor promoting phenotypic variation and plasticity. In order to obtain a description of gene expression status at an early stage of ear development in a maize (Zea mays L.) F1 hybrid as relative to its parental inbreds, we compared gene expression profiles in immature ears of elite inbred lines B73 and H99 to one of their F1 hybrids (B73xH99) using cDNA microarray technology. Results show several genes expressed at a significantly different level between both inbred lines and their hybrid. In addition, gene expression non-additivity in the hybrid was detected on a broad scale, consisting of both dominance and over-dominance components, indicating that complex non-additive interactions at the molecular level exist in the developing ear of the studied maize hybrid. Non-additively regulated genes belong to a wide range of molecular functions, indicating that several regulatory and metabolic patterns are possibly affected during ear development in the investigated hybrid. We discuss the possibility that observed gene expression non-additivity in immature ear might be an early molecular manifestation of hybrid vigor, the most exploited factor for maize agronomic improvement. We directly contrasted both B73 and H99 inbred lines vs. their F1 heterotic hybrid transcriptomes in immature ear. Our experimental design consisted of 10 cDNA microarray hybridizations, 5 for each combination of hybrid vs. inbred genotypes, involving 20 separate labeling reactions. Labeling dyes were swapped in two of the five replicates for each combination. In each hybridization, control channel was assigned to the F1 hybrid, and differences in transcriptional levels between B73 and H99 were inferred using the hybrid as a common reference sample in an indirect experimental design. To take in account variability in transcript population among individuals, total RNA coming from different isolations, each collected on multiple individuals, were mixed before poly(A+) RNA purification. All hybridizations on microarray slides were then performed using cDNA independently labeled from the poly(A+) RNA purification product for each genotype. Base-two logarithms of expression ratios were subjected to one-class response significance analysis in SAM v. 2.20 software [Tusher et al. 2001]. For each EST the estimates of additive parameter "a" and dominance parameter "d" (middle-parent heterosis) were obtained as a = (L2 – L1)/2 d = (L1 + L2)/2 (where L1 and L2 are mean base-two logarithms of expression ratios of F1 vs. B73 and F1 vs. H99, respectively). Positive values of "a" indicate expression values bigger in B73 than in H99. The dominance/additivity ratio (d/|a|) was also calculated [Falconer 1989]. Evaluation of statistical significance of parameters was done by calculating standard errors of the estimates "a" and "d" as standard errors of linear functions of the means. Significance testing was done correspondingly, using an F-test for linear contrasts. P-values for the families of tests corresponding to each parameter were subjected to global error analyses using a method based on fitting mixture distribution [Allison et al. 2002], allowing to estimate the false discovery rates (FDR) and false negative rates (FNR). Confidence intervals for d/|a| ratios were obtained by Fieller’s method [Piepho and Emrich 2005], allowing to classify the genes into different dominance type classes.

ORGANISM(S): Zea mays

SUBMITTER: Giorgio Pea

PROVIDER: E-GEOD-2771 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Heterosis is the phenomenon whereby the progeny of particular inbred lines have enhanced agronomic performance relative to both parents. Although several hypotheses have been proposed to explain this fundamental biological phenomenon, the responsible molecular mechanisms have not been determined. The maize inbred lines B73 and Mo17 produce a heterotic F1 hybrid. Global patterns of gene expression were compared in seedlings of these three genotypes using a microarray that contains 13,999 cDNAs. Using an estimated 15% false discovery rate as a cut-off, 1,367 ESTs (9.8%) were identified as being significantly differentially expressed among genotypes. All possible modes of gene action were observed, including additivity, high- and low-parent dominance, under-dominance, and over-dominance. The largest proportion of the ESTs (78%, 1,062/1,367) exhibited expression patterns that are not statistically distinguishable from additivity. Even so, 22% of the differentially regulated genes exhibited non-additive modes of gene expression. Classified on the basis of significant pair-wise comparisons of genotype means, 181 of these 305 genes exhibited high-parent dominance and 23 exhibited low-parent dominance. In addition, 44 genes exhibited under- or over-dominant gene action. These findings are consistent with the hypothesis that multiple molecular mechanisms contribute to heterosis, including over-dominance Keywords: Genotype Comparison The inbreds B73 (Schnable Lab Accession #660) and Mo17 (Schnable Lab Accession #2618) used in this study were derived by self-pollination from stocks originally obtained from Don Robertson (Iowa State University) and Mike Lee (Iowa State University), respectively. Mo17 was crossed as a female by B73 to generate the F1. Kernels from three different seed sources (ears) per genotype were used in the experimental design. Prior to microarray analyses genotypes were confirmed by genotyping DNA extracted from each seed source using co-dominant IDP genetic markers that distinguish B73 from Mo17 (Fu et al., in preparation). Ten biological replications of B73, Mo17, and their F1 (Mo17xB73) were grown under highly controlled conditions in a randomized complete block design. For each replication, the B73, Mo17, and F1 samples were hybridized to three two-color cDNA microarrays using a loop design such that each loop included all pairwise comparisons between genotypes. RNA pools for each genotype were alternately labeled providing dye balance within each loop. After hybridization, one biological replicate was removed due to poor quality. The final analysis incorporated 27 microarray slides (3 slides for each of nine high-quality biological replicates).

Project description:Hybrid progeny can enjoy increased fitness and stress tolerance relative to their ancestral species, a phenomenon known as hybrid vigor. Though this phenomenon has been documented throughout the Eukarya, evolution of hybrid populations has yet to be explored experimentally in the lab. To fill this knowledge gap we created a pool of Saccharomyces cerevisiae and S. bayanus homoploid and aneuploid hybrids, and then investigated how selection in the form of incrementally increased temperature or ethanol impacted hybrid genome structure and adaptation. During 500 generations of continuous ammonia-limited, glucose-sufficient culture, temperature was raised from 25C to 46??C. This selection invariably resulted in nearly-complete loss of the S. bayanus genome, although the dynamics of genome loss differed among independent replicates. Temperature-evolved isolates were significantly more thermal tolerant and exhibited greater phenotypic plasticity than parental species and founding hybrids. By contrast, when the same hybrid pool was subjected to increases in exogenous ethanol from 0% to 14%, selection favored euploid S. cerevisiae x S. bayanus hybrids. Ethanol-evolved isolates exhibited significantly greater ethanol tolerance relative only to S. bayanus and one of the founding hybrids tested. Adaptation to thermal and ethanol stress manifested as heritable changes in cell wall structure demonstrated by resistance to zymolyase or micafungin treatment. This is the first study to show experimentally that the fate of interspecific hybrids critically depends on the type of selection they encounter during the course of evolution. Array-CGH was performed on the S. cerevisiae parent strain CEN.PK (GSY2160), the S. bayanus parent strain CBS7001 (GSY2161) and on the F1 interspecific hybrid resulting from mating the 2 parents (GSY2168). Additionally, three rare viable spores obtained after sporulation of the F1 were assayed by array-CGH (F2a, F2b, F2c). A large pool of F2 spores (and probably some number of F1 hybrid cells) were subjected to gradually increasing temperatures, in three independent vessels, with populations sampled at various generation times. Likewise, the same pool was used to found populations in an additional three independent vessels, which were then subjected to gradually increasing ethanol concentrations (at constant temperature). Array-CGH was performed on three different clones from each of the three temperature vessels at the final 500 generation time point (T500 clones). Biological replicates of the T500 clones were performed (T500-new). Two self-self array-CGH hybridization controls were also performed (self-control). Array-CGH was performed on one clone from each of the three ethanol vessels taken at the 400 generation timepoint (EtOH400gen clones).

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Gene expression non-additivity in immature ears of a heterotic F1 maize hybrid

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