| Maize is one of the most productive crops and widely used as genetic model plant all over the world. The plant produces two distinct inflorescences commonly referred to as tassel and ear, which is different from other grasses such as rice and wheat. The tassel arises from the apex of the mature plant, while ears originate from axillary bud apices. One obvious difference in morphology between the two inflorescences is presence (tassel) or absence (ear) of a variable number of long branches originating at the base. The wide range of natural variation among different inbred lines was used to determine quantitative trait loci (QTL) underlying a variety of phenotypes by association mapping, and many genes associated with maize ear development have been identified by genetic and molecular studies. However, knowledge of maize ear development is still limited and most genes involved in this process are still unknown.In the study, we used deep-sequencing to identify, profile, and describe differentially expressed genes at four developmental stages (growth points-elongation phase, spikelet differentiation phase, floret primordium differentiation phase and floret organ differentiation phase) of B73. Identification and characterization of this important class of regulatory genes in maize may improve our understanding of molecular mechanisms controlling flower development.The results were showed as follows.1. To genome-wide ccomparative analysis of gene expression profiles, we generated four libraries from developmental ears of maize mentioned above. The high-throughput sequencing of these libraries resulted in 3,931,920 (270,298 distinct),4,066,158 (278,664 distinct),4,054,326 (256,999 distinct) and 4,055,577 (238,767 distinct) tags. Copy number of most of the distinct tags (over 78%) ranged from 1 to 5 (1≤frequency≤5).2. We used SOAP2 software to map all distinct tags to the maize reference genome (B73 RefGen_v2), and identified 24,480 (75.23%)-24,999 (43.79%) genes expressed in each pairwise comparison. Our analysis revealed 18,573 genes expressed in all samples and 24,440 genes expressed collectively in the present study.3. Significantly changed genes were detected between every two samples by a strict Bayesian algorithm developed by Audic et al., and a total of 6,800 (27.9%) genes were identified with significant differential expression (FDR<0.001 and absolute value of log2Ratio≥1). To validate differentially expressed genes detected in this study, qRT-PCR was carried out using 32 randomly selected genes. The expression profiles of 32 genes were generally consistent with DGE data.4. We used Gene Ontology annotation to assign genes to functional categories and grouped genes by expression dynamics using the K-Means clustering algorithm. We identified 18 clusters which contained genes ranging from 56 (K3)-766 (K13). Genes for cellular component organization or biogenesis were greatly enriched in cluster K1, including 135 genes as well as cluster K7. Gene responsible for response to stress showed continuously falling expression and were greatly enriched in cluster K2.56 biological macromolecular synthesis-related genes were found in K3. Genes with peak expression in Stage IV (clusters K4,63 genes) were found to be required for triose phosphate transport and response to hexose stimulus. The ear gene expression. was lowest in Stage Ⅲ (clusters K5,105 genes) and was nearly exclusively restricted to genes involved in cellular aromatic compound metabolic process. Few genes encoding enzymes for cellular process, response to inorganic substance, fatty acid metabolic process, gene expression, and biosynthetic process were also greatly enriched in clusters K8-K18. No significant enriched GO was over-represented in K6.5. Biological pathways influenced by drought were also evaluated by enrichment analysis of all differentially expressed genes. A total of 10 pathways were affected based on the above mentioned 18 clusters (P< 0.05), and differentially expressed genes with pathway annotation were listed according to enrichment priority. The expression level of genes in cluster K1 was lowest in Stage I and then dramatically increased with stage-course, and mildly declined in Stage IV, which were primarily related to phagosome and ribosome metabolism. Genes in the K2 cluster were mainly annotated as cutin, suberine and wax biosynthesis, biosynthesis of secondary metabolites, phenylalanine metabolism, limonene and pinene degradation, and phenylpropanoid biosynthesis. However, few genes encoding enzymes for phenylalanine, tyrosine and tryptophan biosynthesis, aminoacyl-tRNA biosynthesis and ribosome metabolism were also greatly enriched in clusters K3-K18.6. A primary objective was to identify genes that encode transcription factors (TF) and resolve the dynamics of accumulation of TFs during ear development in our DGE data. A total of 1,522 TFs were identified. Of these TFs,1,055 were expressed in all four developmental stages. Furthermore,41,39,28, and 40 TFs were specifically expressed in Stage Ⅰ, Stage Ⅱ, Stage Ⅲ, and Stage Ⅳ, respectively. Of these TFs expressed in four developmental stages mentioned above,630 were differentially expressed during time-points and belong to 24 TF families. These TFs associated with functions in drought tolerance (MYB, NAC, and ERF), while others played roles in development and meristem maintenance or identity (HD-ZIP, NF-YA, NAC, GRAS, and TCP), defense/stress signaling pathways (HSP, WRKY, and bZIP), hormone-mediated or stress-mediated signaling by auxin (AUX/IAA), brassinosteroids (BES), or ethylene and stress (AP2/ERF).7. To describe differentially expressed genes at four developmental stages, most of the top 20 differentially expressed genes in stage Ⅱ compared to stage I are unknown proteins or hypothetical proteins. Of these, GRMZM2G120182 encodes extensin, the significantly up-regulated expression of which might promote the development of spikelet meristem cell wall. Similarly, most of the top 20 differentially expressed genes in stage Ⅲ relative to stage Ⅱ are unknown or hypothetical proteins. Of these, AC187157.4_FG005 and GRMZM2G151319 encode HD-Zip Ⅲ protein and carboxyl-terminal proteinase, respectively. The significantly up-regulated expression of AC187157.4FG005 might act as a differentiation-promoting transcription factor of the vascular meristems in floret primordium differentiation phase, and the other, GRMZM2G151319, might involved in carboxyl-terminal protein processing. As described in stage Ⅱ and Ⅳ, most of the top 20 differentially expressed genes in stage IV relative to stage Ⅲ are unknown or hypothetical proteins. Interestingly, GRMZM2G333049 encoding a VQ motif family protein, the significantly down-regulated expression of which might regulate the development of floret organ, and then regulate endosperm growth and seed size. |
PDF Full Text Request