Endosperm is an absorptive structure that supports embryo development or seedling germination in angiosperms. The endosperm of cereals is a main source of food, feed, and industrial raw materials worldwide. However, the genetic networks that regulate endosperm cell differentiation remain largely unclear. As a first step toward characterizing these networks, we profiled the mRNAs in five major cell types of the differentiating endosperm and in the embryo and four maternal compartments of the maize (Zea mays) kernel. Comparisons of these mRNA populations revealed the diverged gene expression programs between filial and maternal compartments and an unexpected close correlation between embryo and the aleurone layer of endosperm. Gene coexpression network analysis identified coexpression modules associated with single or multiple kernel compartments including modules for the endosperm cell types, some of which showed enrichment of previously identified temporally activated and/or imprinted genes. Detailed analyses of a coexpression module highly correlated with the basal endosperm transfer layer (BETL) identified a regulatory module activated by MRP-1, a regulator of BETL differentiation and function. These results provide a high-resolution atlas of gene activity in the compartments of the maize kernel and help to uncover the regulatory modules associated with the differentiation of the major endosperm cell types.

Development of the cereal endosperm involves cell differentiation processes that enable nutrient uptake from the maternal plant, accumulation of storage products, and their utilization during germination. However, little is known about the regulatory mechanisms that link cell differentiation processes with those controlling storage product synthesis and deposition, including the activation of zein genes by the maize (Zea mays) bZIP transcription factor Opaque-2 (O2). Here, we mapped in vivo binding sites of O2 in B73 endosperm and compared the results with genes differentially expressed in B73 and B73o2. We identified 186 putative direct O2 targets and 1677 indirect targets, encoding a broad set of gene functionalities. Examination of the temporal expression patterns of O2 targets revealed at least two distinct modes of O2-mediated gene activation. Two O2-activated genes, bZIP17 and NAKED ENDOSPERM2 (NKD2), encode transcription factors, which can in turn coactivate other O2 network genes with O2. NKD2 (with its paralog NKD1) was previously shown to be involved in regulation of aleurone development. Collectively, our results provide insights into the complexity of the O2-regulated network and its role in regulation of endosperm cell differentiation and function.

Soybean DCL2 favors long inverted repeat-derived double-stranded RNA as its substrate, and generates primary 22-nt siRNAs to regulate seed coat color and silence transposable elements.

These data indicate that the UPP and the ALP are interrelated and that dysfunction of the ALP would also result in dysfunction of the UPP and severely compromise the capacity of eliminating misfolded and other forms of damaged proteins.

The endosperm of angiosperms is a zygotic seed organ that stores nutrient reserves to support embryogenesis and seed germination. Cereal endosperm is also a major source of human calories and an industrial feedstock. Maize opaque endosperm mutants commonly exhibit opaque, floury kernels, along with other abnormal seed and/or non-seed phenotypes. The opaque endosperm phenotype is sometimes accompanied by a soft kernel texture and increased nutritional quality, including a higher lysine content, which are valuable agronomic traits that have drawn attention of maize breeders. Recently, an increasing number of genes that underlie opaque mutants have been cloned, and their characterization has begun to shed light on the molecular basis of the opaque endosperm phenotype. These mutants are categorized by disruption of genes encoding zein or non-zein proteins localized to protein bodies, enzymes involved in endosperm metabolic processes, or transcriptional regulatory proteins associated with endosperm storage programs.

Plant small RNAs are important regulatory elements that fine-tune gene expression and maintain genome integrity by silencing transposons. Reproductive organs of monocots produce abundant phased, small interfering RNAs (phasiRNAs). The 21-nt reproductive phasiRNAs triggered by miR2118 are highly enriched in pre-meiotic anthers, and have been found in multiple eudicot species, in contrast with prior reports of monocot specificity. The 24-nt reproductive phasiRNAs are triggered by miR2275, and are highly enriched during meiosis in many angiosperms. Here, we report the widespread presence of the 21-nt reproductive phasiRNA pathway in eudicots including canonical and non-canonical microRNA (miRNA) triggers of this pathway. In eudicots, these 21-nt phasiRNAs are enriched in pre-meiotic stages, a spatiotemporal distribution consistent with that of monocots and suggesting a role in anther development. Although this pathway is apparently absent in well-studied eudicot families including the Brassicaceae, Solanaceae and Fabaceae, our work in eudicots supports an earlier singular finding in spruce, a gymnosperm, indicating that the pathway of 21-nt reproductive phasiRNAs emerged in seed plants and was lost in some lineages.

Endosperm is a product of double fertilization, and provides nutrients and signals to the embryo during seed development in flowering plants. Early stages of endosperm development are critical for the development of its storage capacity through synthesis and accumulation of starch and storage proteins. Here we report on the isolation and sequencing of mRNAs from the central portion of the starchy endosperm of Zea mays (maize) B73 at 6 days after pollination. We detected a high level of correlation among the four biological replicates of RNAs isolated using laser-capture microdissection of the cell type. Because the assayed developmental stage precedes the synthesis and accumulation of the major storage proteins and starch in the endosperm, our dataset likely include mRNAs for genes that are involved in control and establishment of these storage programs. The mRNA-Seq data has been deposited in Gene Expression Omnibus (accession number GSE58504).

Several protein families play a role in the biogenesis and function of small RNAs (sRNAs) in plants. Among these are Dicer-like (DCL), RNA-directed RNA polymerase (RDR), and Argonaute (AGO) proteins, as well as accessory proteins such as double-stranded RNA-binding (DRB), SERRATE (SE), and SUPPRESSION OF SILENCING 3 (SGS3) that are co-factors of DCL and RDR proteins. Here, we present phylogenetic analyses of seven sRNA pathway protein families performed on 198 species in the Viridiplantae. We show that the eudicot DCL3 protein is orthologous to the monocot DCL5, indicating that the monocot DCL3 has diverged and specialized in heterochromatic siRNA production. We found 167 highly conserved amino acids conserved among monocot DCL3/DCL4/DCL5 sequences. Amino acids conserved between DCL3 and DCL5 were located in the Helicase and RNase domains while amino acids conserved between DCL4 and DCL5 were mostly in the dsRNA binding and PAZ domains. Absent in DCL4, we found protein regions common to DCL3/DCL5 near both extremities of the PAZ domain which can potentially increase the distance from the RNase III catalytic center and thus determine the product size. We identified multiple duplication events of AGOs that had been lost, retained or further duplicated in sub-groups, indicating that the evolution of AGOs is complex in monocots, suggesting that functional studies in model species might not be easily translated to other species. This work provides an evolutionarily coherent and standardized annotation for gene families involved in plant sRNA biogenesis.