serine threonine kinase1 (stk1) and serine threonine kinase2 (stk2) are closely related maize paralogous genes predicted to encode serine/threonine protein kinases. Pollen mutated in stk1 or stk2 competes poorly with normal pollen, pointing to a defect in pollen tube germination or growth. Both genes are expressed in pollen, but not in most other tissues. In germination media, STK1 and STK2 fluorescent fusion proteins localize to the plasma membrane of the vegetative cell. RNA-seq experiments identified 534 differentially expressed genes in stk1 mutant pollen relative to wild type. Gene ontology (GO) molecular functional analysis uncovered several differentially expressed genes with putative ribosome initiation and elongation functions, suggesting that stk1 might affect ribosome function. Of the two paralogs, stk1 may play a more important role in pollen development than stk2, as stk2 mutations have a smaller pollen transmission effect. However, stk2 does act as an enhancer of stk1 because the double mutant combination is only infrequently pollen-transmitted in double heterozygotes. We conclude that the stk paralogs play an essential role in pollen development.

An RNA-Seq experiment was performed using field grown well-watered and naturally rain fed cotton plants to identify differentially expressed transcripts under water-deficit stress. Our work constitutes the first application of the newly published diploid D5 Gossypium raimondii sequence in the study of tetraploid AD1 upland cotton RNA-seq transcriptome analysis. A total of 1,530 transcripts were differentially expressed between well-watered and water-deficit stressed root tissues, in patterns that confirm the accuracy of this technique for future studies in cotton genomics. Additionally, putative sequence based genome localization of differentially expressed transcripts detected A2 genome specific gene expression under water-deficit stress. These data will facilitate efforts to understand the complex responses governing transcriptomic regulatory mechanisms and to identify candidate genes that may benefit applied plant breeding programs.

Background Cotton is the world’s primary fiber crop and is a major agricultural commodity in over 30 countries. Like many other global commodities, sustainable cotton production is challenged by restricted natural resources. In response to the anticipated increase of agricultural water demand, a major research direction involves developing crops that use less water or that use water more efficiently. In this study, our objective was to identify differentially expressed genes in response to water deficit stress in cotton. A global expression analysis using cDNA-Amplified Fragment Length Polymorphism was conducted to compare root and leaf gene expression profiles from a putative drought resistant cotton cultivar grown under water deficit stressed and well watered field conditions. Results We identified a total of 519 differentially expressed transcript derived fragments. Of these, 147 transcript derived fragment sequences were functionally annotated according to their gene ontology. Nearly 70 percent of transcript derived fragments belonged to four major categories: 1) unclassified, 2) stress/defense, 3) metabolism, and 4) gene regulation. We found heat shock protein-related and reactive oxygen species-related transcript derived fragments to be among the major parts of functional pathways induced by water deficit stress. Also, twelve novel transcripts were identified as both water deficit responsive and cotton specific. A subset of differentially expressed transcript derived fragments was verified using reverse transcription-polymerase chain reaction. Differential expression analysis also identified five pairs of duplicated transcript derived fragments in which four pairs responded differentially between each of their two homologues under water deficit stress. Conclusions In this study, we detected differentially expressed transcript derived fragments from water deficit stressed root and leaf tissues in tetraploid cotton and provided their gene ontology, functional/biological distribution, and possible roles of gene duplication. This discovery demonstrates complex mechanisms involved with polyploid cotton’s transcriptome response to naturally occurring field water deficit stress. The genes identified in this study will provide candidate targets to manipulate the water use characteristics of cotton at the molecular level.

BACKGROUND:Cotton (Gossypium spp.) is produced in over 30 countries and represents the most important natural fiber in the world. One of the primary factors affecting both the quantity and quality of cotton production is water. A major facilitator of water movement through cell membranes of cotton and other plants are the aquaporin proteins. Aquaporin proteins are present as diverse forms in plants, where they function as transport systems for water and other small molecules. The plant aquaporins belong to the large major intrinsic protein (MIP) family. In higher plants, they consist of five subfamilies including plasma membrane intrinsic proteins (PIP), tonoplast intrinsic proteins (TIP), NOD26-like intrinsic proteins (NIP), small basic intrinsic proteins (SIP), and the recently discovered X intrinsic proteins (XIP). Although a great deal is known about aquaporins in plants, very little is known in cotton.RESULTS:From a molecular cloning effort, together with a bioinformatic homology search, 71 upland cotton (G. hirsutum) aquaporin genes were identified. The cotton aquaporins consist of 28 PIP and 23 TIP members with high sequence similarity. We also identified 12 NIP and 7 SIP members that showed more divergence. In addition, one XIP member was identified that formed a distinct 5th subfamily. To explore the physiological roles of these aquaporin genes in cotton, expression analyses were performed for a select set of aquaporin genes from each subfamily using semi-quantitative reverse transcription (RT)-PCR. Our results suggest that many cotton aquaporin genes have high sequence similarity and diverse roles as evidenced by analysis of sequences and their expression.CONCLUSION:This study presents a comprehensive identification of 71 cotton aquaporin genes. Phylogenetic analysis of amino acid sequences divided the large and highly similar multi-gene family into the known 5 aquaporin subfamilies. Together with expression and bioinformatic analyses, our results support the idea that the genes identified in this study represent an important genetic resource providing potential targets to modify the water use properties of cotton.

Background: In metazoans, microRNAs, or miRNAs, constitute a growing family of small regulatory RNAs that are usually 19-25 nucleotides in length. They are processed from longer precursor RNAs that fold into stem-loop structures by the ribonuclease Dicer and are thought to regulate gene expression by base pairing with RNAs of protein-coding genes. In Arabidopsis thaliana, mutations in CARPEL FACTORY (CAF), a Dicer homolog, and those in a novel gene, HEN1, result in similar, multifaceted developmental defects, suggesting a similar function of the two genes, possibly in miRNA metabolism.Results: To investigate the potential functions of CAF and HEN1 in miRNA metabolism, we aimed to isolate miRNAs from Arabidopsis and examine their accumulation during plant development in wild-type plants and in hen1-1 and caf-1 mutant plants. We have isolated 11 miRNAs, some of which have potential homologs in tobacco, rice, and maize. The putative precursors of these miRNAs; have the capacity to form stable stem-loop structures. The accumulation of these miRNAs; appears to be spatially or temporally controlled in plant development, and their abundance is greatly reduced in caf-1 and hen1-1 mutants. HEN1 homologs are found in bacterial, fungal, and metazoan genomes.Conclusions: miRNAs are present in both plant and animal kingdoms. An evolutionarily conserved mechanism involving a protein, known as Dicer in animals and CAF in Arabidopsis, operates in miRNA metabolism. HEN1 is a new player in miRNA accumulation in Arabidopsis, and HEN1 homologs in metazoans may have a similar function. The developmental defects associated with caf-1 and hen1-1 mutations and the patterns of miRNA accumulation suggest that miRNAs play fundamental roles in plant development.

We show here that, although genes constitute only a small percentage of the maize genome, it is possible to identify them phenotypically as Ac receptor sites. Simple and efficient Ac transposition assays based on the well-studied endosperm markers bz and wx were used to generate a collection of >1300 independent Ac transposants. The majority of transposed Ac elements are linked to either the bz or the wx donor loci on chromosome 9. A few of the insertions produce obvious visible phenotypes, but most of them do not, suggesting that these populations will be more useful for reverse genetics than for forward transposon mutagenesis. An inverse polymerase chain reaction method was adapted for the isolation of DNA adjacent to the transposed Ac elements (tac sites). Most Ac insertions were into unique DNA. By sequencing tac sites and comparing the sequences to existing databases, insertions were identified in a number of putative maize genes. The expression of most of these genes was confirmed by RNA gel blot analysis. We report here the isolation and characterization of the first 46 tac sites from the two insertion libraries.