No abstract

Syncytial cells in soybean (Glycine max cultivar [cv.] Peking) roots infected by incompatible and compatible populations of soybean cyst nematode (SCN [Heterodera glycines]) were collected using laser capture microdissection (LCM). Gene transcript abundance was assayed using Affymetrix soybean GeneChips, each containing 37,744 probe sets. Our analyses identified differentially expressed genes in syncytial cells that are not differentially expressed in the whole root analyses. Therefore, our results show that the mass of transcriptional activity occurring in the whole root is obscuring identification of transcriptional events occurring within syncytial cells. In syncytial cells from incompatible roots at three dpi, genes encoding lipoxygenase (LOX), heat shock protein (HSP) 70, superoxidase dismutase (SOD) were elevated almost tenfold or more, while genes encoding several transcription factors and DNA binding proteins were also elevated, albeit at lower levels. In syncytial cells formed during the compatible interaction at three dpi, genes encoding prohibitin, the epsilon chain of ATP synthase, allene oxide cyclase and annexin were more abundant. By 8 days, several genes of unknown function and genes encoding a germin-like protein, peroxidase, LOX, GAPDH, 3-deoxy-D-arabino-heptolosonate 7-phosphate synthase, ATP synthase and a thioesterase were abundantly expressed. These observations suggest that gene expression is different in syncytial cells as compared to whole roots infected with nematodes. Our observations also show that gene expression is different between syncytial cells that were isolated from incompatible and compatible roots and that gene expression is changing over the course of syncytial cell development as it matures into a functional feeding site.

AU-rich element-binding proteins (ARE-BP) regulate the stability and/or translational efficiency of mRNAs containing cognate binding sites. Many targeted transcripts encode factors that control processes such as cell division, apoptosis, and angiogenesis, suggesting that dysregulated ARE-BP expression could dramatically influence oncogenic phenotypes. Using several approaches, we evaluated the expression of four well-characterized ARE-BPs across a variety of human neoplastic syndromes. AUF1, TIA-1, and HuR mRNAs were not systematically dysregulated in cancers; however, tristetraprolin mRNA levels were significantly decreased across many tumor types, including advanced cancers of the breast and prostate.

The syncytium is a nurse cell formed within the roots of Glycine max by the plant parasitic nematode Heterodera glycines. Its development and maintenance are essential for nematode survival. The syncytium appears to undergo two developmental phases during its maturation into a functional nurse cell. The first phase is a parasitism phase where the nematode establishes the molecular circuitry that during the second phase ensures a compatible interaction with the plant cell. The cytological features of syncytia undergoing susceptible or resistant reactions appear the same during the parasitism phase. Depending on the outcome of any defense response, the second phase is a period of syncytium maintenance (susceptible reaction) or failure (resistant reaction). In the analyses presented here, the localized gene expression occurring at the syncytium during the resistant reaction was studied. This was accomplished by isolating syncytial cells from Glycine max genotype Peking (PI 548402) by laser capture microdissection. Microarray analyses using the Affymetrix soybean GeneChip directly compared Peking syncytia undergoing a resistant reaction to those undergoing a susceptible reaction during the parasitism phase of the resistant reaction. Those analyses revealed lipoxygenase-9 and lipoxygenase-4 as the most highly induced genes in the resistant reaction. The analysis also identified induced levels of components of the phenylpropanoid pathway. These genes included phenylalanine ammonia lyase, chalcone isomerase, isoflavone reductase, cinnamoyl-CoA reductase and caffeic acid O-methyltransferase. The presence of induced levels of these genes implies the importance of jasmonic acid and phenylpropanoid signaling pathways locally at the site of the syncytium during the resistance phase of the resistant reaction. The analysis also identified highly induced levels of four S-adenosylmethionine synthetase genes, the EARLY-RESPONSIVE TO DEHYDRATION 2 gene and the 14-3-3 gene known as GENERAL REGULATORY FACTOR 2. Subsequent analyses studied microdissected syncytial cells at 3, 6 and 9 days post infection (dpi) during the course of the resistant reaction, resulting in the identification of signature gene expression profiles at each time point in a single G. max genotype, Peking.

The development of an infection in soybean [Glycine max L. cultivar (cv.) Peking] roots by incompatible (I) and compatible (C) populations of soybean cyst nematode (SCN) (Heterodera glycines) was assayed using an AffymetriX soybean GeneChip. This time-course microarray analysis, using 37,744 probe sets, measured transcript abundance during I and C. These analyses reveal that infection by individual I and C H. glycines populations influence the transcription of G. max genes differently. A substantial difference in gene expression is present between I and C at 12 h post infection. Thus, G. max can differentiate between I and C nematode populations even before they have begun to select their feeding sites. The microarray analysis identified genes induced earlier in infection during I than C. MA also identified amplitude differences in transcript abundance between I and C reactions. Some of the probe sets measuring increased transcript levels during I represented no apical meristem (NAM) and WRKY transcription factors as well as NBS-LRR kinases. Later during I, heat shock protein (HSPs) probe sets (i.e. HSP90, HSP70, ClpB/HSP101) measured increased transcript abundance. These results demonstrate that G. max roots respond very differently to the different H. glycines races even before their feeding site selection has occurred. The ability of G. max to engage an I reaction, thus, appears to be dependent on the ability of root cells to recognize the different races of H. glycines because these experiments were conducted in the identical G. max genetic background.

Roots of soybean, Glycine max cv. Kent L. Merr., plants susceptible to the soybean cyst nematode (SCN), Heterodera glycines Ichinohe, were inoculated and allowed to develop feeding sites (syncytia) for 8 days. Root samples enriched in syncytial cells were collected using laser capture microdissection (LCM). RNA was extracted and used to make a cDNA library and expressed sequence tags (ESTs) were produced and used for a Gene Ontology (GO) analysis. RT-PCR results indicated enhanced expression of an aquaporin (GmPIP2,2), alpha-tubulin (GmTubA1), beta-tubulin (GmTubB4) and several other genes in syncytium-enriched samples as compared to samples extracted from whole roots. While RT-PCR data showed increased transcript levels of GmPIP2,2 from LCM tissue enriched in syncytial cells, in situ hybridization showed prominent GmPIP2,2 hybridization to RNA in the parenchymal cells tightly juxtaposed to the syncytium. Immunolocalization indicated stronger alpha-tubulin signal within the syncytium as compared to surrounding tissue. However, alpha-tubulin labeling appeared diffuse or clumped. Thus, LCM allowed for the isolation of tissue enriched for syncytial cells, providing material suitable for a variety of molecular analyses.

ORCID IDs: 0000-0002-6902-740X (C.K.); 0000-0001-9969-9381 (Z.L.).Flowers are reproductive organs and precursors to fruits and seeds. While the basic tenets of the ABCE model of flower development are conserved in angiosperms, different flowering plants exhibit different and sometimes unique characteristics. A distinct feature of strawberry (Fragaria spp.) flowers is the development of several hundreds of individual apocarpous (unfused) carpels. These individual carpels are arranged in a spiral pattern on the subtending stem tip, the receptacle. Therefore, the receptacle is an integral part of the strawberry flower and is of significant agronomic importance, being the precursor to strawberry fruit. Taking advantage of next-generation sequencing and laser capture microdissection, we generated different tissue-and stage-transcriptomic profiling of woodland strawberry (Fragaria vesca) flower development. Using pairwise comparisons and weighted gene coexpression network analysis, we identified modules of coexpressed genes and hub genes of tissue-specific networks. Of particular importance is the discovery of a developing receptacle-specific module exhibiting similar molecular features to those of young floral meristems. The strawberry homologs of a number of meristem regulators, including LOST MERISTEM and WUSCHEL, are identified as hub genes operating in the developing receptacle network. Furthermore, almost 25% of the F-box genes in the genome are transiently induced in developing anthers at the meiosis stage, indicating active protein degradation. Together, this work provides important insights into the molecular networks underlying strawberry's unique reproductive developmental processes. This extensive floral transcriptome data set is publicly available and can be readily queried at the project Web site, serving as an important genomic resource for the plant biology research community.

The soybean defense response to the soybean cyst nematode was used as a model to map at cellular resolution its genotype-defined cell fate decisions occurring during its resistant reactions. The defense responses occur at the site of infection, a nurse cell known as the syncytium. Two major genotype-defined defense responses exist, the G. max ([Peking])- and G. max ([PI 88788])-types. Resistance in G. max ([Peking]) is potent and rapid, accompanied by the formation of cell wall appositions (CWAs), structures known to perform important defense roles. In contrast, defense occurs by a potent but more prolonged reaction in G. max ([PI 88788]), lacking CWAs. Comparative transcriptomic analyses with confirmation by Illumina® deep sequencing were organized through a custom-developed application, Pathway Analysis and Integrated Coloring of Experiments (PAICE) that presents gene expression of these cytologically and developmentally distinct defense responses using the Kyoto Encyclopedia of Genes and Genomes (KEGG) framework. The analyses resulted in the generation of 1,643 PAICE pathways, allowing better understanding of gene activity across all chromosomes. Analyses of the rhg1 resistance locus, defined within a 67 kb region of DNA demonstrate expression of an amino acid transporter and an α soluble NSF attachment protein gene specifically in syncytia undergoing their defense responses.