Somatic embryogenesis in Solanum tuberosum from cell suspension cultures: histological analysis and extracellular protein patterns

An embryogenic cell suspension, continuously grown in Murashige and Skoog (MS) medium with 0.5 mg/L of 2,4-dichlorophenoxyacetic acid, was established from friable callus of Solanum tuberosum internode sections. The cell suspension was predominantly composed of cell masses and free embryogenic cells. When transferred to an auxin-free medium with zeatin, somatic embryos (SEs) developed and converted to complete plants when cultured on solid MS medium without growth regulators. The system produced approximately 600 SEs per 50 mL of medium. In this investigation, accumulation of extracellular proteins (EPs) of different molecular weights were found associated to different phases of the embryogenic process. At the initiation of the cell suspension, cell clusters and free cells present in the culture (phase "A") secreted a 78kDa EP, unique to this phase. In phase "B", which is related to embryonic cell determination process, proteins (7-14kDa) were secreted mainly by embryogenic cells. In phase "C", SEs in different developmental stages secreted protein of 32 kDa, which appeared as a particular feature of the phase. EPs of phase "D", secreted by torpedo and mature embryos, had molecular weights between 20 and 50 kDa. Further studies will be necessary to identify these proteins and link them to previously identified somatic embryogenesis-related proteins. Histological analysis of the potato embryogenesis in liquid media showed unicellular origin of the SE.     

超级增强子在肿瘤研究中的进展

超级增强子是由多个相邻近的普通增强子组成的、驱动调控细胞身份基因表达的一个大簇,该区域富集高密度的转录因子、辅因子及增强子相关表观修饰。超级增强子所驱动的异常转录基因对维持肿瘤细胞特性至关重要。肿瘤细胞通过组装自身超级增强子,显著促进多种癌基因表达,从而增强肿瘤细胞的增殖、侵袭和转移的能力;抑制超级增强子的活性,则显著抑制肿瘤细胞的生长和存活。本文对目前报道的肿瘤细胞中超级增强子的结构特征和功能调控,以及靶向超级增强子药物研发现状进行了总结,旨在为研发新的针对超级增强子为靶点的抗肿瘤药物提供理论基础和借鉴。     

超级增强子研究进展

超级增强子是具有转录活性增强子的一个大簇,驱动控制细胞身份基因的表达,在发育和肿瘤等疾病发生过程中起到重要作用。和普通增强子相比,许多肿瘤细胞关键致癌基因是由超级增强子驱动,常见疾病如阿尔茨海默病等相关的变异显著富集于超级增强子。超级增强子在关键致癌基因的鉴定、疾病关联变异位点的发现等领域显示了巨大的应用潜力。本文首先概述了如何在全基因组水平进行增强子的鉴定,随后引入超级增强子的概念和鉴定方法,最后阐述了超级增强子的主要结构和功能特征,并对其在研究中的应用做了展望。     

Analysis of early processes in wound-induced vascular regeneration using TED3 and ZeHB3 as molecular markers

Interruption of the vascular bundles of Zinnia internodes induced transdifferentiation of cells into tracheary elements (TEs) or sieve elements (SEs) within 4 d of wounding. The early stage of the regeneration processes was analyzed using two molecular marker genes, TED3 and ZeHB3, which are expressed specifically in TE precursor cells and immature phloem cells, respectively. An increase in the numbers of TED3 and ZeHB3 mRNA-expressing cells always preceded an increase in the numbers of TEs and SEs formed. The earliest sign of vascular differentiation was the appearance 24 h after wounding of a layer(s) of TED3 mRNA-expressing cells in the inter- and intrafascicular cambial-like regions along the severed vascular bundles. In contrast, the number of ZeHB3 mRNA-expressing cells decreased dramatically along the severed bundles 24 h after wounding, and increased again 36 h after wounding. These results clearly indicate that xylem and phloem differentiation are not synchronized during vascular regeneration. Treatment with 10(-3) M colchicine abolished the expression of ZeHB3 mRNA in pith parenchyma, but not TED3 mRNA; this suggests that cell division is a prerequisite for the transdifferentiation of pith parenchymal cells into immature phloem cells expressing ZeHB3. In contrast, transdifferentiation of pith parenchymal cells to TE precursor cells does not require preceding cell division. However, the inhibition of cell division prevented the formation of both radial files of TEs and the cambial-like layer(s) of TED3 mRNA-expressing cells, and, ultimately, vascular regeneration altogether. These results imply that wound-induced cambial-like activity in and between severed vascular bundles is essential for vascular regeneration.     

Summarizing internal dynamics boosts differential analysis and functional interpretation of super enhancers

Super enhancers (SEs) are broad enhancer domains usually containing multiple constituent enhancers that hold elevated activities in gene regulation. Disruption in one or more constituent enhancers causes aberrant SE activities that lead to gene dysregulation in diseases. To quantify SE aberrations, differential analysis is performed to compare SE activities between cell conditions. The state-of-art strategy in estimating differential SEs relies on overall activities and neglect the changes in length and structure of SEs. Here, we propose a novel computational method to identify differential SEs by weighting the combinatorial effects of constituent-enhancer activities and locations (i.e. internal dynamics). In addition to overall activity changes, our method identified four novel classes of differential SEs with distinct enhancer structural alterations. We demonstrate that these structure alterations hold distinct regulatory impact, such as regulating different number of genes and modulating gene expression with different strengths, highlighting the differentiated regulatory roles of these unexplored SE features. When compared to the existing method, our method showed improved identification of differential SEs that were linked to better discernment of cell-type-specific SE activity and functional interpretation.     

Interaction of plant growth regulators and organic C and N components in the formation and maturation of Abies alba somatic embryos

To promote SE maturation, the influence of different media components on different developmental stages was quantitatively evaluated. Advanced maturation was achieved with a sequence of culture media (prematuration medium and maturation medium) that contained various carbohydrates, organic nitrogen compounds and plant growth regulators. Application of lactose, BA, L-glutamine and casein hydrolysate in the prematuration medium enhanced the total number of SEs and promoted advanced differentiation. The highest number of late torpedo stage SEs was observed on maturation medium supplemented with 200 mM lactose and 29 mM sucrose. Lactose and sorbitol favoured SE maturation up to the early cotyledonary stage. With application of PEG and high ABA concentrations (20–40 M), only early torpedo stages were formed. The number of late torpedo stage SEs was significantly higher on hormone free media or with lower ABA concentrations (0–5 M). Formation of early and late cotyledonary SEs was significantly enhanced by adding BA in the maturation medium: neither Zeatin nor 2iP were effective. In addition, low sucrose concentrations in the proliferation medium (29 mM compared to 58 mM) also favoured the formation of cotyledonary SE in the maturation medium.     

NK Cell Phenotypic Modulation in Lung Cancer Environment

Background

Nature killer (NK) cells play an important role in anti-tumor immunotherapy. But it indicated that tumor cells impacted possibly on NK cell normal functions through some molecules mechanisms in tumor microenvironment.

Materials and methods

Our study analyzed the change about NK cells surface markers (NK cells receptors) through immunofluorescence, flow cytometry and real-time PCR, the killed function from mouse spleen NK cell and human high/low lung cancer cell line by co-culture. Furthermore we certificated the above result on the lung cancer model of SCID mouse.

Results

We showed that the infiltration of NK cells in tumor periphery was related with lung cancer patients'' prognosis. And the number of NK cell infiltrating in lung cancer tissue is closely related to the pathological types, size of the primary cancer, smoking history and prognosis of the patients with lung cancer. The expression of NK cells inhibitor receptors increased remarkably in tumor micro-environment, in opposite, the expression of NK cells activated receptors decrease magnificently.

Conclusions

The survival time of lung cancer patient was positively related to NK cell infiltration degree in lung cancer. Thus, the down-regulation of NKG2D, Ly49I and the up-regulation of NKG2A may indicate immune tolerance mechanism and facilitate metastasis in tumor environment. Our research will offer more theory for clinical strategy about tumor immunotherapy.     

Influence of the transposable element neighborhood on human gene expression in normal and tumor tissues

Transposable elements (TEs) are genomic sequences able to replicate themselves, and to move from one chromosomal position to another within the genome. Many TEs contain their own regulatory regions, which means that they may influence the expression of neighboring genes. TEs may also be activated and transcribed in various cancers. We therefore tested whether gene expression in normal and tumor tissues is influenced by the neighboring TEs. To do this, we associated all human genes to the nearest TEs. We analyzed the expression of these genes in normal and tumor tissues using SAGE and EST data, and related this to the presence and type of TEs in their vicinity. We confirmed that TEs tend to be located in antisense orientation relative to their hosting genes. We found that the average number of tissues where a gene is expressed varies depending on the type of TEs located near the gene, and that the difference in expression level between normal and tumor tissues is greatest for genes that host SINE elements. This deregulation increases with the number of SINE copies in the gene vicinity. This suggests that SINE elements might contribute to the cascade of gene deregulation in cancer cells.     

Enhancement of somatic embryogenesis and plant regeneration in Japanese larch (<Emphasis Type="Italic">Larix leptolepis</Emphasis>)

Different nitrogen sources, abscisic acid (ABA), gellan gum at various concentrations, and osmotica were evaluated for their effects on maturation of somatic embryo (SE) in Japanese larch (Larix leptolepis). Different concentrations of l-glutamine or casein hydrolysate (CH) in the medium were also compared. The highest number of matured embryos was obtained with ½ Litvay (LM) medium supplemented with 1.71 mM l-glutamine and 250 mg l^?1 CH. In terms of osmoticum effect, the highest number of cotyledonary SEs was produced in medium containing 0.2 M maltose. As for the effects of ABA and gellan gum concentration, the highest number of cotyledonary SEs was achieved on a medium containing 60 μM ABA and 0.8% gellan gum. In addition, the best plantlet conversion frequency (35.5%) was obtained with SEs derived from the treatment with 60 μM ABA and 0.8% gellan gum.     

Cryopreservation of somatic embryos from <Emphasis Type="Italic">Petiveria alliacea</Emphasis> L. by different techniques based on vitrification

Petiveria alliacea L. is a medicinal plant originating from the Amazon region. This study describes an efficient cryopreservation protocol for somatic embryos (SEs) produced from roots of P. alliacea based on the comparison of vitrification, encapsulation-dehydration, and D cryo-plate techniques. With the vitrification technique, SEs treated with PVS2 solution (0.4 M sucrose, 3.3 M glycerol, 2.4 M ethylene glycol, and 1.9 M DMSO) for 30 min displayed high viability (85%) and intermediate proliferation recovery (about 12 adventitious SEs produced from original SEs [SEs/SE] after 90 d of culture). With the encapsulation-dehydration technique, lower viability (70%) and very low proliferation recovery (about two SEs/SE) were achieved with cryopreserved SEs dehydrated for 10 min in a laminar air flow cabinet. The D cryo-plate technique led to high viability (85%) and proliferation recovery (19 SEs/SE) of cryopreserved SEs after 90 min dehydration. In the experimental conditions tested, the D cryo-plate method was the most efficient technique for cryopreservation of P. alliacea SEs.