The promoter of aBrassica napus polygalacturonase gene directs pollen expression ofβ-glucuronidase in transgenicBrassica plants

A 647-bp 5-flanking fragment obtained from genomic clone Sta 44G(2) belonging to a family of polygalacturonase genes expressed inBrassica napus pollen was fused to the-glucuronidase (GUS) marker gene. This fusion construct was introduced intoB. napus plants viaAgrobacterium tumefaciens transformation. Analysis of the transgenicB. napus plants revealed that this promoter fragment is sufficient to direct GUS expression specifically in the anther and that GUS activity increases in pollen during maturation.Abbreviation GUS -Glucuronidase     

The promoter of the Arabidopsis thaliana SUC2 sucrose-H+ symporter gene directs expression of β-glucuronidase to the phloem: Evidence for phloem loading and unloading by SUC2

The Arabidopsis thaliana (L.) Heynh. SUC2 gene encodes a plasma-membrane sucrose-H⁺ symporter. The DNA sequence of the SUC2 promoter has been determined. Using a translational fusion of this promoter to the N-terminus of -glucuronidase (GUS) and the GUS histochemical assay, the tissue specificity of the SUC2 promoter was studied in Arabidopsis plants transformed with this fusion construct. The SUC2 promoter directed expression of GUS activity with high specificity to the phloem of all green tissues of Arabidopsis such as rosette leaves, stems, and sepals. During leaf development the expression of SUC2-GUS activity was first seen in the tips of young rosette leaves. In older leaves and during their concomitant sink/source transition, expression proceeded from the tips to the bases of the leaves, indicating that expression of the SUC2 sucrose-H⁺ symporter is tightly coupled to the source-strength of Arabidopsis leaves. Expression of SUC2-GUS activity was also seen, however, in sink tissues such as roots and developing Arabidopsis pods, suggesting that the product of the SUC2 gene might not only be important for phloem loading, but also for phloem unloading. A possible regulatory effect of carbohydrates (glucose and sucrose) on the activity of the SUC2 promoter was studied and excluded, both in excised leaves and young seedlings of transgenic Arabidopsis plants. The overall pattern of SUC2-GUS expression correlated well with that of the Arabidopsis thaliana AHA3 plasma-membrane H⁺ -ATPase which is also expressed in the phloem and most likely represents the primary pump generating the energy for secondary active transporters such as SUC2.Abbreviations GUS -glucuronidase - MS Murashige & Skocgmedium - X-Gluc 5-bromo-4-chloro-3-indolyl--d-glucuronic acid Accession number for SUC2-promoter sequences: The DNA sequence data reported in this paper will appear in the EMBL, GenBank, and DDBJ nucleotide sequence databases under the accession number X79702 (AtSUC2 promoter sequence)We want to thank Günther Peissig for growing the Arabidopsis thaliana plants. This work was supported by the Deutsche Forschungsgemeinschaft (SFB 43/C5) and a grant to N.S. from the Bundesministerium für Forschung und Technologie.     

The sucrose synthase-1 promoter from Citrus sinensis directs expression of the β-glucuronidase reporter gene in phloem tissue and in response to wounding in transgenic plants

Interest in phloem-specific promoters for the engineering of transgenic plants has been increasing in recent years. In this study we isolated two similar, but distinct, alleles of the Citrus sinensis sucrose synthase-1 promoter (CsSUS1p) and inserted them upstream of the β-glucuronidase (GUS) gene to test their ability to drive expression in the phloem of transgenic Arabidopsis thaliana and Nicotiana tabacum. Although both promoter variants were capable of conferring localized GUS expression in the phloem, the CsSUS1p-2 allele also generated a significant level of expression in non-target tissues. Unexpectedly, GUS expression was also instigated in a minority of CsSUS1p::GUS lines in response to wounding in the leaves of transgenic Arabidopsis. Deletion analysis of the CsSUS1p suggested that a fragment comprising nucleotides −410 to −268 relative to the translational start site contained elements required for phloem-specific expression while nucleotides −268 to −103 contained elements necessary for wound-specific expression. Interestingly, the main difference between the two CsSUS1p alleles was the presence of a 94-bp insertion in allele 2. Fusion of this indel to a minimal promoter and GUS reporter gene indicated that it contained stamen and carpel-specific enhancer elements. This finding of highly specific and separable regulatory units within the CsSUS1p suggests that this promoter may have a potential application in the generation of constructs for the use in the development of transgenic plants resistant to a wide variety of target pests.     

The MLH1 −93 promoter variant influences gene expression

The ?93 SNP of MLH1 gene is associated with MLH1 gene methylation in endometrial and colorectal cancers. We undertook luciferase reporter assay and electrophoretic mobility shift assay (EMSA) to test whether the ?93 SNP affects the MLH1 gene expression. The luciferase activity for ?93A plasmid is significantly lower than ?93G plasmid. In EMSA experiments, the ?93A and ?93G probes have different binding affinity to nuclear proteins of JEG3 cells. Our data indicate that ?93 SNP affects MLH1 gene expression by altering protein binding to the promoter of MLH1 gene.     

High-level expression of sucrose inducible sweet potato sporamin gene promoter: β-glucuronidase fusion gene in transgenic Nicotiana plumbaginifolia hairy roots

In Vitro Cellular & Developmental Biology - Plant - We developed transgenic Nicotiana plumbaginifolia hairy roots with sucrose-inducible minimal promoter (Spomin)-β-glucuronidase (GUS)...     

Insight into lymphoid development by gene expression profiling of avian B cells

The avian immune system provides an excellent model to track B-cell development from prebursal stem cells throughout B-cell differentiation and maturation. Bursal B cells are uniquely positioned at the crossroads of B-cell development, having properties of both stem cells and of mature B cells, as demonstrated by their ability to reconstruct the bursal B-cell compartment and to express and diversify the B-cell receptor at their cell surface. To understand avian B-cell development better, we determined the gene expression profile of different B-cell stages using a bursal expressed sequence tag array. The expression profile of bursal B cells reveals the presence of factors associated with B-cell signaling and defines novel B-cell-specific genes. Genes associated with proliferation, apoptosis, DNA repair and recombination are abundantly expressed. The expression profile of the DT40 cell line is most similar to bursal B cells rather than to other stages of B-cell development, confirming the suitability of DT40 for studies of B-cell physiology. Interestingly, prebursal stem cells express genes involved in B-cell receptor signaling, although they express only low levels of immunoglobulin genes. This suggests that B-cell receptor-mediated selection is present before bursal colonization. The gene expression signatures of germinal centers and cells of the Harderian gland indicate that evolutionarily conserved genetic programs regulate B-cell activation and terminal differentiation.Electronic Supplementary Material Supplementary material is available in the online version of this article atK. Koskela, P. Kohonen and P. Nieminen contributed equally to this work     

The promoter of Milk vetch dwarf virus component 8 confers effective gene expression in both dicot and monocot plants

The activity of a predicted promoter, PMC8, from Milk vetch dwarf virus was evaluated by comparing it with the cauliflower mosaic virus 35S RNA promoter (P35S) and PNCR, a promoter from Soybean chlorotic mottle virus. When the GUS fusion gene was introduced into tobacco, PMC8 showed a similar expression profile to P35S but with a more intense expression in proliferating tissues. The usefulness of PMC8 was confirmed by driving NPTII for selection of kanamycin-resistant tobacco plants with improved transformation efficiency. PMC8 was also effective in transgenic rice plants. Thus, PMC8 is useful as an alternative to P35S in both dicotyledonous and monocotyledonous plants, especially for gene expression in proliferating tissues.     

A hybrid bovine ß-casein/bGH gene directs transgene expression to the lung and mammary gland of transgenic mice

We investigated spatial and temporal expression of bGH controlled by two different sizes (1.8kb and 15kb) of 5-flanking sequences of the bovine ß-casein in transgenic mice. In the 1.8-kb promoter-containing mice, bGH expression was specifically confined to lung and mammary gland at lactation. While mammary gland expression was highly variable depending on the lines, lung expression was relatively constant with a high level in most lines. Moreover, this dual-tissue specificity of bGH expression was consistently retained in all of the 15kb-promoter-containing mice, although a low ectopic expression was sometimes detected in salivary gland or brain. During mammary gland development in the 1.8-kb promoter-containing mice was mammary gland expression first detected at lactation, following the bovine rather than murine pattern of ß-casein expression. In contrast, lung expression was almost constant regardless of mammary gland developmental state or sex. Therefore, it can be concluded that a combination of the bovine ß-casein promoter and bGH gene directs a distinct dual-tissue specific bGH expression with different regulatory mechanisms between mammary gland and lung and as little as 1.8-kb promoter is sufficient for the proper regulation of the bovine ß-casein gene in mammary gland.     

Primary structure and expression of mRNAs encoding basic chitinase and 1,3-β-glucanase in potato

Infection of potato leaves (Solanum tuberosum L. cv. Datura) by the late blight fungus Phytophthora infestans, or treatment with fungal elicitor leads to a strong increase in chitinase and 1,3--glucanase activities. Both enzymes have been implicated in the plant's defence against potential pathogens. In an effort to characterize the corresponding genes, we isolated complementary DNAs encoding the basic forms (class I) of both chitinase and 1,3--glucanase, which are the most abundant isoforms in infected leaves. Sequence analysis revealed that at least four genes each are expressed in elicitor-treated leaves. The structural features of the potato chitinases include a hydrophobic signal peptide at the N-terminus, a hevein domain which is characteristic of class I chitinases, a proline- and glycine-rich linker region which varies among all potato chitinases, a catalytic domain, and a C-terminal extension. The potato 1,3--glucanases also contain a N-terminal hydrophobic signal peptide and a C-terminal extension, the latter comprising a potential glycosylation site. RNA blot hybridization experiments showed that basic chitinase and 1,3--glucanase are strongly and coordinately induced in leaves in response to infection, elicitor treatment, ethylene treatment, or wounding. In addition to their activation by stress, both types of genes are regulated by endogenous factors in a developmental and organ-specific manner. Appreciable amounts of chitinase and 1,3--glucanase mRNAs were found in old leaves, stems, and roots, as well as in sepals of healthy, untreated plants, whereas tubers, root tips, and all other flower organs (petals, stamen, carpels) contained very low levels of both mRNAs. In young leaves and stems, chitinase and 1,3--glucanase were differentially expressed. While chitinase mRNA was abundant in these parts of the plant, 1,3--glucanase mRNA was absent. DNA blot analysis indicated that in potato, chitinase and 1,3--glucanase are encoded by gene families of considerable complexity.     

A truncated tRNA synthetase directs a “translational trickle” of gene expression

Comment on: Yao P, et al. Cell 2012; 149:88-100.