How Drosophila combats microbial infection: a model to study innate immunity and host-pathogen interactions

During the past year, dramatic progress has been achieved in our understanding of Drosophila immune reactions. The completion of the Drosophila genome sequencing project, microarray analysis and the use of genetic screens have led to the identification of several new genes required to combat microbial infection, filling in some important gaps in the understanding of innate immunity. At the same time, this insect was used as a model for the study of host-pathogen interactions. The recent major advances on the mechanisms by which this insect defends itself against intrusion of pathogens are discussed in this review.     

Novel diamino imidazole and pyrrole-containing polyamides: Synthesis and DNA binding studies of mono- and diamino-phenyl-ImPy*Im polyamides designed to target 5'-ACGCGT-3'

Pyrrole- and imidazole-containing polyamides are widely investigated as DNA sequence selective binding agents that have potential use as gene control agents. The key challenges that must be overcome to realize this goal is the development of polyamides with low molar mass so the molecules can readily diffuse into cells and concentrate in the nucleus. In addition, the molecules must have appreciable water solubility, bind DNA sequence specifically, and with high affinity. It is on this basis that the orthogonally positioned diamino/dicationic polyamide Ph-ImPy*Im 5 was designed to target the sequence 5'-ACGCGT-3'. Py* denotes the pyrrole unit that contains a N-substituted aminopropyl pendant group. The DNA binding properties of diamino polyamide 5 were determined using a number of techniques including CD, ΔT(M), DNase I footprinting, SPR and ITC studies. The effects of the second amino moiety in Py* on DNA binding affinity over its monoamino counterpart Ph-ImPyIm 3 were assessed by conducting DNA binding studies of 3 in parallel with 5. The results confirmed the minor groove binding and selectivity of both polyamides for the cognate sequence 5'-ACGCGT-3'. The diamino/dicationic polyamide 5 showed enhanced binding affinity and higher solubility in aqueous media over its monoamino/monocationic counterpart Ph-ImPyIm 3. The binding constant of 5, determined from SPR studies, was found to be 1.5 × 10(7)M(-1), which is ～3 times higher than that for its monoamino analog 3 (4.8 × 10(6)M(-1)). The affinity of 5 is now approaching that of the parent compound f-ImPyIm 1 and its diamino equivalent 4. The advantages of the design of diamino polyamide 5 over 1 and 4 are its sequence specificity and the ease of synthesis compared to the N-terminus pyrrole analog 2.     

IL-12 overexpression in mice as a model for Sjögren lung disease

Interleukin-12 (IL-12), a Th1 proinflammatory cytokine, is reported to be increased in Sj?gren syndrome. To evaluate the effects of local Th1/Th2 deregulation, we generated a transgenic mouse model that overexpresses IL-12 in the lungs. IL-12 transgenic mice developed bronchial and alveolar abnormalities strikingly similar to those found in the lungs of Sj?gren patients. Pathologically, lung abnormalities began at approximately 4 mo of age and were characterized by lymphocytic infiltrates around the bronchi, intraluminal periodic acid Schiff-positive debris, increased cell proliferation in the alveolar region, and increased interstitial and alveolar macrophages. Functionally, these abnormalities translated into decreased mucociliary clearance (P<0.05 vs. wild-type littermates) and increased oxidative stress (P<0.01). The pathological and functional abnormalities were accompanied by significant changes in lung natural killer (NK) cells. The number of NK cells was fourfold higher in IL-12 transgenic than wild-type lungs (20% of all lymphoid cells vs. 5%) during the first month of life. NK cells then decreased within a narrow window of time (from 30 to 50 days of age), reaching a nadir of approximately 2% on day 50, and remained at these low levels thereafter. This new mouse model highlights the role of IL-12 in the initiation of Sj?gren syndrome.     

Regulation of alpha-amylase-encoding gene expression in germinating seeds and cultured cells of rice.

Four alpha-amylase-encoding cDNA (alpha Amy-C) clones were isolated from a cDNA library derived from poly(A)+RNA of gibberellic acid (GA3)-treated rice aleurone layers. Nucleotide sequence analysis indicates that the four cDNAs were derived from different alpha Amy genes. Expression of the individual alpha Amy gene in germinating seeds and cultured suspension cells of rice was studied using gene-specific probes. In germinating seeds, expression of the alpha Amy genes is positively regulated by GA3 in a temporally coordinated but quantitatively distinct manner. In cultured suspension cells, in contrast, expression of the alpha Amy genes is negatively and differentially regulated by sugars present in the medium. In addition, one strong and one weak carbohydrate-starvation-responsive alpha Amy genes have been identified. Interactions between the promoter region (HS501) of a rice alpha Amy gene and GA3-inducible DNA binding proteins in rice aleurone cells were also studied. A DNA mobility-shift assay showed that the aleurone proteins interact with two specific DNA fragments within HS501. One fragment is located between nt -131 to -170 and contains two imperfect directly repeated pyrimidine elements and a putative GA3-response element. The other fragment is located between nt -92 to -130 that contains a putative enhancer sequence. The interactions between aleurone proteins and these two fragments are sequence-specific and GA-responsive.