Embryonic Lethality in Homozygous Human Her-2 Transgenic Mice Due to Disruption of the Pds5b Gene

The development of antigen-targeted therapeutics is dependent on the preferential expression of tumor-associated antigens (TAA) at targetable levels on the tumor. Tumor-associated antigens can be generated de novo or can arise from altered expression of normal basal proteins, such as the up-regulation of human epidermal growth factor receptor 2 (Her2/ErbB2). To properly assess the development of Her2 therapeutics in an immune tolerant model, we previously generated a transgenic mouse model in which expression of the human Her2 protein was present in both the brain and mammary tissue. This mouse model has facilitated the development of Her2 targeted therapies in a clinically relevant and suitable model. While heterozygous Her2^+/- mice appear to develop in a similar manner to wild type mice (Her2^-/-), it has proven difficult to generate homozygous Her2^+/+ mice, potentially due to embryonic lethality. In this study, we performed whole genome sequencing to determine if the integration site of the Her2 transgene was responsible for this lethality. Indeed, we report that the Her2 transgene had integrated into the Pds5b (precocious dissociation of sisters) gene on chromosome 5, as a 162 copy concatemer. Furthermore, our findings demonstrate that Her2^+/+ mice, similar to Pds5b^-/- mice, are embryonic lethal and confirm the necessity for Pds5b in embryonic development. This study confirms the value of whole genome sequencing in determining the integration site of transgenes to gain insight into associated phenotypes.     

Modeling of cell adhesion and deformation mediated by receptor–ligand interactions

The current work is devoted to studying adhesion and deformation of biological cells mediated by receptors and ligands in order to enhance the existing models. Due to the sufficient in-plane continuity and fluidity of the phospholipid molecules, an isotropic continuum fluid membrane is proposed for modeling the cell membrane. The developed constitutive model accounts for the influence of the presence of receptors on the deformation and adhesion of the cell membrane through the introduction of spontaneous area dilation. Motivated by physics, a nonlinear receptor–ligand binding force is introduced based on charge-induced dipole interaction. Diffusion of the receptors on the membrane is governed by the receptor–ligand interaction via Fick’s Law and receptor-ligand interaction. The developed model is then applied to study the deformation and adhesion of a biological cell. The proposed model is used to study the role of the material, binding, spontaneous area dilation and environmental properties on the deformation and adhesion of the cell.     

Effects of selection for the timing of vegetative phase transition on corn borer (Lepidoptera: Noctuidae and Crambidae) damage

In maize, Zea mays L., the timing of vegetative phase transition from juvenile to adult vegetative phases can be modified through selection. A reduction in the juvenile vegetative phase has been associated with resistance to diseases and pests. The major maize pest in temperate areas is Ostrinia nubilalis (Hübner) and in Europe Sesamia nonagrioides Lefebvre. The objective of our study was to determine the effects of divergent selection for the timing of vegetative phase transition in maize on resistance to corn borers. Three cycles of divergent selection for early and late phase transition in a field corn synthetic and in a sweet corn population were evaluated separately under S. nonagrioides and O. nubilalis artificial infestation. For the field corn experiment, yield and moisture improved with selection for phase transition in both directions, but improvement was due to artifacts of selection, rather than to the change in phase transition. There were no correlated responses for corn borer damage, yield, or grain moisture due to selection for the timing of vegetative phase transition. In the sweet corn experiment, selection for the timing of vegetative phase transition had no significant effects on corn borer damage in sweet corn harvested at the fresh stage. Our results do not support the use of phase transition as an indirect criterion for improving resistance to corn borers in maize. The relationship between phase transition and pest resistance reported by other studies could depend on the genotypes or could be too weak to be detected in a selection program with wild-type maize.     

Polymorphisms in the RANTES gene increase susceptibility to active tuberculosis in Tunisia

RANTES plays a pivotal role in attracting and activating various leukocyte populations that control Mycobacterium tuberculosis infection. The present study investigated the relationship between the RANTES polymorphisms (-28C/G; rs2280788, and -403G/A; rs2107538) and susceptibility to active tuberculosis (TB) in Tunisian populations. A total of 168 patients with pulmonary TB (pTB), 55 with extrapulmonary TB (epTB), and 150 control subjects were studied. Genotype analyses were carried out using polymerase chain reaction-restriction fragment length polymorphism method. We found that the -28 GG genotype was significantly associated with susceptibility to pTB (odds ratio [OR]=11.19; 95% confidence intervals [CI], 5.14-25; P corrected for the number of genotypes [Pc]=10(-8)) and epTB (OR=11.67; 95% CI, 4.74-29.33; Pc=10(-8)). However, the -28 CC genotype was found to be significantly associated with resistance to pTB (OR=0.08; 95% CI, 0.04-0.16; Pc=10(-8)) and epTB development (OR=0.11; 95% CI, 0.05-0.27; Pc=10(-8)). -403A allele was associated with increased risk development of epTB (OR=2.21; 95% CI, 1.18-4.14; p=0.007). G-G and A-C haplotypes and the AG/GC diplotype were associated with increase susceptibility to pTB (OR=7.88, 95% CI, 5.38-11.55; Pc=3.10(-8); OR=2.32, 95% CI, 1.32-4.11; Pc=3.10(-3); OR=13.26, 95% CI, 6.06-29.89; Pc=3.10(-8); respectively) and epTB (OR=6.64, 95% CI, 4-11.05; Pc=3.10(-8); OR=2.6, 95% CI, 1.26-5.35; Pc=12.10(-3); OR=11.26, 95% CI, 4.44-29.28; Pc=3.10(-8); respectively). Collectively, our findings suggested an association of the RANTES -28C/G and -403G/A functional polymorphisms with susceptibility to Mycobacterium tuberculosis infection in Tunisian populations.     

Polymorphic regions affecting human height also control stature in cattle

Orthologous positions of 55 genes associated with height in four human populations were located on the bovine genome. Single nucleotide polymorphisms close to eight of these genes were significantly associated with stature in cattle (Bos taurus and Bos indicus). This suggests that these genes may contribute to controlling stature across mammalian species.     

C-linked antifreeze glycoprotein (C-AFGP) analogues as novel cryoprotectants

Significant cell damage occurs during cryopreservation resulting in a decreased number of viable and functional cells post-thawing. Recent studies have correlated the unsuccessful outcome of regenerative therapies with poor cell viability after cryopreservation. Cell damage from ice recrystallization during freeze-thawing is one cause of decreased viability after cryopreservation. We have assessed the ability of two C-AFGPs that are potent inhibitors of ice recrystallization to increase cell viability after cryopreservation. Our results indicate that a 1-1.5 mg/mL (0.5-0.8 mM) solution of C-AFGP 1 is an excellent alternative to a 2.5% DMSO solution for the cryopreservation of human embryonic liver cells.     

Two Dictyostelium orthologs of the prokaryotic cell division protein FtsZ localize to mitochondria and are required for the maintenance of normal mitochondrial morphology

In bacteria, the protein FtsZ is the principal component of a ring that constricts the cell at division. Though all mitochondria probably arose through a single, ancient bacterial endosymbiosis, the mitochondria of only certain protists appear to have retained FtsZ, and the protein is absent from the mitochondria of fungi, animals, and higher plants. We have investigated the role that FtsZ plays in mitochondrial division in the genetically tractable protist Dictyostelium discoideum, which has two nuclearly encoded FtsZs, FszA and FszB, that are targeted to the inside of mitochondria. In most wild-type amoebae, the mitochondria are spherical or rod-shaped, but in fsz-null mutants they become elongated into tubules, indicating that a decrease in mitochondrial division has occurred. In support of this role in organelle division, antibodies to FszA and FszA-green fluorescent protein (GFP) show belts and puncta at multiple places along the mitochondria, which may define future or recent sites of division. FszB-GFP, in contrast, locates to an electron-dense, submitochondrial body usually located at one end of the organelle, but how it functions during division is unclear. This is the first demonstration of two differentially localized FtsZs within the one organelle, and it points to a divergence in the roles of these two proteins.     

β-Catenin Inactivation Is a Pre-Requisite for Chick Retina Regeneration

In the present study we explored the role of β-catenin in mediating chick retina regeneration. The chick can regenerate its retina by activating stem/progenitor cells present in the ciliary margin (CM) of the eye or via transdifferentiation of the retinal pigmented epithelium (RPE). Both modes require fibroblast growth factor 2 (FGF2). We observed, by immunohistochemistry, dynamic changes of nuclear β-catenin in the CM and RPE after injury (retinectomy). β-catenin nuclear accumulation was transiently lost in cells of the CM in response to injury alone, while the loss of nuclear β-catenin was maintained as long as FGF2 was present. However, nuclear β-catenin positive cells remained in the RPE in response to injury and were BrdU-/p27+, suggesting that nuclear β-catenin prevents those cells from entering the cell cycle. If FGF2 is present, the RPE undergoes dedifferentiation and proliferation concomitant with loss of nuclear β-catenin. Moreover, retinectomy followed by disruption of active β-catenin by using a signaling inhibitor (XAV939) or over-expressing a dominant negative form of Lef-1 induces regeneration from both the CM and RPE in the absence of FGF2. Our results imply that β-catenin protects cells of the CM and RPE from entering the cell cycle in the developing eye, and specifically for the RPE during injury. Thus inactivation of β-catenin is a pre-requisite for chick retina regeneration.