On the tertiary structure of tropomysin tactoids.

Correlation of tropomyosin amino acid sequence with uranyl acetate stained magnesium salt tactoids has been carried out by a computer graphic technique. Contrary to some previous suggestions, the results seem to indicate that there is no chain stagger within the molecule and the molecules are aligned with 178 ± 2 residue overlap in antiparallel array. In addition, the molecules for one particular tactoid appear to have an end overlap of approximately 21 residues.     

Relationship between endogenous cyclic AMP production and steroid hormone secretion in chick adrenal cells

Dispersed chick adrenal cells were incubated with either ACTH, cholera toxin or forskolin. All three agents stimulated cyclic AMP accumulation and secretion of corticosterone and aldosterone by the dispersed cells. The dose-response to ACTH was similar for cyclic AMP and corticosterone but aldosterone secretion appeared to be more sensitive to ACTH stimulation. Concentrations higher than 10(-8) M of ACTH caused suppression of corticosterone output but not of cyclic AMP accumulation or aldosterone secretion. A significant cyclic AMP accumulation occurred within 30 min of exposure to ACTH whereas significant increases in steroid secretion were observed only after 30 min. An early increase (within 30 min) in cyclic AMP accumulation with both cholera toxin and forskolin was not accompanied by any significant stimulation of steroid secretion, which occurred only after 120 min. The results with the avian adrenal cells are consistent with the thesis that steroid production in the adrenocortical cells is stimulated by cyclic AMP-dependent pathways, whereas steroid release may be modulated by others.     

Clonally related CD8+ T cells responsible for rapid population of both diffuse nasal-associated lymphoid tissue and lung after respiratory virus infection

The immune system has evolved to use sophisticated mechanisms to recruit lymphocytes to sites of pathogen exposure. Trafficking pathways are precise. For example, lymphocytes that are primed by gut pathogens can, in some cases, be imprinted with CCR9 membrane receptors, which can influence migration to the small intestine. Currently, little is known about T cell trafficking to the upper respiratory tract or the relationship between effectors that migrate to the diffuse nasal-associated lymphoid tissue (d-NALT), the lower airways, and the lung. To determine whether a T cell primed by Ag from a respiratory pathogen is imprinted for exclusive trafficking to the upper or lower respiratory tract or whether descendents from that cell have the capacity to migrate to both sites, we inoculated mice by the intranasal route with Sendai virus and conducted single-cell-sequencing analyses of CD8(+) T lymphocytes responsive to a K(b)-restricted immunodominant peptide, FAPGNYPAL (Tet(+)). Cells from the d-NALT, lung airways (bronchoalveolar lavage), lung, and mediastinal lymph node were examined 10 d postinfection to determine TCR usage and clonal relationships. We discovered that 1) Tet(+) cells were heterogeneous but preferentially used TCR elements TRAV6, TRAV16, and TRBD1; 2) both N and C termini of Vα and Vβ TCR junctions frequently encompassed charged residues, perhaps facilitating TCR αβ pairing and interactions with a neutral target peptide; and 3) T cells in the d-NALT were often clonally related to cells in the lower respiratory tract.     

Comparative expression profiling of E. coli and S. aureus inoculated primary mammary gland cells sampled from cows with different genetic predispositions for somatic cell score

Background

During the past ten years many quantitative trait loci (QTL) affecting mastitis incidence and mastitis related traits like somatic cell score (SCS) were identified in cattle. However, little is known about the molecular architecture of QTL affecting mastitis susceptibility and the underlying physiological mechanisms and genes causing mastitis susceptibility. Here, a genome-wide expression analysis was conducted to analyze molecular mechanisms of mastitis susceptibility that are affected by a specific QTL for SCS on Bos taurus autosome 18 (BTA18). Thereby, some first insights were sought into the genetically determined mechanisms of mammary gland epithelial cells influencing the course of infection.

Methods

Primary bovine mammary gland epithelial cells (pbMEC) were sampled from the udder parenchyma of cows selected for high and low mastitis susceptibility by applying a marker-assisted selection strategy considering QTL and molecular marker information of a confirmed QTL for SCS in the telomeric region of BTA18. The cells were cultured and subsequently inoculated with heat-inactivated mastitis pathogens Escherichia coli and Staphylococcus aureus, respectively. After 1, 6 and 24 h, the cells were harvested and analyzed using the microarray expression chip technology to identify differences in mRNA expression profiles attributed to genetic predisposition, inoculation and cell culture.

Results

Comparative analysis of co-expression profiles clearly showed a faster and stronger response after pathogen challenge in pbMEC from less susceptible animals that inherited the favorable QTL allele ''Q'' than in pbMEC from more susceptible animals that inherited the unfavorable QTL allele ''q''. Furthermore, the results highlighted RELB as a functional and positional candidate gene and related non-canonical Nf-kappaB signaling as a functional mechanism affected by the QTL. However, in both groups, inoculation resulted in up-regulation of genes associated with the Ingenuity pathways ''dendritic cell maturation'' and ''acute phase response signaling'', whereas cell culture affected biological processes involved in ''cellular development''.

Conclusions

The results indicate that the complex expression profiling of pathogen challenged pbMEC sampled from cows inheriting alternative QTL alleles is suitable to study genetically determined molecular mechanisms of mastitis susceptibility in mammary epithelial cells in vitro and to highlight the most likely functional pathways and candidate genes underlying the QTL effect.     

Development of paratransgenic Artemia as a platform for control of infectious diseases in shrimp mariculture

Aim: To study the accumulation and retention of recombinant proteins in Artemia gut for optimizing paratransgenic disease control in shrimp aquaculture. Methods and Results: Transgenic Escherichia coli expressing fluorescent marker proteins and the transgenic cyanobacterium Synechococcus bacillarus expressing a functional murine single chain antibody, DB3, were fed to Artemia franciscana. Stable expression and retention of several marker molecules (e.g. GFP, DS Red and DB3) up to 10 h after of feeding with E. coli were evident within the gut of Artemia. Engineered strains of S. bacillarus expressing DB3 accumulated within the gut of Artemia with detectable antibody activity for 8–10 h of feeding via ELISA, coincident with the time period of the highest density of transgenic S. bacillarus in the Artemia gut. Conclusions: Artemia fed transgenic bacteria or algae accumulated recombinant proteins for up to 10 h that retained biological activity. Co‐delivery of multiple recombinant proteins simultaneously in the gut of Artemia was also demonstrated. Significance and Impact of the Study: Expression of molecules that target infectious agents of mariculture in shrimp via commonly deployed feed organisms such as Artemia could potentially offer powerful new tools in the ongoing global effort to increase food supply.