Monte Carlo simulations of membrane signal transduction events: Effect of receptor blockers on G-protein activation

Cells have evolved elaborate strategies for sensing, responding to, and interacting with their environment. In many systems, interaction of cell surface receptors with extracellular ligand can activate cellular signal transduction pathways leading to G-protein activation and calcium mobilization. In BC 3 H1 smooth muscle-like cells, we find that the speed of calcium mobilization as well as the fraction of cells which mobilize calcium following phenylephrine stimulation is dependent upon receptor occupation. To determine whether receptor inactivation affects calcium mobilization, we use the receptor antagonist prazosin to block a fraction of cell surface receptors prior to phenylephrine stimulation. For cases of equal receptor occupation by agonist, cells with inactivated or blocked receptors show diminished calcium mobilization following phenylephrine stimulation as compared to cells without inactivated receptors. Ligand/receptor binding and two-dimensional diffusion of receptors and G-proteins in the cell membrane are studied using a Monte Carlo model. The model is used to determine if receptor inactivation affects G-protein activation and thus the following signaling events for cases of equal equilibrium receptor occupation by agonist. The model predicts that receptor inactivation by antagonist binding results in lower G-protein activation not only by reducing the number of receptors able to bind agonist but also by restricting the movement of agonist among free receptors. The latter process is important to increasing the access of bound receptors to G-proteins.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43997/1/10439_2006_Article_BF00000009.pd

Neutrophils activate plasmacytoid dendritic cells by releasing self-DNA-peptide complexes in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a severe and incurable autoimmune disease characterized by chronic activation of plasmacytoid dendritic cells (pDCs) and production of autoantibodies against nuclear self-antigens by hyperreactive B cells. Neutrophils are also implicated in disease pathogenesis; however, the mechanisms involved are unknown. Here, we identified in the sera of SLE patients immunogenic complexes composed of neutrophil-derived antimicrobial peptides and self-DNA. These complexes were produced by activated neutrophils in the form of web-like structures known as neutrophil extracellular traps (NETs) and efficiently triggered innate pDC activation via Toll-like receptor 9 (TLR9). SLE patients were found to develop autoantibodies to both the self-DNA and antimicrobial peptides in NETs, indicating that these complexes could also serve as autoantigens to trigger B cell activation. Circulating neutrophils from SLE patients released more NETs than those from healthy donors; this was further stimulated by the antimicrobial autoantibodies, suggesting a mechanism for the chronic release of immunogenic complexes in SLE. Our data establish a link between neutrophils, pDC activation, and autoimmunity in SLE, providing new potential targets for the treatment of this devastating disease

THE PROTEIN SEQUENCE OF GLUTAMATE DEHYDROGENASE FROM Sulfolobus solfataricus A THERMOACIDOPHILIC ARCHAEBACTERIUM. IS THE PRESENCE OF N-ε-METHYLLYSINE RELATED TO THERMOSTABILITY ?

The complete amino acid sequence of glutamate dehydrogenase from the thermoacidophilic archaebacterium Sulfolobus solfataricus has been determined. The sequence was reconstructed by automated sequence analysis of peptides obtained after cleavage by trypsin, cyanogen bromide, Staphylococcus aureus V8 protease and pepsin. The enzyme subunit is composed of 421 amino acid residues yielding a molecular mass of 46.078 kDa. The presence of N-epsilon-methyllysine in six positions of the sequence was observed. Comparison of the sequence of glutamate dehydrogenase from S. solfataricus with the other known primary structures of the corresponding enzyme from different sources, gives an overall identity of 9.2% and shows a symmetrical evolutionary distance of this archaebacterial protein from the two groups of vertebrate on one side and eubacterial and low eucaryote enzymes on the other side. The occurrence of specific substitutions and a possible role for N-epsilon-methylation of lysine residues are discussed in view of current hypotheses on the molecular basis of thermal adaptation of proteins