Structural Insight into the Rotational Switching Mechanism of the Bacterial Flagellar Motor

Structural analysis of a clockwise-biased rotation mutant of the bacterial flagellar rotor protein FliG provides a new model for the arrangement of FliG subunits in the motor, and novel insights into rotation switching

PGH1, the Precursor for the Anti-Inflammatory Prostaglandins of the 1-series, Is a Potent Activator of the Pro-Inflammatory Receptor CRTH2/DP2

Prostaglandin H1 (PGH1) is the cyclo-oxygenase metabolite of dihomo-γ-linolenic acid (DGLA) and the precursor for the 1-series of prostaglandins which are often viewed as “anti-inflammatory”. Herein we present evidence that PGH1 is a potent activator of the pro-inflammatory PGD2 receptor CRTH2, an attractive therapeutic target to treat allergic diseases such as asthma and atopic dermatitis. Non-invasive, real time dynamic mass redistribution analysis of living human CRTH2 transfectants and Ca2+ flux studies reveal that PGH1 activates CRTH2 as PGH2, PGD2 or PGD1 do. The PGH1 precursor DGLA and the other PGH1 metabolites did not display such effect. PGH1 specifically internalizes CRTH2 in stable CRTH2 transfectants as assessed by antibody feeding assays. Physiological relevance of CRTH2 ligation by PGH1 is demonstrated in several primary human hematopoietic lineages, which endogenously express CRTH2: PGH1 mediates migration of and Ca2+ flux in Th2 lymphocytes, shape change of eosinophils, and their adhesion to human pulmonary microvascular endothelial cells under physiological flow conditions. All these effects are abrogated in the presence of the CRTH2 specific antagonist TM30089. Together, our results identify PGH1 as an important lipid intermediate and novel CRTH2 agonist which may trigger CRTH2 activation in vivo in the absence of functional prostaglandin D synthase

Salmonella typhimurium fliG and fliN mutations causing defects in assembly, rotation, and switching of the flagellar motor.

FliG, FliM, and FliN are three proteins of Salmonella typhimurium that affect the rotation and switching of direction of the flagellar motor. An analysis of mutant alleles of FliM has been described recently (H. Sockett, S. Yamaguchi, M. Kihara, V. M. Irikura, and R. M. Macnab, J. Bacteriol. 174:793-806, 1992). We have now analyzed a large number of mutations in the fliG and fliN genes that are responsible for four different types of defects: failure to assembly flagella (nonflagellate phenotype), failure to rotate flagella (paralyzed phenotype), and failure to display normal chemotaxis as a result of an abnormally high bias to clockwise (CW) or counterclockwise (CCW) rotation (CW-bias and CCW-bias phenotypes, respectively). The null phenotype for fliG, caused by nonsense or frameshift mutations, was nonflagellate. However, a considerable part of the FliG amino acid sequence was not needed for flagellation, with several substantial in-frame deletions preventing motor rotation but not flagellar assembly. Missense mutations in fliG causing paralysis or abnormal switching occurred at a number of positions, almost all within the middle one-third of the gene. CW-bias and CCW-bias mutations tended to segregate into separate subclusters. The null phenotype of fliN is uncertain, since frameshift and nonsense mutations gave in some cases the nonflagellate phenotype and in other cases the paralyzed phenotype; in none of these cases was the phenotype a consequence of polar effects on downstream flagellar genes. Few positions in FliN were found to affect switching: only one gave rise to the CW mutant bias and only four gave rise to the CCW mutant bias. The different properties of the FliM, FliG, and FliN proteins with respect to the processes of assembly, rotation, and switching are discussed