Ursinus College Bulletin Vol. 8, No. 8

A digitized copy of the May 1892 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1075/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 5

A digitized copy of the February 1892 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1072/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 4

A digitized copy of the January 1892 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1071/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 5

A digitized copy of the February 1892 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1072/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 3

A digitized copy of the December 1891 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1070/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 2

A digitized copy of the November 1891 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1069/thumbnail.jp

Ursinus College Bulletin Vol. 8, No. 1

A digitized copy of the October 1891 Ursinus College Bulletin.https://digitalcommons.ursinus.edu/ucbulletin/1068/thumbnail.jp

Molecular basis of association of receptor activity-modifying protein 3 with the family B G protein-coupled secretin receptor

The three receptor activity-modifying proteins (RAMPs) have been recognized as being important for the trafficking and function of a subset of family B G protein-coupled receptors, although the structural basis for this has not been well established. In the current work, we use morphological fluorescence techniques, bioluminescence resonance energy transfer, and bimolecular fluorescence complementation to demonstrate that the secretin receptor associates specifically with RAMP3, but not with RAMP1 or RAMP2. We use truncation constructs, peptide competition experiments, and chimeric secretin-GLP1 receptor constructs to establish that this association is structurally specific, dependent on the intramembranous region of the RAMP and TM6 and TM7 of this receptor. There were no observed changes in secretin-stimulated cAMP, intracellular calcium, ERK1/2 phosphorylation, or receptor internalization in receptor-bearing COS or CHO-K1 cells in the presence or absence of exogenous RAMP transfection, although the secretin receptor trafficks normally to the cell surface in these cells in a RAMP-independent manner, resulting in both free and RAMP-associated receptor on the cell surface. RAMP3 association with this receptor was shown to be capable of rescuing a receptor mutant (G241C) that is normally trapped intracellularly in the biosynthetic machinery. Similarly, secretin receptor expression had functional effects on adrenomedullin activity, with increasing secretin receptor expression competing for RAMP3 association with the calcitonin receptor-like receptor to yield a functional adrenomedullin receptor. These data provide important new insights into the structural basis for RAMP3 interaction with a family B G protein-coupled receptor, potentially providing a highly selective target for drug action. This may be representative of similar interactions between other members of this receptor family and RAMP proteins

Long-Distance Delivery of Bacterial Virulence Factors by Pseudomonas aeruginosa Outer Membrane Vesicles

Bacteria use a variety of secreted virulence factors to manipulate host cells, thereby causing significant morbidity and mortality. We report a mechanism for the long-distance delivery of multiple bacterial virulence factors, simultaneously and directly into the host cell cytoplasm, thus obviating the need for direct interaction of the pathogen with the host cell to cause cytotoxicity. We show that outer membrane–derived vesicles (OMV) secreted by the opportunistic human pathogen Pseudomonas aeruginosa deliver multiple virulence factors, including β-lactamase, alkaline phosphatase, hemolytic phospholipase C, and Cif, directly into the host cytoplasm via fusion of OMV with lipid rafts in the host plasma membrane. These virulence factors enter the cytoplasm of the host cell via N-WASP–mediated actin trafficking, where they rapidly distribute to specific subcellular locations to affect host cell biology. We propose that secreted virulence factors are not released individually as naked proteins into the surrounding milieu where they may randomly contact the surface of the host cell, but instead bacterial derived OMV deliver multiple virulence factors simultaneously and directly into the host cell cytoplasm in a coordinated manner