Cytomegalovirus promotes intestinal macrophage-mediated mucosal inflammation through induction of Smad7.

Intestinal macrophages in healthy human mucosa are profoundly down-regulated for inflammatory responses (inflammation anergy) due to stromal TGF-β inactivation of NF-κB. Paradoxically, in cytomegalovirus (CMV) intestinal inflammatory disease, one of the most common manifestations of opportunistic CMV infection, intestinal macrophages mediate severe mucosal inflammation. Here we investigated the mechanism whereby CMV infection promotes macrophage-mediated mucosal inflammation.

Phosphorylation of Golgi Peripheral Membrane Protein Grasp65 Is an Integral Step in the Formation of the Human Cytomegalovirus Cytoplasmic Assembly Compartment.

Unlabelled: Human cytomegalovirus (HCMV) is the largest member of the Herpesviridae and represents a significant cause of disease. During virus replication, HCMV alters cellular functions to facilitate its replication, including significant reorganization of the secretory and endocytic pathways of the infected cell. A defining morphologic change of the infected cell is the formation of a membranous structure in the cytoplasm that is designated the virion assembly compartment (AC), which consists of virion structural proteins surrounded by cellular membranes.

p115-SNARE interactions: a dynamic cycle of p115 binding monomeric SNARE motifs and releasing assembled bundles.

Tethering factors regulate the targeting of membrane-enclosed vesicles under the control of Rab GTPases. p115, a golgin family tether, has been shown to participate in multiple stages of ER/Golgi transport. Despite extensive study, the mechanism of action of p115 is poorly understood.

Cytomegalovirus microRNAs.

The discovery that animals, plants and DNA viruses encode microRNAs (miRNAs) has transformed our understanding of the regulation of gene expression. miRNAs are ubiquitous small non-coding RNAs that regulate gene expression post-transcriptionally, generally by binding to sites within the 3' untranslated regions (UTR) of messenger RNA (mRNA) transcripts. To date, over 250 viral miRNAs have been identified primarily in members of the herpesvirus family.

Cytomegalovirus miRNAs target secretory pathway genes to facilitate formation of the virion assembly compartment and reduce cytokine secretion.

Herpesviruses, including human cytomegalovirus (HCMV), encode multiple microRNAs (miRNA) whose targets are just being uncovered. Moreover, miRNA function during the virus life cycle is relatively unknown. We find that HCMV miRs UL112-1, US5-1, and US5-2 target multiple components of the host secretory pathway, including VAMP3, RAB5C, RAB11A, SNAP23, and CDC42.

Tethering factor P115: a new model for tether-SNARE interactions.

The membrane tethering factor p115 has been shown to have important functions in ER to Golgi traffic and Golgi biogenesis. The multidomain structure of p115 allows for interactions with a diverse array of proteins that govern cargo movement at the ER-Golgi interface. Within its C-terminal region p115 contains four coiled-coil domains (CC1-CC4).

Identification of a functional domain within the p115 tethering factor that is required for Golgi ribbon assembly and membrane trafficking.

The tethering factor p115 (known as Uso1p in yeast) has been shown to facilitate Golgi biogenesis and membrane traffic in cells in culture. However, the role of p115 within an intact animal is largely unknown. Here, we document that depletion of p115 by using RNA interference (RNAi) in C.

Depletion of beta-COP reveals a role for COP-I in compartmentalization of secretory compartments and in biosynthetic transport of caveolin-1.

We have utilized small interfering RNA (siRNA)-mediated depletion of the beta-COP subunit of COP-I to explore COP-I function in organellar compartmentalization and protein traffic. Reduction in beta-COP levels causes the colocalization of markers for the endoplasmic reticulum (ER)-Golgi intermediate compartment (ERGIC), Golgi, trans-Golgi network (TGN), and recycling endosomes in large, globular compartments. The lack of spatial differentiation of these compartments is not due to a general collapse of all cellular organelles since markers for the early endosomes and lysosomes do not redistribute to the common structures.

Dissecting the role of the ARF guanine nucleotide exchange factor GBF1 in Golgi biogenesis and protein trafficking.

COPI recruitment to membranes appears to be essential for the biogenesis of the Golgi and for secretory trafficking. Preventing COPI recruitment by expressing inactive forms of the ADP-ribosylation factor (ARF) or the ARF-activating guanine nucleotide exchange factor GBF1, or by treating cells with brefeldin A (BFA), causes the collapse of the Golgi into the endoplasmic reticulum (ER) and arrests trafficking of soluble and transmembrane proteins at the ER. Here, we assess COPI function in Golgi biogenesis and protein trafficking by preventing COPI recruitment to membranes by removing GBF1.

Activation of ADP-ribosylation factor regulates biogenesis of the ATP7A-containing trans-Golgi network compartment and its Cu-induced trafficking.

ATP7A (MNK) regulates copper homeostasis by translocating from a compartment localized within the trans-Golgi network to the plasma membrane (PM) in response to increased copper load. The mechanisms that regulate the biogenesis of the MNK compartment and the trafficking of MNK are unclear. Here we show that the architecture of the MNK compartment is linked to the structure of the Golgi ribbon.

On and off membrane dynamics of the endoplasmic reticulum-golgi tethering factor p115 in vivo.

The mechanisms regulating membrane recruitment of the p115 tethering factor in vivo are unknown. Here, we describe cycling of p115 between membranes and cytosol and document the effects of Golgi matrix proteins, Rab1, and soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP) receptors (SNAREs) on this process. Rapid membrane/cytosol exchange is shown by swift (t1/2 approximately 20 s) loss of Golgi-localized p115-green fluorescent protein (GFP) after repeated photobleaching of cell periphery and rapid (t1/2 approximately 13 s) fluorescence recovery after photobleaching Golgi-localized p115-GFP.

Mutations in COCH that result in non-syndromic autosomal dominant deafness (DFNA9) affect matrix deposition of cochlin.

The COCH gene mutated in autosomal dominant sensorineural deafness (DFNA9) encodes cochlin, a major constituent of the inner ear extracellular matrix. Sequence analysis of cochlin from DFNA9 patients identified five distinct single-amino-acid mutations within a conserved region (the LCCL domain) of cochlin. To define the molecular basis of DFNA9, we have generated myc-tagged wild-type and mutant cochlins and explored their behavior in transient transfection systems.