Increased gene and protein expression of the novel eNOS regulatory protein NOSTRIN and a variant in alcoholic hepatitis.

Background & Aims: Increased intrahepatic resistance in cirrhosis is associated with reduced endothelial NO synthase (eNOS) activity and exacerbated by superimposed inflammation. NOSTRIN induces intracellular translocation of eNOS and reduces NO generation. Our aims were to quantify and compare hepatic expression of eNOS, NOSTRIN, NOSIP, and caveolin-1 in alcoholic cirrhosis with or without superimposed alcoholic hepatitis and in normal livers.

Reggie-1 and reggie-2 localize in non-caveolar rafts in epithelial cells: cellular localization is not dependent on the expression of caveolin proteins.

Reggie-1 and reggie-2 are highly conserved and widely expressed proteins associated with membrane rafts. The molecular function of reggies remains to be clarified, but recent data indicate that they are involved in various cellular processes such as insulin signaling, phagocytosis and actin remodeling. However, there is discrepancy in the literature if reggies are associated with caveolae or non-caveolar rafts.

Polarized transport of Alzheimer amyloid precursor protein is mediated by adaptor protein complex AP1-1B.

Alzheimer amyloid precursor protein (APP) is the precursor for the Abeta peptide involved in pathogenesis of Alzheimer's disease. The soluble ectodomain fragment of APP (sAPP) functions as a growth factor for epithelial cells, suggesting an important function for APP outside neuronal tissue. Previous studies have shown that in polarized epithelial cells, APP is targeted to the basolateral domain.

Translocation of endothelial nitric-oxide synthase involves a ternary complex with caveolin-1 and NOSTRIN.

Recently, we characterized a novel endothelial nitric-oxide synthase (eNOS)-interacting protein, NOSTRIN (for eNOS-trafficking inducer), which decreases eNOS activity upon overexpression and induces translocation of eNOS away from the plasma membrane. Here, we show that NOSTRIN directly binds to caveolin-1, a well-established inhibitor of eNOS. Because this interaction occurs between the N terminus of caveolin (positions 1-61) and the central domain of NOSTRIN (positions 323-434), it allows for independent binding of each of the two proteins to eNOS.

Subcellular targeting and trafficking of nitric oxide synthases.

Unlike most other endogenous messengers that are deposited in vesicles, processed on demand and/or secreted in a regulated fashion, NO (nitric oxide) is a highly active molecule that readily diffuses through cell membranes and thus cannot be stored inside the producing cell. Rather, its signalling capacity must be controlled at the levels of biosynthesis and local availability. The importance of temporal and spatial control of NO production is highlighted by the finding that differential localization of NO synthases in cardiomyocytes translates into distinct effects of NO in the heart.

FCH/Cdc15 domain determines distinct subcellular localization of NOSTRIN.

NOSTRIN, an NO synthase binding protein, belongs to the PCH family of proteins, exposing a typical domain structure. While its SH3 domain and the C-terminal coiled-coil region cc2 have been studied earlier, the function of the N-terminal half comprising a Cdc15 domain with an FCH (Fes/CIP homology) region followed by a coiled-coil stretch cc1 is unknown. Here, we show that the FCH region is necessary and sufficient for membrane association of NOSTRIN, whereas the Cdc15 domain further specifies subcellular distribution of the protein.

NOSTRIN functions as a homotrimeric adaptor protein facilitating internalization of eNOS.

Intracellular trafficking of endothelial nitric oxide synthase (eNOS) between different compartments is incompletely understood. Recently, we described a novel eNOS-interacting protein, NOSTRIN, which upon overexpression drives eNOS away from the plasma membrane towards intracellular compartments. Sequence similarity of NOSTRIN and pacsins/syndapins suggested a role for NOSTRIN in endocytosis.

The receptor-bound N-terminal ectodomain of the amyloid precursor protein is associated with membrane rafts.

The soluble N-terminal ectodomain of amyloid precursor protein (sAPP), resulting from alpha-secretase-mediated proteolytic processing, has been shown to function as a growth factor for epithelial cells, including keratinocytes and thyrocytes. Extracellularly applied sAPP binds to a cell surface receptor and exhibits a patchy binding pattern reminiscent of that observed for raft proteins. Here we show that (i) the receptor-bound sAPP resides in a detergent-insoluble membrane microdomain which cofractionates in density gradients with cholesterol-rich membrane rafts and caveolae; (ii) the sAPP-binding microdomains are different from caveolae; and (iii) sAPP is capable of binding to isolated rafts and inducing tyrosine phosphorylation of some raft proteins.

AP-4 binds basolateral signals and participates in basolateral sorting in epithelial MDCK cells.

Adaptors are heterotetrameric complexes that mediate the incorporation of cargo into transport vesicles by interacting with sorting signals present in the cytosolic domain of transmembrane proteins. Four adaptors, AP-1 (beta 1, gamma, mu 1A or mu 1B, sigma 1), AP-2 (beta 2, alpha, mu 2, sigma 2), AP-3 (beta 3 , delta, mu 3, sigma 3) or AP-4 (beta 4, epsilon, mu 4, sigma 4), have been characterized. AP-1 and AP-3 mediate sorting events at the level of the TGN and/or endosomes, whereas AP-2 functions in endocytic clathrin coated vesicle formation; no function is known so far for AP-4.