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Background/Aims: Trafficking, brush border membrane (BBM) retention, and signal-specific regulation of the Na+/H+ exchanger NHE3 is regulated by the Na+/H+ Exchanger Regulatory Factor (NHERF) family of PDZ-adaptor proteins, which enable the formation of multiprotein complexes. It is unclear, however, what determines signal specificity of these NHERFs. Thus, we studied the association of NHE3, NHERF1 (EBP50), NHERF2 (E3KARP), and NHERF3 (PDZK1) with lipid rafts in murine small intestinal BBM. Methods: Detergent resistant membranes ('lipid rafts') were isolated by floatation of Triton X-incubated small intestinal BBM from a variety of knockout mouse strains in an Optiprep step gradient. Acid-activated NHE3 activity was measured fluorometrically in BCECF-loaded microdissected villi, or by assessment of CO2/HCO3- mediated increase in fluid absorption in perfused jejunal loops of anethetized mice. Results: NHE3 was found to partially associate with lipid rafts in the native BBM, and NHE3 raft association had an impact on NHE3 transport activity and regulation in vivo. NHERF1, 2 and 3 were differentially distributed to rafts and non-rafts, with NHERF2 being most raft-associated and NHERF3 entirely non-raft associated. NHERF2 expression enhanced the localization of NHE3 to membrane rafts. The use of acid sphingomyelinase-deficient mice, which have altered membrane lipid as well as lipid raft composition, allowed us to test the validity of the lipid raft concept in vivo. Conclusions: The differential association of the NHERFs with the raft-associated and the non-raft fraction of NHE3 in the brush border membrane is one component of the differential and signal-specific NHE3 regulation by the different NHERFs. © 2013 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Key points Cystic fibrosis (CF) is a lethal disease characterized by low rates of epithelial Cl− and HCO3− secretion and obstruction of the airways and gastrointestinal and reproductive organs by sticky mucus. HCO3− secretion has recently been demonstrated to be necessary for mucus hydration., The most frequent CF mutation is F508del. This mutant protein is usually degraded in the proteasome. New therapeutic strategies have been developed which deliver F508del to the plasma membrane., Utilizing transgenic F508del mutant and cystic fibrosis transmembrane conductance regulator (CFTR) knockout mice, apical membrane expression of F508del protein was found to be associated with enhanced stimulation of intestinal HCO3− secretion., The predominant molecular mechanism for enhanced F508del HCO3− stimulation appeared to be the activation of a Cl− recycling pathway, with Cl− exit via membrane-resident F508del protein and Cl− entry in exchange for HCO3− by apical Cl−/HCO3− exchange. In contrast, the predominant molecular mechanism for cAMP-activated HCO3− secretion in WT intestine appears to be HCO3− exit via CFTR itself., Abstract This study investigated whether expression of the common cystic fibrosis transmembrane conductance regulator (CFTR) mutant F508del in the apical membrane of enterocytes confers increased bicarbonate secretory capacity on the intestinal epithelium of F508del mutant mice compared to that of CFTR knockout (KO) mice. CFTR KO mice, F508del mutant mice (F508del) and wild-type (WT) littermates were bred on the FVB/N background. F508del isolated brush border membrane (BBM) contained approximately 5-10% fully glycosylated band C protein compared to WT BBM. Similarly, the forskolin (FSK)-induced, CFTR-dependent short-circuit current (Δ Isc) of F508del mucosa was approximately 5-10% of WT, whereas the HCO3− secretory response () was almost half that of WT in both duodenum and mid-colon studied in vitro and in vivo. While WT intestine retained full FSK-induced in the absence of luminal Cl−, the markedly higher than Δ Isc in F508del intestine was dependent on the presence of luminal Cl−, and was blocked by CFTR inhibitors. The Ste20-related proline-alanine-rich kinases (SPAK/OSR1), which are downstream of the with-no-lysine (K) protein kinases (WNK), were rapidly phosphorylated by FSK in WT and F508del, but significantly more slowly in CFTR KO intestine. In conclusion, the data demonstrate that low levels of F508del membrane expression in the intestine of F508del mice significantly increased FSK-induced HCO3− secretion mediated by Cl−/HCO3− exchange. However, in WT mucosa FSK elicited strong SPAK/OSR1 phosphorylation and Cl−-independent HCO3− efflux. This suggests that therapeutic strategies which deliver F508del to the apical membrane have the potential to significantly enhance epithelial HCO3− secretion. [ABSTRACT FROM AUTHOR]

An abstract of the article "cGMP-dependent kinase 2, Na+/H+ regulatory factor 2, and Na+/H+ exchanger isoform 3 assemble within lipid rafts in murine small intestinal brush border membrane," by Min Luo and colleagues is presented.

Background/aims: Trafficking, brush border membrane (BBM) retention, and signal-specific regulation of the Na+/H+ exchanger NHE3 is regulated by the Na+/H+ Exchanger Regulatory Factor (NHERF) family of PDZ-adaptor proteins, which enable the formation of multiprotein complexes. It is unclear, however, what determines signal specificity of these NHERFs. Thus, we studied the association of NHE3, NHERF1 (EBP50), NHERF2 (E3KARP), and NHERF3 (PDZK1) with lipid rafts in murine small intestinal BBM., Methods: Detergent resistant membranes ("lipid rafts") were isolated by floatation of Triton X-incubated small intestinal BBM from a variety of knockout mouse strains in an Optiprep step gradient. Acid-activated NHE3 activity was measured fluorometrically in BCECF-loaded microdissected villi, or by assessment of CO2/HCO3(-) mediated increase in fluid absorption in perfused jejunal loops of anethetized mice., Results: NHE3 was found to partially associate with lipid rafts in the native BBM, and NHE3 raft association had an impact on NHE3 transport activity and regulation in vivo. NHERF1, 2 and 3 were differentially distributed to rafts and non-rafts, with NHERF2 being most raft-associated and NHERF3 entirely non-raft associated. NHERF2 expression enhanced the localization of NHE3 to membrane rafts. The use of acid sphingomyelinase-deficient mice, which have altered membrane lipid as well as lipid raft composition, allowed us to test the validity of the lipid raft concept in vivo., Conclusions: The differential association of the NHERFs with the raft-associated and the non-raft fraction of NHE3 in the brush border membrane is one component of the differential and signal-specific NHE3 regulation by the different NHERFs., (© 2013 S. Karger AG, Basel.)

In the title compound, C(10)H(9)IO, the asymmetric unit contains two mol-ecules, in which the iodo-bearing six-membered rings adopt envelope conformations [displacements of the flap atoms = 0.419 (3) and 0.431 (3) Å]. In both mol-ecules, the I atoms are disordered over two set of sites in 0.54 (4):0.46 (4) and 0.71 (3):0.29 (3) ratios. In the crystal, the packing features a weak C-H⋯π inter-action.