The current through the L‐type calcium channel is inhibited and stimulated by distinct dihydropyridines at very low concentrations. The molecular determinants for the high affinity block and stimulation were investigated using chimeras between the class C and E calcium channels. Mutation of three amino acids in the last putative transmembrane segment (IVS6) of the alpha1C subunit decreased the affinity for (+)isradipine 100‐fold without significantly affecting the basic properties of the expressed channel. Mutation of two of these three amino acids completely abolished the stimulatory effect of the calcium channel agonist Bay K 8644. These mutations only slightly affected the blocking efficacy of mibefradil and the phenylalkylamine devapamil. Three distinct but adjacently located amino acids mediated the high affinity block by devapamil. These results suggest that the IVS6 segment of the alpha1C subunit is critical for the high affinity interaction between the L‐type calcium channel and the calcium channel agonist Bay K 8644 and the two antagonists isradipine and devapamil.

The Ca2+ channel subunits alpha 1C-a and alpha 1C-b were stably expressed in Chinese hamster ovary (CHO) and human embryonic kidney (HEK) 293 cells. The peak Ba2+ current (IBa) of these cells was not affected significantly by internal dialysis with 0.1 mM cAMP-dependent protein kinase inhibitor peptide (mPKI), 25 microM cAMP-dependent protein kinase catalytic subunit (PKA), or a combination of 25 microM PKA and 1 microM okadaic acid. The activity of the alpha 1C-b channel subunit expressed stably in HEK 293 cells was depressed by 1 microM H 89 and was not increased by superfusion with 5 microM forskolin plus 20 microM isobutyl-methylxanthine (IBMX). The alpha 1C-a.beta 2.alpha 2/delta complex was transiently expressed in HEK 293 cells; it was inhibited by internal dialysis of the cells with 1 microM H 89, but was not affected by internal dialysis with mPKI, PKA or microcystin. Internal dialysis of cells expressing the alpha 1C-a.beta 2.alpha 2/delta channel with 10 microM PKA did not induce facilitation after a 150-ms prepulse to +50 mV. The Ca2+ current (ICa) of cardiac myocytes increased threefold during internal dialysis with 5 microM PKA or 25 microM microcystin and during external superfusion with 0.1 microM isoproterenol or 5 microM forskolin plus 50 microM IBMX. These results indicate that the L-type Ca2+ channel expressed is not modulated by cAMP-dependent phosphorylation to the same extent as in native cardiac myocytes.

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Integrins of the ␤7 subfamily, ␣4␤7 and ␣E␤7, contribute to lymphocyte homing and to the development of protective or autoreactive immune responses at mucosal sites. The ␤ subunits of integrins are considered important for regulation of stimulated cell adhesion and adhesion-dependent signal transduction. Using a yeast interaction trap screen, a human WD repeat protein, termed WAIT-1, was isolated that interacts with the integrin ␤7 cytoplasmic tail and is homologous to mouse EED and Drosophila ESC proteins. WAIT-1 also binds to the cytoplasmic domains of ␣4 and ␣E but not to those of integrin ␤1, ␤2, and ␣L subunits. Association of WAIT-1 and ␤7-integrin was confirmed by coprecipitation from transiently transfected 293 cells. The binding site for WAIT-1 was mapped to a short membrane-proximal region of the ␤7 cytoplasmic tail with Tyr-735 being of critical importance. Northern blot analysis revealed multiple WAIT-1-related transcripts with differential expression in circulating leukocytes, tissue-resident cells of diverse origin, and lymphoid malignancies. These results suggest that WAIT-1, together with the recently identified RACK1, may define a novel subfamily of WD repeat proteins that interact with distinct subsets of integrin cytoplasmic tails and may act as specific regulators of integrin function.Expression of ␤7-integrins, ␣4␤7 and ␣E␤7, is largely confined to leukocytes. Although ␣4␤7-integrin is homogeneously expressed on natural killer cells, eosinophils, and naive T and B cells, its distribution on effector/memory T and B cells is restricted to a subset of gut-homing lymphocytes (1-6). Peripheral blood monocytes do not express ␤7-integrins, but ␣4␤7 and ␣E␤7 are up-regulated upon stimulation of macrophage differentiation with phorbol ester or interferon-␥ (7). Recent studies have indicated that, in addition, ␣4␤7-integrin is induced on endothelial cells after treatment with proinflammatory mediators such as tumor necrosis factor (8). In contrast to ␣4␤7, expression of the ␣E␤7-integrin is confined to a subset of gutassociated T lymphocytes and dendritic cells (4,6,9).␤7-Integrins are considered important for the development and function of gut-associated lymphoid tissues. Interaction of ␣4␤7 with MAdCAM-1 allows for tissue-specific migration of circulating lymphocytes into the lamina propria and Peyer's patches of the gut (10, 11), whereas ␣E␤7 may retain intraepithelial lymphocytes within the gut epithelium through binding of E-cadherin on epithelial cells (12,13). Lack of ␤7-integrins severely impairs the development of the gut immune system, as Peyer's patches are absent or hypoplastic, and fewer intraepithelial lymphocytes are detected in ␤7-integrin-deficient mice (14). Moreover, gut-homing ␣4␤7 ϩ CD4 T cells specifically harbor cellular memory for intestinal antigens, suggesting that ␣4␤7 helps to target and segregate intestinal versus systemic immune responses (15).Integrins of the ␤7 family are involved in the development and/or progression of diseases such as colitis (16,17), diabetic insul...