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Biomedical subjects

C A Haskell

Publications and source records attributed to C A Haskell.

7 recordsLinked to original sources
PubMed2001

Tumor necrosis factor-alpha-converting enzyme mediates the inducible cleavage of fractalkine.

Fractalkine (FK, CX3CL1) is a novel multidomain protein expressed on the surface of endothelial cells. As a full-length transmembrane protein, FK binds cells expressing CX3CR1, its cognate receptor, with high affinity. Proteolytic cleavage of FK releases a soluble form that is a potent chemoattractant for monocytes, T cells, and natural killer cells. Activation of protein kinase C dramatically increases the rate of this cleavage. Regulation of FK cleavage is critical for maintaining the balance between the immobilized and soluble forms, but the protease responsible has not been identified. Here we report that tumor necrosis factor-alpha-converting enzyme (TACE) is primarily responsible for the inducible cleavage of FK. After transfection into host cells, the proteolytic cleavage of FK was blocked by TACE-specific inhibitors and was not detected in cells genetically altered to remove TACE activity. In contrast, the constitutive cleavage of FK was not mediated by TACE and proceeded normally in TACE-null fibroblasts. We conclude that TACE is primarily responsible for the inducible cleavage of FK. These studies identify a potentially important link between local generation of potent cytokines and control of the balance between the cell adhesion and chemotactic properties of FK.

ADAM Proteins↗
PubMed2001

Targeted deletion of CX(3)CR1 reveals a role for fractalkine in cardiac allograft rejection.

Fractalkine (Fk) is a structurally unusual member of the chemokine family. To determine its role in vivo, we generated mice with a targeted disruption of CX(3)CR1, the receptor for Fk. CX(3)CR1(-/-) mice were phenotypically indistinguishable from wild-type mice in a pathogen-free environment. In response to antibody-induced glomerulonephritis, CX(3)CR1(-/-) and CX(3)CR1(+/+) mice had similar levels of proteinuria and injury. CX(3)CR1(-/-) and CX(3)CR1(+/+) mice also developed similar levels of disease in myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis. We performed heterotopic MHC class I/II cardiac transplants from BALB/c mice into C57BL/6 mice. In the absence of cyclosporin A (CsA), there was no difference in graft survival time between CX(3)CR1(-/-) and CX(3)CR1(+/+) recipient mice. However, in the presence of subtherapeutic levels of CsA, graft survival time was significantly increased in the CX(3)CR1(-/-) mice. Characterization of cells infiltrating the grafts revealed a selective reduction in natural killer cells in the CX(3)CR1(-/-) recipients in the absence of CsA and a reduction in macrophages, natural killer cells, and other leukocytes in the presence of CsA. We conclude that Fk plays an important role in graft rejection. The development of CX(3)CR1 antagonists may allow reductions in the doses of immunosuppressive drugs used in transplantation.

Animals↗
PubMed2001

Messenger RNA localization and further characterisation of the putative tachykinin receptor NK4 (NK3B).

We have previously shown that a cloned receptor, highly homologous to the NK3 tachykinin peptide receptor, encodes a novel functional tachykinin receptor NK4. Examining sites of receptor mRNA expression by Northern blot we show that NK4 mRNA is expressed in numerous rat tissues, in contrast to the NK3 receptor which has been shown to have a distribution principally in nervous tissues. We have localised the NK4 receptor mRNA in rat brain and spinal cord using in situ hybridisation. NK4 receptor mRNA is widely expressed in neurons in the rat central nervous system, including cerebral cortex, hippocampus, hypothalamus and dorsal horn of the spinal cord. During peripheral inflammation of the hindpaw, NK4 mRNA shows complex patterns of regulation. We have also investigated some pharmacological properties of this receptor expressed ectopically in Xenopus oocytes. We show that the functional antagonism of dynorphin at the NK4 receptor is reversed by the non-specific opioid antagonist naloxone and that tachykinin-evoked responses at the NK4 receptor are inhibited by the non-peptide NK3 receptor antagonist SR142801 in a concentration dependent manner.

Animals↗
PubMed2000

Unique role of the chemokine domain of fractalkine in cell capture. Kinetics of receptor dissociation correlate with cell adhesion.

The chemokine fractalkine (FK) has two structural features that make it unique in the chemokine family: a CX(3)C motif and an extended carboxyl terminus that anchors it to the cell surface. This mucin-like stalk or an equivalent spacer is required for FK to mediate the adhesion of cells expressing its receptor, CX(3)CR1. To determine whether the ability of FK to act as a cell adhesion molecule is due to the unique presentation of a chemokine domain on a stalk or to properties of the chemokine domain itself, we created a series of chimeras in which other soluble chemokines (RANTES (regulated on activation normal T cell expressed), monocyte chemoattractant protein 1, macrophage inflammatory protein 1 beta, secondary lymphoid tissue chemokine, and interleukin 8) were fused to the mucin stalk. When tested in a static-cell adhesion assay, many of these chemokine chimeras demonstrated activity equivalent to that of FK. In flow assays, however, none of the chimeras captured cells as efficiently as FK. Interestingly, FK captured cells expressing either CX(3)CR1 or the viral receptor US28. Cells bound to FK without rolling or detaching, whereas the interleukin 8 and monocyte chemoattractant protein 1 chimeras induced primarily cell rolling and detaching, respectively. In binding studies, FK has a significantly slower off-rate from its receptors than any of the other chemokine chimeras had for their cognate receptors. We conclude that presentation of a chemokine atop a mucin-like stalk is not, in and of itself, sufficient to capture cells. The unique ability of FK to mediate adhesion under flow may be a function of its slow receptor off-rate.

Animals↗
PubMed1999

Molecular uncoupling of fractalkine-mediated cell adhesion and signal transduction. Rapid flow arrest of CX3CR1-expressing cells is independent of G-protein activation.

Fractalkine is a novel multidomain protein expressed on the surface of activated endothelial cells. Cells expressing the chemokine receptor CX3CR1 adhere to fractalkine with high affinity, but it is not known if adherence requires G-protein activation and signal transduction. To investigate the cell adhesion properties of fractalkine, we created mutated forms of CX3CR1 that have little or no ability to transduce intracellular signals. Cells expressing signaling-incompetent forms of CX3CR1 bound rapidly and with high affinity to immobilized fractalkine in both static and flow assays. Video microscopy revealed that CX3CR1-expressing cells bound more rapidly to fractalkine than to VCAM-1 (60 versus 190 ms). Unlike VCAM-1, fractalkine did not mediate cell rolling, and after capture on fractalkine, cells did not dislodge. Finally, soluble fractalkine induced intracellular calcium fluxes and chemotaxis, but it did not activate integrins. Taken together these data provide strong evidence that CX3CR1, a seven-transmembrane domain receptor, mediates robust cell adhesion to fractalkine in the absence of G-protein activation and suggest a novel role for this receptor as an adhesion molecule.

Animals↗
PubMed1996

Functional characterization by heterologous expression of a novel cloned tachykinin peptide receptor.

An orphan receptor resembling the neurokinin 3 tachykinin receptor (NK3), initially claimed to be an atypical opioid receptor, is shown herein to respond potently to the physiological NK3 receptor ligand, neurokinin B. This 'NK4' receptor did not give functional responses in Xenopus oocytes to opioid agonists. However, NK4 receptor activation was inhibited by nanomolar concentrations of dynorphin. The NK4 receptor is therefore a tachykinin receptor which is functionally antagonized by an endogenous opioid peptide.

3T3 Cells↗