Expression and targeting of CX3CL1 (Fractalkine) in renal tubular epithelial cells

The chemokine CX(3)CL1 plays a key role in glomerulonephritis and can act as both chemoattractant and adhesion molecule. CX(3)CL1 also is upregulated in tubulointerstitial injury, but little is known about the subcellular distribution and function of CX(3)CL1 in renal tubular epithelial cells (RTEC). Unexpectedly, it was found that CX(3)CL1 is expressed predominantly on the apical surface of tubular epithelium in human renal transplant biopsy specimens with acute rejection or acute tubular necrosis. For studying the targeting of CX(3)CL1 in polarized RTEC, MDCK cells that expressed untagged or green fluorescent protein-tagged CX(3)CL1 were generated. The chemokine was present on the apical membrane and in subapical vesicles. Apical targeting of CX(3)CL1 was not due to signals that were conferred by its intracellular domain, to associations with lipid rafts, or to O-glycosylation but, rather, depended on N-linked glycosylation of the protein. With the use of fluorescence recovery after photobleaching, it was found that CX(3)CL1 is immobile in the apical membrane. However, CX(3)CL1 partitioned with the triton-soluble rather than -insoluble cellular fraction, indicating that it is not associated directly with the actin cytoskeleton or with lipid rafts. Accordingly, disruption of rafts through cholesterol depletion did not render CX(3)CL1 mobile. For exploration of potential functions of apical CX(3)CL1, binding of CX(3)CR1-expressing leukocytes to polarized RTEC was examined. Leukocyte adhesion to the luminal surface was enhanced significantly when CX(3)CL1 was present. These data demonstrate that CX(3)CL1 is expressed preferentially on the apical membrane of RTEC and suggest a novel function for the chemokine in recruitment and retention of leukocytes in tubulointerstitial inflammation.