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.