Interleukin (IL)-17 contributes to inflammatory response in part by promoting enhanced expression of chemokines such as CXCL1 by prolonging the half-life of this constitutively unstable mRNA. While IL-17 is a weak stimulus for transcription of the CXCL1 gene, it strongly potentiates message accumulation via stabilization when the mRNA is transcribed in cells stimulated with TNF. In myeloid cells, LPS-induced CXCL1 mRNA stabilization is dependent upon AUUUA-containing sequence motifs that are recognized by the RNA binding protein Tristetraprolin (TTP). Using deletion and site specific mutagenesis, we now report that IL-17-mediated stabilization of CXCL1 mRNA in non-myeloid cells depends upon a sequence that does not contain the AUUUA motif. Furthermore, a specific two nucleotide mutation within this region markedly abrogates sensitivity for IL-17-mediated stabilization. Consistent with this finding, the IL-17-sensitive sequence does not exhibit increased instability in the presence of TTP, and CXCL1 mRNA remains unstable and can be stabilized in response to treatment with IL-17 in embryo fibroblasts (MEFs) from mice in which the TTP gene has been deleted. While the RNA binding protein KSRP has been shown to participate in regulating the instability of human CXCL8 mRNA, RNAi-based reduction in KSRP does not impact the instability mediated by the IL-17-sensitive sequence motif. These findings suggest that IL-17-mediated chemokine mRNA stabilization in non-myeloid cells utilizes a mechanism that is distinct from that operating to control AU-rich mRNA stability in myeloid cells.