TI - PHOSphorylation regulates human T cell leukemia virus type 1 Rex function . AB - Background Human T-cell leukemia virus type 1 ( HTLV-1 ) is a pathogenic complex deltaretrovirus , which is the causative agent of adult T-cell leukemia/lymphoma ( ATL ) and HTLV-1-associated myelopathy/tropical spastic paraparesis . In addition to the structural and enzymatic viral gene products , HTLV-1 encodes the positive regulatory proteins Tax and Rex along with viral accessory proteins . Tax and Rex proteins orchestrate the timely expression of viral genes important in viral replication and cellular transformation . Rex is a nucleolar-localizing shuttling protein that acts post-transcriptionally by binding and facilitating the export of the unspliced and incompletely spliced viral mRNAs from the nucleus to the cytoplasm . HTLV-1 Rex ( Rex-1 ) is a PHOSphoprotein and general protein kinase inhibition correlates with reduced function . Therefore , it has been proposed that Rex-1 function may be regulated through site -specific PHOSphorylation . Results We conducted a phosphoryl mapping of Rex-1 over-expressed in transfected 293 T cells using a combination of affinity purification and liquid chromatography tandem mass spectrometry . We achieved 100% physical coverage of the Rex-1 polypeptide and identified five novel PHOSphorylation sites at Thr-22 , Ser-36 , Thr-37 , Ser-97 and Ser-106 . We also confirmed evidence of two previously identified residues , Ser-70 and Thr-174 , but found no evidence of PHOSphorylation at Ser-177 . The functional significance of these PHOSphorylation events was evaluated using a Rex reporter assay and site -directed mutational analysis . Our results indicate that PHOSphorylation at Ser-97 and Thr-174 is critical for Rex-1 function . Conclusion We have mapped completely the site -specific PHOSphorylation of Rex-1 identifying a total of seven residues ; Thr-22 , Ser-36 , Thr-37 , Ser-70 , Ser-97 , Ser-106 and Thr-174 . Overall , this work is the first to completely map the PHOSphorylation sites in Rex-1 and provides important insight into the regulation of Rex-1 function .