Human T-cell Lymphotropic Virus Type 1 Tax Induction of Biologically Active NF-κB Requires IκB Kinase-1-mediated Phosphorylation of RelA/p65

Activation of the NF- Kappa B/Rel family of transcription factors proceeds through a catalytic complex containing I Kappa B kinase (IKK)-1 and IKK2. Targeted disruption of each of the IKK genes suggests that these two kinases may mediate distinct functions in the activation pathway. In our studies of the human T-cell lymphotropic virus type 1 (HTLV-1) Tax oncoprotein, we have uncovered a new function of IKK1 required for complete activation of the NF- Kappa B transcriptional program. In IKK1 super(-/-) murine embryonic fibroblasts (MEFs), Tax normally induced early NF- Kappa B activation events. However, NF- Kappa B induced by Tax in these IKK1 super(-/-) cells was functionally impaired. In IKK1 super(-/-) (but not wild-type) MEFs, Tax failed to activate several different Kappa B reporter constructs or to induce the endogenous I Kappa B alpha gene. In contrast, Tax normally activated the cAMP-responsive element-binding protein/activating transcription factor pathway, leading to full stimulation of an HTLV-1 long terminal repeat reporter construct in IKK1 super(-/-) cells. Furthermore, reconstitution of IKK1 super(-/-) cells with kinase-proficient (but not kinase-deficient) forms of IKK1 restored the Tax induction of full NF- Kappa B transactivation. We further found that the defect in NF- Kappa B action in IKK1 super(-/-) cells correlated with a failure of Tax to induce phosphorylation of the RelA/p65 subunit of NF- Kappa B at Ser super(529) and Ser super(536). Such phosphorylation of RelA/p65 was readily detected in wild-type MEFs. Phosphorylation of Ser super(536) was required for a complete response to Tax expression, whereas phosphorylation of Ser super(529) appeared to be less critical. Together, these findings highlight distinct roles for the IKK1 and IKK2 kinases in the activation of NF- Kappa B in response to HTLV-1 Tax. IKK2 plays a dominant role in signaling for I Kappa B alpha degradation, whereas IKK1 appears to play an important role in enhancing the transcriptional activity of NF- Kappa B by promoting RelA/p65 phosphorylation.