Molecular Mechanism of hTid-1, the Human Homolog of Drosophila Tumor Suppressor l(2)Tid, in the Regulation of NF-κB Activity and Suppression of Tumor Growth

Abstract

hTid-1, a human homolog of the Drosophila tumor suppressor l(2)Tid and a novel DnaJ protein, regulates the activity of nuclear factor B (NF-B), but its mechanism is not established. We report here that hTid-1 strongly associated with the cytoplasmic protein complex of NF-B-IB through direct interaction with IB␣/␤ and the IKK␣/␤ subunits of the IB kinase complex. These interactions resulted in suppression of the IKK activity in a J-domain-dependent fashion and led to the cytoplasmic retention and enhanced stability of IB. Overexpression of hTid-1 by using recombinant baculovirus or adenovirus led to inhibition of cell proliferation and induction of apoptosis of human osteosarcoma cells regardless of the p53 expression status. hTid-1 is a human homologue of the Drosophila tumor suppressor Tid56 encoded by the lethal(2)tumorous imaginal discs, or l(2)tid, gene (26). Loss of the l(2)tid gene causes malignancy of the anlagen of the adult organs, the imaginal discs in Drosophila (26). hTid-1 was initially identified as a cellular target for the viral oncogenic protein E7 derived from human papillomavirus type 16 (HPV16) (36). hTid-1 also serves as the intracellular target for the viral transforming protein Tax from human T-cell leukemia virus type 1 (HTLV-1) and represses the Tax-induced transactivation of nuclear factor B (NF-B) (7,8). Subsequent studies demonstrated that hTid-1 interacts with the viral nuclear protein UL9 from herpes simplex virus type 1 (HSV-1) in enhancing the binding of UL9 to the viral genome to facilitate viral replication (14) and that hTid-1 forms a protein complex with Jak2 tyrosine kinase, adversely affecting its kinase activity (35).The htid1 gene encodes two spliced variants of hTid-1, hTid-1 L , and hTid-1 S (37). The full-length hTid-1 L (long) comprises 480 amino acids, whereas hTid-1 S (short), generated via alternative splicing, produces a predicted protein of 453 amino acids that lacks the C-terminal 33 amino acids of hTid-1 L but contains an additional 6 amino acids (KRSTGN) (37). Aside from differences at the C termini, both hTid-1 variants share identical structural motifs, including an N-terminal mitochondria-processing signal peptide, an N-terminal conserved signature J domain, and a central cysteine-rich motif. hTid-1 was classified as a member of the DnaJ protein family and a molecular cochaperone based on its signature J domain and ability to interact with the heat shock protein 70 chaperone (Hsp70) (25,27). Currently, the biological functions of these two forms of hTid-1 are unclear. The two variants were reported to exhibit opposing effects on induction of apoptosis in the human osteosarcoma cell line U2OS in response to tumor necrosis factor-alpha (TNF-␣) and mitomycin C (37), but the mechanism by which hTid-1-mediated apoptosis and antiapoptosis is unknown. Whether hTid-1 functions as a human tumor suppressor is also unknown. The early reports that hTid-1 serves as an intracellular target for viral oncogenic proteins Tax and E7 suggest a role for hTid-1 as a tumor suppre...

The Role of the Phylogenetically Conserved Cochaperone Protein Droj2/DNAJA3 in NF-κB Signaling

“…With regard to the NF-B pathway with DNAJ proteins, Cheng et al (63,64) have demonstrated that overexpression of DNAJA3 suppresses IB phosphorylation by IKK␤ and, thereby, concluded that DNAJA functions as a negative regulator of NF-B. Our study also indicates a role for DNAJA3 in IB phosphorylation, but our conclusion is opposite that of previous studies.…”

The Tumor Suppressor hTid1 Inhibits STAT5b Activity via Functional Interaction

“…Interestingly, interaction with Smad7 was shown to alter the development of chicken embryos by inhibiting the signaling induced by BMP. Although the DnaJ cysteine-rich domain is essential for the hTid1/STAT5b interaction, we cannot rule out the contribution of the DnaJ domain amino terminus in regulating the expression of STAT5b because this domain plays a role in the targeted degradation of intracellular proteins, as has been observed for the IB-IKK complex (42). Although the physiological function of hTid1 in hematopoietic cells remains currently poorly documented, our findings and published reports suggest that hTid1 may regulate the proliferation and/or survival of hematopoietic cells (60).…”

“…The mechanisms involved in hTid1-mediated growth suppression remain elusive because hTid1 modulates the activity of many proteins involved in signal transduction and hence in the survival/apoptosis and/or cell proliferation. Indeed, hTid1 regulates the activity and/or stability of the tyrosine kinase JAK2, the IKK-IK␤ complex, and therefore the activation of NFB as well as the HIF1␣ transcription factor (42,43,63). In addition, hTid1 interacts with the tumor suppressor APC (adenomatous polyposis coli) and regulates the activity of p53 by affecting its subcellular localization (64,65).…”

“…For example, it was shown that hTid1 allowed the degradation of the transcription factor HIF1␣ by the Von Hippel-Lindau protein (43). Similarly, hTid1 may prolong the half-life of IB kinases or act on the degradation of the receptor Erb2 in breast carcinoma cells (42,56). It is therefore conceivable that hTid1 can regulate the stability/degradation of STAT5b.…”

“…hTid1 Inhibits Expression of STAT5b in Ba/F3 Cells-It has been previously shown that hTid1 as a co-chaperone molecule regulates the stability and activity of different viral and cellular proteins (42)(43)(44). We therefore analyzed by Western blot whether hTid1 could affect expression of STAT5b in Ba/F3 cells (Fig.…”