Np9 Protein of Human Endogenous Retrovirus K Interacts with Ligand of Numb Protein X

Abstract

We have recently identified Np9 as a novel nuclear protein produced by the human endogenous retrovirus K and were able to document the exclusive presence of np9 transcript in tumors and transformed cells. With the aim of studying whether Np9 has a role in tumorigenesis, a systematic search for interacting proteins was performed. Here, we identify the RING-type E3 ubiquitin ligase LNX (ligand of Numb protein X) as an Np9-interacting partner. We furthermore show that the interaction involves N-and C-terminal domains of both proteins and can affect the subcellular localization of LNX. LNX has been reported to target the cell fate determinant and Notch antagonist Numb for proteasome-dependent degradation, thereby causing an increase in transactivational activity of Notch. We document that LNX-interacting Np9, like Numb, is unstable and degraded via the proteasome pathway and that ectopic Numb can stabilize recombinant Np9. Combined, these findings point to the possibility that Np9 affects tumorigenesis through the LNX/Numb/Notch pathway.Up to 8% of the human genome consists of retrovirusderived sequences (19). They are the result of numerous infections and subsequent integration of proviral elements into the germ line of human ancestors during the past 40 million years (2). Most of the recent human endogenous retrovirus (HERV) elements are defective due to the accumulation of mutations. Nonetheless, retroviral genes are frequently expressed in normal and transformed human tissues. The function of these genes in normal human cell physiology is still unclear (32). So far, only the expression of the env gene of HERV-W is known to have a crucial function in normal organismal physiology, namely human trophoblast cell fusion and differentiation (5,12,23). In contrast, retroviral gene expression has been linked to several human diseases, including tumorigenesis. In particular, expression of HERV-K seems to be strongly associated with transformation. Early hints pointing in this direction came from the observation that expression of the HERV-K proteins Gag and Env, as well as antibodies directed against these proteins, could be specifically detected in patients suffering from germ cell tumors (17,24,30,31). Other studies implicate a contribution of HERV-K10 gag RNA expression to the development of leukemia (9) or show that env as well as the expression of subgenomic env transcripts correlates with human breast cancer (35,36).It is unclear, at the present, how these proteins may support tumorigenesis. The Rev-like regulatory protein Rec (20,21)

Quantitation of HERV-K env gene expression and splicing in human breast cancer

“…This result provides support for the idea that cORF rather than full-length env protein is responsible for cell transformation. Recently, a novel gene, np9, resulting from use of a unique HERV-K type 1-specific splice donor site, was confirmed to be expressed in various tumor tissues and transformed cell lines but not in normal, nontransformed cells, suggesting that the protein may function in tumorigenesis (Armbruester et al, 2002). It remains to be seen whether the variants that we have identified here will possess tumorigenic properties similar to the np9 variant just reported.…”

Retroelements and the human genome: New perspectives on an old relation

“…Armbruester et al (24) found np9 transcripts in most transformed human cell lines by using an RT-PCR method. Interestingly, nearly all healthy and malignant tissues from humans express full-length HERV-K type 1 mRNAs, but no np9 transcripts are generated in normal human tissues.…”

“…Recently, a HERV-K type 1-specific transcript coding for a 9-kDa protein, designated Np9, has been identified (24). This transcript represents a kind of type 1 substitute for the Rec sequence of type 2 proviruses.…”

“…23). The nucleotide stretch missing in type 1 sequences harbors a splice donor site for the generation of doubly spliced transcripts coding for the two exons of the Rec protein (23,24). Because of the absence of this splice donor site, (25).…”

Regulation of the Human Endogenous Retrovirus K (HML-2) Transcriptome by the HIV-1 Tat Protein

“…In fact, Np9 and Rec have been shown to be highly expressed in transformed tissues (Np9), to cause carcinoma in situ in mice (Rec), and to be associ-ated with the Notch pathway and pathways affecting c-myc expression (Np9 and both, respectively) (13,15,16,(62)(63)(64)(65). Our laboratory has noted high expression levels of both proteins during HIV-1-associated lymphoma (unpublished observations).…”