TOE1 is an inhibitor of HIV-1 replication with cell-penetrating capability

Abstract

Target of Egr1 (TOE1) is a nuclear protein localized primarily in nucleoli and Cajal bodies that was identified as a downstream target of the immediate early gene Egr1. TOE1 displays a functional deadenylation domain and has been shown to participate in spliceosome assembly. We report here that TOE1 can function as an inhibitor of HIV-1 replication and show evidence that supports a direct interaction of TOE1 with the viral specific transactivator response element as part of the inhibitory mechanism. In addition, we show that TOE1 can be secreted by activated CD8 + T lymphocytes and can be cleaved by the serine protease granzyme B, one of the main components of cytotoxic granules. Both full-length and cleaved TOE1 can spontaneously cross the plasma membrane and penetrate cells in culture, retaining HIV-1 inhibitory activity. Antiviral potency of TOE1 and its cell-penetrating capability have been identified to lie within a 35-amino-acid region containing the nuclear localization sequence.W e previously reported the cloning of a gene encoding the Target Of Egr1 (TOE1) protein as a target for the immediate early transcription factor Egr1 using ChIP-based methodology and its partial characterization as an inhibitor of cell growth (1). Mechanistically, TOE1 overexpression correlates with up-regulation of the cyclin-dependent kinase inhibitor p21 and produces an accumulation of cells in the G2/M phase of the cell cycle. Subsequently, we found that TOE1 is able to bind to the C-terminal tetramerization domain of the p53 tumor suppressor protein and enhances its transcriptional activity, further underscoring the potential importance of a role for TOE1 in the cellular growth inhibition through cell cycle modulation (2).A previous report from another group studying RNA turnover and investigating human homologs for the yeast Ccr4p and Caf1p/ Pop2p found that TOE1 possesses RNA deadenylase catalytic activity in vitro (3). Interestingly, only transfection of TOE1 with a deletion in its nuclear localization signal, restricting cytoplasmic localization of TOE1, displayed deadenylase activity in cultured cells.Although heterokaryon experiments using HeLa and NIH 3T3 cells detected nucleocytoplasmic shuttling, it remains uncertain whether retention of TOE1 in the cytoplasm occurs to permit effective deadenylation of RNA substrates. An additional report from another group studying Cajal body structure and function identified a role for TOE1 in Cajal body formation, as well as in RNA splicing, although its precise role in RNA splicing is yet to be determined (4). Together, the combination of the published work points to the involvement of TOE1 in the regulation of multiple cellular functions including those involving growth regulation and RNA metabolism.With regard to human disease pathology, clues for a biological role for TOE1 were revealed in a study by investigators in search for the identity of an endogenous noncytotoxic proteinaceous factor secreted from CD8 + T lymphocytes able to control the replication of HIV-1 in infected...

Discovery of another anti-HIV protein in the search for the CD8+ cell anti-HIV Factor

“…This finding, however, was not confirmed by subsequent DNA microarray procedures (22,23). The de Belle research group based their conclusion on TOE1 and CAF from the effect of TOE1 on HIV transcription, its surprising secretion by CD8+ lymphocytes, and its ability to penetrate cells and block HIV replication (17).…”

“…Notably, in their report, Sperandio et al demonstrate that TOE1 is secreted by activated CD8+ T lymphocytes in a full length and cleaved form: Both have antiviral activity (17). The anti-HIV action results from the binding of TOE1 to the TAR sequence in the HIV promotor.…”

“…In PNAS, the research group led by Ian de Belle reports the surprising finding that a nuclear protein, TOE1 (target of EGR1, early growth response 1), could be CAF (17). TOE1 is found primarily in nucleoli and Cajal bodies and is involved in a variety of intracellular events, including deadenylase activity and spliceosome assembly (18,19).…”

“…The small cleaved TOE1 product has a 35-amino acid region comprising the NLS that retains the anti-HIV inhibitory activity involving TAR. This shortened form has the advantage of not being taken up by endosomal-like structures and thus can be readily available for activity (17). See companion article on page E3392.…”

“…The serine protease granzyme B, found in CD8+ cell cytotoxic granules, appears responsible for the cleaved TOE1 products (17). Similarly, CAF seems to undergo cleavage by a serine protease, but it is not Granzyme B (27).…”