Biochemistry as a Tool TOR and TORC Protein Interactions

Biochemical analysis of TOR binding partners is hindered by the fact that AtTOR accumulates in only a limited set of plant tissues. This set includes embryos, endosperm, shoot and root meristems, and cells recently emergent from the meristem (Menand et al. 2002). None of these cell types are conducive to biochemical analysis. AtTOR transcripts accumulate in a broader range of tissues (Robaglia et al. 2004). The discrepancy in TOR transcript accumulation and protein accumulation may be due to a small regulatory ORF in the TOR mRNA 5' untranslated region (Menand et al. 2004; Robaglia et al. 2004). Alternately, a TOR-specific microRNA, mi34, conserved from Arabidopsis to rice (Bonnet et al. 2004) may regulate TOR translational regulation.

To overcome this obstacle, researchers (Mahfouz et al. 2006) have employed a tobacco transfection system (Voinnet et al. 2003) to test for interactions among TOR complex proteins. In this system, AtRaptor1B was shown to interact with the AtTOR N-terminal HEAT repeats (Mahfouz et al. 2006). This interaction is consistent with TOR-Raptor interactions in other systems, and is the only direct biochemical evidence to date for a plant TORC1 complex.

AtS6K1, one of two Arabidopsis ribosomal protein S6 kinases, interacts with AtRaptor1B in vivo in transfected tobacco (Mahfouz et al. 2006). In this system, AtS6K1 kinase activity was inhibited by osmotic stress. Cotransfec-tion with AtRaptor1B restored AtS6K1 kinase activity under osmotic stress (Mahfouz et al. 2006).

The only other known binding partner of AtRaptor1B, identified thorough a targeted yeast two-hybrid assay, is AML1 (Anderson and Hanson 2005). As mentioned above, Mei2p interacts with fission yeast Mip1p/Raptor (Shinozaki-Yabana et al. 2000), suggesting that AML1 and Mei2p are conserved TOR substrates, and AML1 overexpression suppresses some fission yeast meiosis signaling defects, suggesting a conserved function (Hirayama et al. 1997).

In summary, work using a tobacco transfection system indicates that a Raptor-TOR interaction mediated by the TOR HEAT repeats is conserved in plants. This TOR-Raptor interaction is the hallmark of the nutrient- and rapamycin-sensitive complex TORC1. Two proteins, AtS6K1 and AML1, have been shown to interact with AtRaptor1B, marking them as conserved TOR effectors.

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