TABLE 2.
Constructs made for the mammalian two-hybrid assaysa
Procedure | Figure | Human, mouse | pGAL4.3 | pACT (protein X) | pBIND (protein Y) | pcDNA3 (vector) | Interaction |
---|---|---|---|---|---|---|---|
A | 7C | Human, mouse | mCherry | Empty | CEP | − | |
Human, mouse | mCherry | ALIX | CEP | ++ | |||
Human, mouse | mCherry | TSG | CEP | + | |||
Human, mouse | mCherry | TX | CEP | +++ | |||
7D | Human, mouse | mCherry | Empty | CEP | − | ||
Human, mouse | mCherry | TX | CEP | +++ | |||
Human, mouse | mCherry | Empty | ALIX | − | |||
Human, mouse | mCherry | TX | ALIX | − | |||
Human, mouse | mCherry | Empty | TSG | +, − | |||
Human, mouse | mCherry | TX | TSG | +, − | |||
7E | Human, mouse | mCherry | Empty | TX | − | ||
Human, mouse | mCherry | CEP | TX | +++ | |||
Human, mouse | mCherry | Empty | TX-AAAX3A | − | |||
Human, mouse | mCherry | CEP | TX-AAAX3A | − | |||
7F | Mouse | mCherry | Empty | CEP | − | ||
Mouse | mCherry | TX | CEP | +++ | |||
Mouse | mCherry | TX-APPX3Y | CEP | +++ | |||
Mouse | mCherry | TX-GAPX3Y | CEP | − | |||
Mouse | mCherry | TX-GPAX3Y | CEP | ++ | |||
Mouse | mCherry | TX-GPPX3A | CEP | − | |||
Mouse | mCherry | TX-AAAX3A | CEP | − | |||
B | 8A and 9, left | Mouse, human | mCherry | Empty | CEP | NA | − |
Mouse, human | mCherry | ALIX | CEP | Empty | ++ | ||
Mouse, human | mCherry | ALIX | CEP | TX | + | ||
Mouse, human | mCherry | ALIX | CEP | TX-AAAX3A | ++ | ||
8B and 9, center | Mouse | mCherry | Empty | CEP | NA | − | |
Mouse | mCherry | TSG | CEP | Empty | + | ||
Mouse | mCherry | TSG | CEP | TX | − | ||
Mouse | mCherry | TSG | CEP | TX-AAAX3A | + | ||
8C and 9, right | Mouse | mCherry | Empty | CEP | NA | − | |
Mouse | mCherry | TX | CEP | Empty | +++ | ||
Mouse | mCherry | TX | CEP | ALIX | +++ | ||
Mouse | mCherry | TX | CEP | TSG | +++ |
The table shows the species from which each protein is derived. In most cases the experiments were performed using both human and mouse TX, ALIX, TSG, and CEP. Boldfacing indicates experiments that are presented in Fig. 7C to F, Fig. 8A to C, and Fig. 9. “Empty” refers to the original pACT and pcDNA vector without insert. For procedure A, the two vectors were cotransfected with the pGL4-mCherry vector. For procedure B, the three vectors were cotransfected with the pGL4-mCherry vector. Under the “pcDNA3 (vector)” column, no additional vector is indicated by “NA.” A pBIND-YFP vector with CEP was used for mammalian two-hybrid analyses instead of a pBIND vector for Fig. 9. The terms “+” and “−” indicate positive and negative interactions, and the number of “+” symbols indicates the strength of the interaction.