TABLE 1.
Summary of all mutant overexpression EhAgo cell lines used in this study and effect on protein localization and sRNA bindinga
| Cell line | Mutation(s) | Protein localization | sRNA binding |
|---|---|---|---|
| Wild type | |||
| Myc-EhAgo2-1 | None | Cytoplasm/perinuclear ring | Yes |
| Myc-EhAgo2-2 | None | Nucleus | Yes |
| Myc-EhAgo2-3 | None | Cytoplasm/perinuclear ring | Yes |
| PAZ mutants | |||
| Myc-EhAgo2-1PAZ-mut | Y267A, Y268A | Cytoplasm | No |
| DHFR-Myc-EhAgo2-2PAZ-mut | Y267A, Y268A | Nucleus | No |
| Myc-EhAgo2-3PAZ-mut | Y259A, Y260A | Cytoplasm | No |
| EhAgo2-2 C-terminal mutants | |||
| Myc-EhAgo2-2ΔDR | 783–937 deletion | Cytoplasm | Yes |
| Myc-EhAgo2-2ΔNLS-DR | 761–937 deletion | Cytoplasm | Yes |
| EhAgo2-2 PIWI mutants | |||
| Myc-EhAgo2-2D545A | D545A | Nucleus | Yes |
| Myc-EhAgo2-2N747A | N747A | Nucleus | Yes |
| Myc-EhAgo2-2D545A;N747A | D545A, N747A | Nucleus | Yes |
| Chimeric protein | |||
| Myc-EhAgo2-3 plus C-terminal tail region of EhAgo2-2 |
Full-length EhAgo2-3 fused with EhAgo2-2 tail (761–937) |
Nucleus | Yes |
a
Listed are cell lines, specific mutations, protein localizations determined by anti-Myc IFA, and sRNA binding ability determined by anti-Myc IPs and pCp labeling of sRNAs.