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A
RNA immunoprecipitation (RIP) results showing the interaction of BTG4 with indicated transcripts, in the presence or absence of PABPN1L (full length, RRM‐deleted, or R171A mutant). HeLa cells were co‐transfected with plasmids expressing FLAG‐tagged PABPN1L and HA‐tagged BTG4 for 48 h before immunoprecipitating with an anti‐HA antibody. RNAs recovered from the immunoprecipitants were subjected to RT–qPCR of the indicated transcripts. Fold change values of both input and IP samples were normalized by RIP results of HA‐BTG4 and FLAG‐PABPN1L co‐expression groups. n = 3 biological replicates. Error bars, SEM. The P‐value represents the two‐tailed Student's t‐test comparing the RIP results of BTG4 with the indicated transcripts in the presence of PABPN1L, *P < 0.05, **P < 0.01, and ***P < 0.001.
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B
Diagrams of mouse BTG4 constructs.
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C, D
Co‐IP and Western blot results showing interactions between PABPN1L and BTG4. Lysates from HeLa cells expressing HA‐BTG4 (WT or mutants shown in (B)) and FLAG‐PABPN1L were immunoprecipitated with an anti‐HA antibody. The immunoprecipitated proteins are detected by Western blot with the indicated antibodies.
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E
Diagrams of mouse BTG2 and BTG4 constructs, and co‐IP results showing that PABPN1L binds to BTG4 but not BTG2.
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F
Diagrams and co‐IP results showing BTG4 binding to PABPN1L. Lysates from HeLa cells expressing HA‐BTG4 and FLAG‐PABPN1L (full length and C‐terminal deleted) were immunoprecipitated with an anti‐FLAG antibody. The immunoprecipitated proteins are detected by Western blot with the indicated antibodies.
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G
Diagrams of PABPN1 and PABPN1L constructs, and co‐IP results showing that PABPN1L, but not PABPN1, binds to BTG4.
Data information: For all Western blot results, at least three independent experiments were done with consistent results.
