The La-related proteins LARP4B, LARP4, and LARP1 interact with an overlapping set of mRNAs. (A) Comparison of interacting mRNAs of the LARP4B, LARP4, and LARP1 proteins. To identify mRNAs bound by LARP4 and LARP1, the FLAG/HA-tagged proteins were immunoprecipitated from cell extract of stably overexpressing HEK293 cell lines (lanes 6 and 10). For control a nonoverexpressing HEK293 cell line was used (lanes 8 and 12). The copurified RNA was analyzed by RT-PCR, which revealed an overlapping set of target mRNAs for the La-related proteins 4B, 4, and 1 (LARP4B immunoprecipitation identical to Fig. 2A). Western blot analysis confirmed the immunoprecipitation of the antigens. (B) Effect of LARP4B, LARP4, and LARP1 on luciferase reporter expression. LARP4B, LARP4, and LARP1 and combinations thereof were knocked down and a reporter plasmid encoding a firefly luciferase was transiently transfected. Relative luciferase activity of the prepared extracts was determined (left panel). Results were deduced from at least three independent biological replicates. A Western blot analysis confirmed the knockdown of LARP4B, LARP4, and LARP1 (right panel). Numbers indicate knockdown efficiency.
