Rbp9 is an RNA binding protein expressed mainly in the central nervous system of adult Drosophila melanogaster. Rbp9 shares a high degree of sequence similarity with human neural proteins referred to as Hu antigens. Hu antigens bind to U-rich mRNA destabilizing elements with a high affinity and, thus, have been implicated as regulators of mRNA stability. Using in vitro RNA binding assays, we found that Rbp9 binds strongly to poly U sequences. We then employed a Selex system to identify a consensus Rbp9 binding site (UUUXUUUU). Information obtained from the Selex results allowed the detection of two repeats of the Rbp9 consensus binding sequence in the 3' untranslated region of extramacrochaetae mRNA. UV crosslinking experiments demonstrated that Rbp9 interacts specifically with emc mRNA. The requirement of Rbp9 protein in the down regulation of emc mRNA was confirmed by northern (RNA) analysis, which revealed that the level of emc mRNA increased 10-fold in rbp9 mutant flies. Taken together with the in vitro RNA binding results, the genetic evidence obtained strongly supports the hypothesis that Rbp9 functions as a regulator of RNA stability.
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