Molecular aspects of RNA-recognition by the neuronal mRNA transport factor Staufen 2

Abstract

Staufen (Stau) is a double-stranded RNA-binding protein (dsRBP) originally identified as an mRNA transport factor and germ-cell marker in Drosophila. The mammalian homolog Staufen 2 (Stau2) is specifically expressed in neurons, where it clusters in RNA granules and moves along microtubules from cell bodies to dendrites. Stau2 is an important core factor for neuronal mRNA transport and has been implicated in synaptic plasticity, memory and learning. Stau2 knockdown in rats results in reduced numbers of mature dendritic spines and impairs long-term depression of synapses. All Stau proteins contain four to five dsRNA-binding domains (RBDs). For all Stau proteins RBD3 and RBD4 are thought to be the only active RNA-binding domains, whereas RBDs 1, 2 and 5 are considered to be pseusoRBDs, which retained the fold but no activity for RNA binding. To date, the mechanistic basis of Stau2 function in the cell is not well understood. The translational repressor Pumilio 2 (Pum2) acts in the same pathways as Stau2 by forming joint messenger ribonucleoproteins (mRNPs). The 3’ untranslated region (UTR) of the Stau2 target mRNA Regulator of G-protein signaling 4 (Rgs4) comprises two predicted Staufen-recognized structures (SRSs) and two Pum2 binding sites in close proximity, suggesting a functional complex of Stau2, Pum2 and Rgs4. In this work, it could be confirmed that Stau2 and Pum2 both bind to the Rgs4 3’UTR. However, a functional complex of Stau2 and Pum2 with Rgs4 that could mediate their role in common biological processes could not be confirmed. This work shows that in Stau2 also the non-canonical RBDs 1 and 2 exhibit RNA-binding activity. Their RNA-binding activity was characterized by biochemical and biophysical methods and an important role for both domains in RNA-recognition is demonstrated. Although RBDs 3 and 4 alone achieve high affinity binding, RBDs 1 and 2 are required to form stable complexes with RNA. This feature might be of great importance for biological target recognition and the transport of RNAs over longer distances.