Abstract
Sm and Sm-like proteins represent an evolutionarily conserved family with key roles in RNA metabolism. Sm-based regulation is diverse and can range in scope from eukaryotic mRNA splicing to bacterial quorum sensing, with at least one step in these processes being mediated by an RNA-associated molecular assembly built on Sm proteins. Despite the availability of several 3D-structures of Sm-like archaeal proteins (SmAPs), their function has remained elusive. The aim of this study was to shed light on the function of SmAP1 and SmAP2 of the crenarchaeon Sulfolobus solfataricus (Sso). Using co-purification followed by RNASeq different classes of non-coding RNAs and mRNAs were identified that co-purified either with both paralogues or solely with Sso-SmAP1 or Sso-SmAP2. The large number of associated intron-containing tRNAs and tRNA/rRNA modifying RNAs may suggest a role of the two Sso-SmAPs in tRNA/rRNA processing. Moreover, the 3D structure of Sso-SmAP2 was elucidated. Like Sso-SmAP1, Sso-SmAP2 forms homoheptamers. The binding of both proteins to distinct RNA substrates is discussed in terms of surface conservation, structural differences in the RNA binding sites and differences in the electrostatic surface potential of the two Sso-SmAP proteins. Taken together, this study may hint to common and different functions of both Sso-SmAPs in Sso RNA metabolism.
Highlights
The eukaryotic hetero-heptameric Sm proteins bind to single-stranded regions of the U1, U2, U4 and U5 small nuclear RNAs, which are essential for pre-mRNA splicing
E. coli Hfq hexamers have dedicated RNA binding sites, preferably binding uridine-rich stretches of small regulatory RNAs around the central pore on the proximal surface (L3 face) and A-rich sequences on the distal surface (L4 face) [10]
The major structural differences between both Sulfolobus solfataricus (Sso)-Sm-like archaeal proteins (SmAPs) are found in loop L4 and in the N- and C-terminal regions (Figure 1C).The extended N-terminus of Sso-SmAP1 forms a protrusion on the L3 face at the proximal site that results in the formation of distinct grooves between the subunits (Figure S2; left panel), whereas Sso-SmAP2 displays a smoother surface due to the absence of the N-terminal extension (Figure S2; right panel)
Summary
The eukaryotic hetero-heptameric Sm proteins bind to single-stranded regions of the U1, U2, U4 and U5 small nuclear RNAs (snRNAs), which are essential for pre-mRNA splicing. E. coli Hfq hexamers have dedicated RNA binding sites, preferably binding uridine-rich stretches of small regulatory RNAs (sRNAs) around the central pore on the proximal surface (L3 face) and A-rich sequences on the distal surface (L4 face) [10]. The lateral surface of Hfq contributes as well to RNA binding and duplex formation between sRNAs and their target mRNAs [11,12].The dedicated sRNA and mRNA binding surfaces on either site of the Hfq-hexamer may serve to transiently increase the local concentration of two RNA substrates. The inherent capacity of Hfq to induce conformational changes in RNAs together with the observed structural flexibility of RNA ligands bound to Hfq could stochastically facilitate base-pairing [13]
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