Abstract
Activated Akt has been previously implicated in acting on RS domain-containing proteins. However, it has been questioned whether its action is direct or it is mediated by co-existing SR kinase activity. To address this issue we studied in detail the phosphorylation of Lamin B Receptor (LBR) by Akt. Using synthetic peptides and a set of recombinant proteins expressing mutants of the LBR RS domain we now demonstrate that while all serines of the RS domain represent more or less equal phosphoacceptor sites for SRPK1, Ser80 and Ser82 are mainly targeted by Akt. 3D-modeling combined with molecular dynamics (MD) simulations show that amongst short, overlapping LBR RS-containing peptides complying with the minimum Akt recognition consensus sequence, only those bearing phosphosites either at Ser80 or Ser82 are able to fit into the active site of Akt, at least as effectively as its known substrate, GSK3-β. Combined our results provide evidence that Akt kinases directly phosphorylate an RS domain-containing protein and that both the residues N-terminal the phosphosite and at position +1 are essential for Akt specificity, with the latter substrate position being compatible with the arginine residue of RS-repeats.
Highlights
The mammalian genome encodes for more than 100 RS domain-containing proteins, almost half of which are implicated in regulating mRNA processing and the remaining half in a variety of other cellular functions [1]
The RS domain of turkey Lamin B Receptor (LBR) (Fig 1A, sequence underlined) is composed of five consecutive RS repeats which are all targeted by SRPK1 [21, 22]
We used a combination of biochemical approaches and 3D-modeling followed by molecular dynamics (MD) simulations to shed light on the phosphorylation of the RS domain of Lamin B Receptor by activated Akt
Summary
The mammalian genome encodes for more than 100 RS domain-containing proteins, almost half of which are implicated in regulating mRNA processing and the remaining half in a variety of other cellular functions [1]. Arg-Ser dipeptide repeats vary in both length and position and have been implicated in protein-protein interactions as well as in non-sequence specific interactions with RNA molecules [2, 3]. The most studied regulation of RS domain-containing proteins is through phosphorylation of serine residues within the RS domains, catalyzed mainly by PLOS ONE | DOI:10.1371/journal.pone.0154198. RS Domain Phosphorylation by SRPK1 and Akt analysis, decision to publish, or preparation of the manuscript The most studied regulation of RS domain-containing proteins is through phosphorylation of serine residues within the RS domains, catalyzed mainly by PLOS ONE | DOI:10.1371/journal.pone.0154198 April 22, 2016
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