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Arginine methylation of hnRNPK negatively modulates apoptosis upon DNA damage through local regulation of phosphorylation.

Yang JH, Chiou YY, Fu SL, Shih IY, Weng TH, Lin WJ, Lin CH - Nucleic Acids Res. (2014)

Bottom Line: In addition, increased hnRNPK expression has been associated with tumor development and progression.In the present study, we demonstrated that the methylation of two essential arginines, Arg296 and Arg299, on hnRNPK inhibited a nearby Ser302 phosphorylation that was mediated through the pro-apoptotic kinase PKCδ.While such elevated apoptosis can be diminished through addition with wild-type hnRNPK, we further demonstrated that this increased apoptosis occurred through both intrinsic and extrinsic pathways and was p53 independent, at least in part.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences and Institute of Genome Sciences, National Yang Ming University, Taipei 11221, Taiwan.

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Arg296 and Arg299 are conserved methylation sites located near the PKCδ-mediated phosphorylation site of hnRNPK (a) Alignment of the arginine methylation sites within the Akt consensus phosphorylation motif (RXRXXS/T) in two known Akt substrates, FOXO1 and BAD. The conserved arginine methylation sites by PRMT1 in AKT substrates are shaded in gray, and an asterisk indicates the Akt phosphorylation sites. (b) Alignment of Arg296 and Arg299 methylation sites within the PKCδ consensus phosphorylation motif (RXXSRXR) of human hnRNPK with two other vertebrate hnRNPKs. The conserved arginine methylation sites by PRMT1 in hnRNPK are shaded in gray, and an asterisk indicates the known PKCδ phosphorylation sites.
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Figure 4: Arg296 and Arg299 are conserved methylation sites located near the PKCδ-mediated phosphorylation site of hnRNPK (a) Alignment of the arginine methylation sites within the Akt consensus phosphorylation motif (RXRXXS/T) in two known Akt substrates, FOXO1 and BAD. The conserved arginine methylation sites by PRMT1 in AKT substrates are shaded in gray, and an asterisk indicates the Akt phosphorylation sites. (b) Alignment of Arg296 and Arg299 methylation sites within the PKCδ consensus phosphorylation motif (RXXSRXR) of human hnRNPK with two other vertebrate hnRNPKs. The conserved arginine methylation sites by PRMT1 in hnRNPK are shaded in gray, and an asterisk indicates the known PKCδ phosphorylation sites.

Mentions: The interplay between diverse PTMs, including methylation, phosphorylation and acetylation, has been shown to regulate histone protein functions (37,38). In addition, arginine methylation has also been proposed to regulate protein–protein interactions (39–41). Accordingly, we further investigated the methylated arginine-containing sequences of hnRNPK via bioinformatics to determine whether Arg296 and Arg299 methylation have functions distinct from other methylable arginine residues. Several studies have revealed that arginine methylation inhibits the nearby phosphorylation level on the same protein and thus regulates phosphorylation-mediated functions (34,42–44), i.e. human FOXO1 and Bcl2-associated agonist of cell death (BAD) . Further comparison of the methylated arginine-containing sequences among FOXO1, BAD and hnRNPK revealed that Arg296 and Arg299 in hnRNPK are only three residues away from Ser302, similar to the locations of these residues in FOXO1 and BAD (Figure 4a). Ser302 phosphorylation is mediated through PKCδ (24). In addition, it has been suggested that Arg299 is an essential residue in the consensus substrate sequence of PKCδ (RXXSXSR), as shown in Figure 4b (24). Therefore, we further determined whether hnRNPK methylation at Arg296 and Arg299 regulates the interaction of this protein with PKCδ. The results of the co-immunoprecipitation of WT or 2RK hnRNPK with PKCδ in U2OS cells revealed that the loss of hnRNPK arginine methylation on Arg296 and Arg299 increased the interaction of this protein with PKCδ (Supplementary Figure S1).


Arginine methylation of hnRNPK negatively modulates apoptosis upon DNA damage through local regulation of phosphorylation.

Yang JH, Chiou YY, Fu SL, Shih IY, Weng TH, Lin WJ, Lin CH - Nucleic Acids Res. (2014)

Arg296 and Arg299 are conserved methylation sites located near the PKCδ-mediated phosphorylation site of hnRNPK (a) Alignment of the arginine methylation sites within the Akt consensus phosphorylation motif (RXRXXS/T) in two known Akt substrates, FOXO1 and BAD. The conserved arginine methylation sites by PRMT1 in AKT substrates are shaded in gray, and an asterisk indicates the Akt phosphorylation sites. (b) Alignment of Arg296 and Arg299 methylation sites within the PKCδ consensus phosphorylation motif (RXXSRXR) of human hnRNPK with two other vertebrate hnRNPKs. The conserved arginine methylation sites by PRMT1 in hnRNPK are shaded in gray, and an asterisk indicates the known PKCδ phosphorylation sites.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150800&req=5

Figure 4: Arg296 and Arg299 are conserved methylation sites located near the PKCδ-mediated phosphorylation site of hnRNPK (a) Alignment of the arginine methylation sites within the Akt consensus phosphorylation motif (RXRXXS/T) in two known Akt substrates, FOXO1 and BAD. The conserved arginine methylation sites by PRMT1 in AKT substrates are shaded in gray, and an asterisk indicates the Akt phosphorylation sites. (b) Alignment of Arg296 and Arg299 methylation sites within the PKCδ consensus phosphorylation motif (RXXSRXR) of human hnRNPK with two other vertebrate hnRNPKs. The conserved arginine methylation sites by PRMT1 in hnRNPK are shaded in gray, and an asterisk indicates the known PKCδ phosphorylation sites.
Mentions: The interplay between diverse PTMs, including methylation, phosphorylation and acetylation, has been shown to regulate histone protein functions (37,38). In addition, arginine methylation has also been proposed to regulate protein–protein interactions (39–41). Accordingly, we further investigated the methylated arginine-containing sequences of hnRNPK via bioinformatics to determine whether Arg296 and Arg299 methylation have functions distinct from other methylable arginine residues. Several studies have revealed that arginine methylation inhibits the nearby phosphorylation level on the same protein and thus regulates phosphorylation-mediated functions (34,42–44), i.e. human FOXO1 and Bcl2-associated agonist of cell death (BAD) . Further comparison of the methylated arginine-containing sequences among FOXO1, BAD and hnRNPK revealed that Arg296 and Arg299 in hnRNPK are only three residues away from Ser302, similar to the locations of these residues in FOXO1 and BAD (Figure 4a). Ser302 phosphorylation is mediated through PKCδ (24). In addition, it has been suggested that Arg299 is an essential residue in the consensus substrate sequence of PKCδ (RXXSXSR), as shown in Figure 4b (24). Therefore, we further determined whether hnRNPK methylation at Arg296 and Arg299 regulates the interaction of this protein with PKCδ. The results of the co-immunoprecipitation of WT or 2RK hnRNPK with PKCδ in U2OS cells revealed that the loss of hnRNPK arginine methylation on Arg296 and Arg299 increased the interaction of this protein with PKCδ (Supplementary Figure S1).

Bottom Line: In addition, increased hnRNPK expression has been associated with tumor development and progression.In the present study, we demonstrated that the methylation of two essential arginines, Arg296 and Arg299, on hnRNPK inhibited a nearby Ser302 phosphorylation that was mediated through the pro-apoptotic kinase PKCδ.While such elevated apoptosis can be diminished through addition with wild-type hnRNPK, we further demonstrated that this increased apoptosis occurred through both intrinsic and extrinsic pathways and was p53 independent, at least in part.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences and Institute of Genome Sciences, National Yang Ming University, Taipei 11221, Taiwan.

Show MeSH
Related in: MedlinePlus