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The multifunctional poly(A)-binding protein (PABP) 1 is subject to extensive dynamic post-translational modification, which molecular modelling suggests plays an important role in co-ordinating its activities.

Brook M, McCracken L, Reddington JP, Lu ZL, Morrice NA, Gray NK - Biochem. J. (2012)

Bottom Line: Intriguingly, PABP1 contains glutamate and aspartate methylations, modifications of unknown function in eukaryotes, as well as lysine and arginine methylations, and lysine acetylations.The latter dramatically alter the pI of PABP1, an effect also observed during the cell cycle, suggesting that different biological processes/stimuli can regulate its modification status, although PABP1 also probably exists in differentially modified subpopulations within cells.Modelling using available structures implicates these modifications in regulating interactions with individual PAM2 (PABP-interacting motif 2)-containing proteins, suggesting a direct link between PABP1 modification status and the formation of distinct mRNP (messenger ribonucleoprotein) complexes that regulate mRNA fate in the cytoplasm.

View Article: PubMed Central - PubMed

Affiliation: MRC Centre for Reproductive Health/MRC Human Reproductive Sciences Unit, Queen's Medical Research Institute, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, Scotland, UK. matt.brook@ed.ac.uk

ABSTRACT
PABP1 [poly(A)-binding protein 1] is a central regulator of mRNA translation and stability and is required for miRNA (microRNA)-mediated regulation and nonsense-mediated decay. Numerous protein, as well as RNA, interactions underlie its multi-functional nature; however, it is unclear how its different activities are co-ordinated, since many partners interact via overlapping binding sites. In the present study, we show that human PABP1 is subject to elaborate post-translational modification, identifying 14 modifications located throughout the functional domains, all but one of which are conserved in mouse. Intriguingly, PABP1 contains glutamate and aspartate methylations, modifications of unknown function in eukaryotes, as well as lysine and arginine methylations, and lysine acetylations. The latter dramatically alter the pI of PABP1, an effect also observed during the cell cycle, suggesting that different biological processes/stimuli can regulate its modification status, although PABP1 also probably exists in differentially modified subpopulations within cells. Two lysine residues were differentially acetylated or methylated, revealing that PABP1 may be the first example of a cytoplasmic protein utilizing a 'methylation/acetylation switch'. Modelling using available structures implicates these modifications in regulating interactions with individual PAM2 (PABP-interacting motif 2)-containing proteins, suggesting a direct link between PABP1 modification status and the formation of distinct mRNP (messenger ribonucleoprotein) complexes that regulate mRNA fate in the cytoplasm.

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Post-translational modification of PABP1 is not restricted to arginine methylation(A) HeLa cells were either left untreated (Control) or treated with 20 μM AdOX. Cell extracts were subjected to OFFGEL isoelectric fractionation using a pH 3–10 linear immobilized pH gradient (IPG) and fractions immunoblotted for PABP1 and α-tubulin. (B) Cell extracts from Prmt4+/+ and Prmt4−/− MEFs were fractionated and immunoblotted as described in (A) using a pH 6–11 gradient and GAPDH as a control. A longer exposure of the PABP1 blots is shown to visualize low abundance highly modified forms of PABP1. (A) α-Tubulin [40] and (B) GAPDH [41] exhibit expected pI distributions comprising unmodified and modified forms.
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Figure 3: Post-translational modification of PABP1 is not restricted to arginine methylation(A) HeLa cells were either left untreated (Control) or treated with 20 μM AdOX. Cell extracts were subjected to OFFGEL isoelectric fractionation using a pH 3–10 linear immobilized pH gradient (IPG) and fractions immunoblotted for PABP1 and α-tubulin. (B) Cell extracts from Prmt4+/+ and Prmt4−/− MEFs were fractionated and immunoblotted as described in (A) using a pH 6–11 gradient and GAPDH as a control. A longer exposure of the PABP1 blots is shown to visualize low abundance highly modified forms of PABP1. (A) α-Tubulin [40] and (B) GAPDH [41] exhibit expected pI distributions comprising unmodified and modified forms.

Mentions: Since the pI of unmodified human PABP1 is pH 9.52, its distribution in HeLa cells (between ~pH 7.6 and 10) is consistent with multiple PTMs. This distribution was only slightly altered following AdOX treatment (Figure 3A), indicating the presence of PTMs other than arginine methylation. Similarly, in both Prmt4+/+ and Prmt4−/− MEFs a small proportion of PABP1 is detected in fractions containing proteins with significantly lower pI values (Figure 3B; e.g. fractions 1–4 represent a pI range ~pH 6.0–7.7). Since its pI distribution is not significantly affected by arginine methylation, PABP1 appears to be modified by multiple PTM species.


The multifunctional poly(A)-binding protein (PABP) 1 is subject to extensive dynamic post-translational modification, which molecular modelling suggests plays an important role in co-ordinating its activities.

Brook M, McCracken L, Reddington JP, Lu ZL, Morrice NA, Gray NK - Biochem. J. (2012)

Post-translational modification of PABP1 is not restricted to arginine methylation(A) HeLa cells were either left untreated (Control) or treated with 20 μM AdOX. Cell extracts were subjected to OFFGEL isoelectric fractionation using a pH 3–10 linear immobilized pH gradient (IPG) and fractions immunoblotted for PABP1 and α-tubulin. (B) Cell extracts from Prmt4+/+ and Prmt4−/− MEFs were fractionated and immunoblotted as described in (A) using a pH 6–11 gradient and GAPDH as a control. A longer exposure of the PABP1 blots is shown to visualize low abundance highly modified forms of PABP1. (A) α-Tubulin [40] and (B) GAPDH [41] exhibit expected pI distributions comprising unmodified and modified forms.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3298439&req=5

Figure 3: Post-translational modification of PABP1 is not restricted to arginine methylation(A) HeLa cells were either left untreated (Control) or treated with 20 μM AdOX. Cell extracts were subjected to OFFGEL isoelectric fractionation using a pH 3–10 linear immobilized pH gradient (IPG) and fractions immunoblotted for PABP1 and α-tubulin. (B) Cell extracts from Prmt4+/+ and Prmt4−/− MEFs were fractionated and immunoblotted as described in (A) using a pH 6–11 gradient and GAPDH as a control. A longer exposure of the PABP1 blots is shown to visualize low abundance highly modified forms of PABP1. (A) α-Tubulin [40] and (B) GAPDH [41] exhibit expected pI distributions comprising unmodified and modified forms.
Mentions: Since the pI of unmodified human PABP1 is pH 9.52, its distribution in HeLa cells (between ~pH 7.6 and 10) is consistent with multiple PTMs. This distribution was only slightly altered following AdOX treatment (Figure 3A), indicating the presence of PTMs other than arginine methylation. Similarly, in both Prmt4+/+ and Prmt4−/− MEFs a small proportion of PABP1 is detected in fractions containing proteins with significantly lower pI values (Figure 3B; e.g. fractions 1–4 represent a pI range ~pH 6.0–7.7). Since its pI distribution is not significantly affected by arginine methylation, PABP1 appears to be modified by multiple PTM species.

Bottom Line: Intriguingly, PABP1 contains glutamate and aspartate methylations, modifications of unknown function in eukaryotes, as well as lysine and arginine methylations, and lysine acetylations.The latter dramatically alter the pI of PABP1, an effect also observed during the cell cycle, suggesting that different biological processes/stimuli can regulate its modification status, although PABP1 also probably exists in differentially modified subpopulations within cells.Modelling using available structures implicates these modifications in regulating interactions with individual PAM2 (PABP-interacting motif 2)-containing proteins, suggesting a direct link between PABP1 modification status and the formation of distinct mRNP (messenger ribonucleoprotein) complexes that regulate mRNA fate in the cytoplasm.

View Article: PubMed Central - PubMed

Affiliation: MRC Centre for Reproductive Health/MRC Human Reproductive Sciences Unit, Queen's Medical Research Institute, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, Scotland, UK. matt.brook@ed.ac.uk

ABSTRACT
PABP1 [poly(A)-binding protein 1] is a central regulator of mRNA translation and stability and is required for miRNA (microRNA)-mediated regulation and nonsense-mediated decay. Numerous protein, as well as RNA, interactions underlie its multi-functional nature; however, it is unclear how its different activities are co-ordinated, since many partners interact via overlapping binding sites. In the present study, we show that human PABP1 is subject to elaborate post-translational modification, identifying 14 modifications located throughout the functional domains, all but one of which are conserved in mouse. Intriguingly, PABP1 contains glutamate and aspartate methylations, modifications of unknown function in eukaryotes, as well as lysine and arginine methylations, and lysine acetylations. The latter dramatically alter the pI of PABP1, an effect also observed during the cell cycle, suggesting that different biological processes/stimuli can regulate its modification status, although PABP1 also probably exists in differentially modified subpopulations within cells. Two lysine residues were differentially acetylated or methylated, revealing that PABP1 may be the first example of a cytoplasmic protein utilizing a 'methylation/acetylation switch'. Modelling using available structures implicates these modifications in regulating interactions with individual PAM2 (PABP-interacting motif 2)-containing proteins, suggesting a direct link between PABP1 modification status and the formation of distinct mRNP (messenger ribonucleoprotein) complexes that regulate mRNA fate in the cytoplasm.

Show MeSH
Related in: MedlinePlus