1. Introduction
Discovering Relationships Between Lysine Ubiquitylation
and Acetylation Sites from Proteomic Datasets
Jake Elwood†
, Jennifer Maki
†
, and Dana M. Freund‡
†
The College of Saint Scholastica Department of Chemistry
‡
University of Minnesota Department of Horticultural Science
Results
Conclusions and future work
Results
For the last four decades, post translational modifications (PTMs) have been
associated with altering the structure and function of nuclear associated proteins.
However, recent research has shown that PTMs play a large role in increasing
the proteomic diversity of non-nuclear proteins as well, and that different PTMs
often occur on the same protein1. This research focuses on the potential
relationships between the ubiquitylation (Ub) and the newly found acetylation
(Ac) PTMs that occur in non-nuclear Arabidopsis thaliana proteins.
Ubiquitylation and acetylation are post translational modifications that both occur
on lysine residues. Ac of non-nuclear proteins is a new area of research,
especially in the plant biology field. Ac of proteins can alter enzyme activities and
protein functions, while Ub is well known for marking proteins for degradation by
the proteasome as well as many other cellular functions. Recently, it has been
suggested that competition may exist between Ub and Ac of specific lysine
residues. It may be that the neutralization of the lysine side chain caused by Ac
prevents Ub from taking place. If proven, this Ac-Ub competition has the potential
to affect processes such as the regulation of the cell cycle and protein
degradation. The purpose of this study was to manually data mine previously
published Ac and Ub sites in Arabidopsis thaliana to determine the amount of
overlap. Sites identified in three publications were compared: Finkemeier et al.
(2011) Plant Physiology, 155:1779-90; Kim et al. (2013) The Plant Cell, 25: 1523-
40; and Wu et al. (2011) Plant Physiology, 155:1769-78.
Background
Acknowledgements
The National Science Foundation grants IOS-1400818 & IOS-1238812
Drs. Adrian D. Hegeman & Jerry D. Cohen, University of Minnesota Department of
Horticultural Science
Function of non-nuclear acetylation
• Down regulation of ATP synthase
activity3
• Role in Energy metabolism3
• Affects Rubisco activity4
• Potential role in protein degradation4
• Links apoptosis to metabolism5
• Protein stability5
Function of non-nuclear ubiquitylation
• Developmental progression6
• Protein trafficking6
• Hormone Signaling6
• Proteolysis7
Protein Name
Site #
(Ac) Peptide (Ac)
Site #
(Ub) Peptide (Ub)
Phosphoglycerate kinase 1
K146 K.GVTPK(ac)FSLAPLVPR.L
ATP synthase subunit beta
K154
K178
K.LSIFETGIK(ac)VVDLLAPYRR.G
IGLFGGAGVG(ac)KTVLI(ox)MELINNIAK
Glutathione synthetase
K66 K.MESQK(ac)PIFDLEKLDDEFVQK.L
Glutamine synthetase
K223 K.WPLGWPVGAFPGPQGPYYCGVGADK(ac)I
WGR.D
Protein argonaute 1 K837 RSTGH(ac)KPLR
Phosphatidylinositol/
phosphatidylcholine transfer
protein SFH4
K283 SFLDP(ac)KTVSK K380 VSDTSTAK(ub)SGS
ELEEMASPK
Elongation factor 1-alpha 4
K36
K79
K178
K187
K227
K306
LGGID(ac)KR
GITIDIALW(ac)KFETTK
IGLFGGAGVG(ac)KTVLI(ox)MELINNIAK
VGYNPD(ac)KIPFVPISGFEGDNMIER
GPTLLEALDQINEP(ac)KRPSDKPLR
NVAV(ac)KDLK
K438
K441
DPTGAK(ub)VTK(ub)
AAVK
ADP-ribosylation factor 2-A
K16 K(ac)EMRILMVGLDAAGK
60S ribosomal protein L10a-2
K90 MGLSNMDVEAL(ac)KK
To date, 121 acetylated and approximately 1200 ubquitylated non-nuclear Arabidopsis thaliana
protein sites have been recorded. Thus far, nine of the identified proteins experience both, Ac and
Ub, post translational modifications.
References
1. Varshavsky, Alexander (2006) The early history of the ubiquitin field. Protein Science 15(3), 647-654
2. Wu, X., Oh, M.-H., Schwarz, E. M., Larue, C. T., Sivaguru, M., Imai, B. S., Yau P. M., Ort, D.R., & Huber, S. C. (2011) Lysine acetylation is a
widespread protein modification for diverse proteins in Arabidopsis. Plant physiol, 155(4), 1769–78
3. König, A.-C., Hartl, Markus, Pham, P. A., et al., (2014) The Arabidopis Class II Sirtuin Is a Lysine Deacetylase and Interacts with Mitochondrial
Energy Metabolism. Plany physiol, 164(3), 1401-14
4. Finkemeier, I., Laxa, M., Miguet, L., Howden, A. J. M., &Sweetlove, L. J. (2011) Proteins of diverse function and subcellular location are lysine
acetylated in Arabidopsis. Plant physiol, 155(4), 1779–90
5. Xing, S., Yves, P. (2012) The protein acetylome and the regulation of metabolism. Trends in Plant Science, 17(7), 423-30
6. Saracco, SA., Hansson, M., Scalf, M., Walker, JM., Smith, LM., Vierstra, RD. (2009) Tandem affinity purification and mass spectrometric
analysis of ubiquitylated proteins in Arabidopsis. Plant J. 59(2), 344-58
7. Book, AF., Gladman, NP., Lee, SS., Scalf, M., Smith, LM., Vierstra, RD. (2010) Affinity purification of the Arabidopsis 26 S proteasome reveals
a diverse array of plant proteolytic complexes. J Biol Chem, 285(33), 25554-69
8. Kim, D.-Y., Scalf, Mark., Smith, Lloyd., Vierstra, Richard. (2013) Advanced Proteomic Analyses Yield Deep Catalog Ubiquitylation Targets in
Arabidopsis. The Plant Cell Preview 25(5), 1523-1540
9. www.uniprot.org
The findings of non-nuclear proteins that contain both, Ub and Ac, post translational
modifications is encouraging. The discovery that the majority of these proteins were
located within energy associated organelles contributes to the hypothesis that Ub and Ac
cross talk has a potential influence on metabolic and energy associated pathways.
In the future, more Arabidopsis thaliana protein acetylation data is needed, as only
around 120 acetylated Arabidopsis sites were compared to a list of approximately 1200
known Ub proteins. An increase in Ub data is also necessary to further investigate
whether or not acetylation has a direct or indirect influence on ubiquitylation. Many of the
published proteins lacked information indicating the specific ubiquitylated residue.
The findings of this research are to be submitted to
http://www.p3db.org/acetylation_datasets.php.
Table 1. Proteins that were shown to undergo ubiquitylation and acetylation
Figure 1. Overlapping proteins were categorized via molecular function, subcellular
location, and functional categories.