Editorial :
Lysine Methylation of Non-Histone Proteins
Byron Baron
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Introduction
Proteomics, or the large-scale study of protein structure and
function, has contributed greatly to our understanding of cellular biology and
disease. Over time, it has become apparent that the proteome is spatially,
temporally, and chemically dynamic allowing for the same protein to perform
very different functions and fufill completely unrelated roles in a cell
through small chemical changes. This can be described as epiproteomics (just as
epigenetic are changes to DNA not encoded in the DNA sequence) and mainly
covers Post-Translational Modifications (PTMs) such as phosphorylation,
ubiquitination, acetylation or methylation, among over 200 or more [1]. These
vary depending on cell type, signalling, stress condition, micro-environment and
so on, leading to some sort of change in protein properties, which
consequently, either directly or indirectly, alters the function.
One of the PTMs that is gaining new interest for various reasons
is methylation onlysine in non-histone proteins. Lysine can undergo mono
(Kme1), di (Kme2) or tri (Kme3) methylation on its epsilon amine and these
reactions are catalysed by proteinlysine methyltransferases (PKMTs). Initially,
lysine methylation was extensively studied on various histone residues, playing
important roles in the regulation (both activation and repression) of chromatin
packing and gene transcription [2].
Subsequently, it has been found to play many important roles in
non-histone proteins (mainly transcription factors and chaperones) by impacting
function via domain activity, interaction strength to target proteins or DNA,
localisation and protein stability or half-life. Moreover, as lysine can
undergo various alterations, methylation can compete with other PTMs, adding another level of regulation
[3]. A good example to illustrate this point is the tumour suppressor p53, in
which methylation of different lysine residues or different degrees of
methylation (mono or di) leads to alteration of very different properties from
suppresion of gene transcription to increased affinity for 53BP1[3].
More recently, two very
interesting phenomena have been identified. The first is the report of Heat
Shock Protein 70 (HSP70) acting as a transcription factor following lysine
methylation, translocation to the nucleus and enhanced the kinase activity of Aurora
kinase B (AURKB) [4] and the second is the isolation of a group of PKMTs that do
not act on histones but appear to act mainly on specific categories of
nonhistone proteins such as transcription factors and chaperones [5].
As expected, dysregulation of PKMTs leads to disease, for example,
SUV39H1 [6] and EZH2 [7] have been shown to be over-expressed in a variety of
human tumors and a gain-of-function mutation in EZH2 has been shown to increase
catalytic activity and lead to tumorigenesis [8]. Interestingly, truncation
mutations in METTL23 have
been linked to Intellectual Disability (ID) [9,10]. This implies
that such enzymes make potential therapeutic targets, and there is an ongoing
search for PRMT-inhibiting small molecules [11].
HSPs have also been linked to various insulin signalling pathways
and Type 2 diabetes mellitus (T2DM), such that HSP70 [12], HSP90 and 78-kDa
glucoseregulated protein (GRP78) [13] were significantly up-regulated in the
skeletal muscles of T2DM patients. We have therefore developed a custon
sandwich ELISA for investigating the presence and quantification of lysine
methylation on HSP27, HSP60 (mitochondrial), HSP70, GRP78 and HSP90 in the
serum of diabetics and matched non-diabetic controls and are currenly going
through our first round of 100 patients.
However, despite recognising the importance of lysine methylation
in both cellular biology and human disease, much is still unknown regarding
target proteins, residue position, degree, enzyme and function. The main
reasons for this are a lack of reliable tools and methods for studying these modifications.
Top-down proteomic methods allow the sequencing of proteins and the
identification of PTMs but the resolution of the mass spectrometry system used
needs to be very high. So far the best studied PTMs by such methods is
phosphorylation and the reason for this is that while the addition of a
phosphate group to a protein increases the mass by 80Da (and gives an overall
negative charge) or an acetyl group adds 42Da (and neutralizes the positive charge
on the lysine residue), the addition of a methyl group adds only 14Da (and does
not change the charge). This makes the mass on charge (m/z) change following
methylation, extremely small. Another hurdle is the limited availability and
low quality of commercial antibodies to detect lysine methylation.
The good news is that these are only short-term inconveniences. The
technological advances in mass spectrometry are extremely fast both in terms of
improved resolution, especially using electron transfer dissociation (ETD), and
in the development of enrichment techniques which improve the stoichiometric presence
of modified species. Similarly, antibodies are constantly being developed and
tested. As interest in the subject picks up,
more and more robust and reliable anti-methyl lysine antibodies will
become available as has happened with anti-phospho and antiacetyl antibodies.
In time, lysine methylation will become part of standard biomarker assays and
diagnostic kits as has already happened with phosphorylation.
References:
1. Krishna RG, Wold F. Post-translational modification of protein
(1993) Adv.Enzymol. Relat. Areas Mol. Biol. 67: 265-298.
2. Aletta JM, Cimato TR, Ettinger MJ. Protein methylation: a
signal event inpost-translational modification (1998) Trends Biochem Sci 23:
89-91.
3. Baron B. The lysine multi-switch: the impact of lysine
methylation ontranscription factor properties (2014) Biohelikon: Cell Biology
2: a13.
4. Cho HS, Shimazu T, Toyokawa G, Daigo Y, Maehara Y, et al.
EnhancedHSP70 lysine methylation promotes proliferation of cancer cells through activation of Aurora kinase (2012) B. Nat. Commun. 3 :1072.
5. Cloutier P, Lavallée-Adam M, Faubert D, Blanchette M, Coulombe
B.Methylation of the DNA/RNA-binding protein Kin17 by METTL22 affects its association with chromatin (2014) Journal of Proteomics 100:
115-124.
6. Kang MY, Lee BB, Kim YH, Chang DK, Kyu Park S. Association of
the SUV39H1 histone methyltransferase with the DNA methyltransferase 1 at mRNA expression level in primary colorectal cancer (2007) Int J
Cancer, 121: 2192-2197.
7. Tsang DP, Cheng AS. Epigenetic regulation of signaling pathways
in cancer: role of the histone methyltransferase EZH2 (2011) J Gastroenterol Hepatol 26: 19-27.
8. Yap DB, Chu J, Berg T, Schapira M, Cheng SW, et al. Somatic
mutations at EZH2 Y641 act dominantly through a mechanism of selectively
altered PRC2 catalytic activity, to increase H3K27 trimethylation (2011)
Blood 117: 2451-2459.
9. Reiff RE, Ali BR, Baron B, Yu TW, Ben-Salem S, et al. METTL23,
a transcriptional partner of GABPA, is essential for human cognition (2014) Hum
Mol Genet 23: 3456-3466.
10. Bernkopf M, Webersinke G, Tongsook C, Koyani CN, Rafiq MA, et
al. Disruption of the methyltransferase-like 23 gene METTL23 causes mild autosomal recessive intellectual disability (2014) Hum Mol Genet
23: 4015-4023.
11. Yost JM, Korboukh I, Liu F, Gao C, Jin J. Targets in
epigenetics: inhibiting the methyl writers of the histone code (2011) Curr Chem
Genomics 5: 72-84.
12. Sreekumar R, Halvatsiotis P, Schminke JC, Nair KS. Gene
Expression Profile in Skeletal Muscle of Type 2 Diabetes and the Effect of
Insulin Treatment (2002) Diabetes 51:1913-1920.
13. Hojlund K, Wrzesinski K, Mose Larsen P, Fey SJ, Roepstorff P,
et al. Proteome analysis reveals phosphorylation of ATP synthase beta -subunit in human skeletal muscle and proteins with potential roles in type
2 diabetes (2003) J Biol Chem 20: 479-487
Keywords
Phosphorylation, Methylation, Transcription, Histone proteins, Tumor supressors
