SMYD2 Structure and Function: A Multispecificity ...

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Invi t ing Innova t ions

J Cytol Molecul Biol October 2014 Volume 1, Issue 2 © All rights are reserved by Yang et al.

Review Article

Journal of Group Avens Publishing

SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase

Invi t ing Innova t ions

Cytology & Molecular Biology Margaret Rice1, Yuanyuan Jiang1, Joshua Holcomb1, Laura Trescott1, Nicholas Spellmon1, Nualpun Sirinupong2 and Zhe Yang1* 1

Keywords:

SMYD2; Heart and Muscle development; Structure and

Function

Abstract SMYD2 belongs to a family of SET and MYND domain-containing proteins. SET domain is an evolutionarily conserved domain responsible for protein lysine methylation. MYND domain is a zinc-finger motif that acts as a protein interaction module in corepressor complex recruitment. SMYD2 methylates histones, suggesting a role in epigenetic gene regulation. SMYD2 also methylates several non-histone proteins, demonstrating broad substrate specificity. Through these methylations, SMYD2 regulates heart and muscle development and is associated with various human cancers. This review is to gather the available information on SMYD2 and discuss the relationship between its function and structure.

Introduction SET and MYND domain-containing protein 2 (SMYD2) is a member of the SMYD family of protein methyltransferases. All five members of this family (SMYD1–5) contain a conserved catalytic SET domain and a zinc-finger MYND motif (Figure 1). The SET domain was originally identified in Drosophila Su(var)3-9, Enhancer of zeste, and Trithorax proteins, while MYND was named after Myeloid translocation protein 8, Nervy, and Deaf-1. SMYD2 is abundantly expressed in heart and skeletal muscle [1-5]. Endogenous SMYD2 is expressed in both the nucleus and cytoplasm in cardiomyocytes [4]. SMYD2 plays an important role in heart and muscle development [15]. Knockdown of SMYD2 in zebrafish leads to developmental delays and severe cardiac and skeletal muscle defects [2,3,5]. SMYD2 also has cardiac-protective functions. Knockdown of SMYD2 in mouse cardiomyocytes stimulates cell apoptosis [4]. In addition, SMYD2 plays a critical role in human embryonic stem cell differentiation and acts as a negative regulator of endodermal differentiation [3]. SMYD2 is involved in transcriptional regulation of cellular proliferation and development. SMYD2 regulates gene transcription by methylating histones or interacting with RNA polymerases [6,7].

Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, USA Nutraceuticals and Functional Food Research and Development Center, Prince of Songkla University, Hat-Yai, Songkhla, Thailand

2

*Address for Correspondence Zhe Yang, Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, 540 East Canfield Street, Detroit, Michigan 48201, USA, Tel: 1-313-577-1294; Fax: 1-313-577-2765; Email: [email protected]

Submission: 27 August 2014 Accepted: 01 October 2014 Published: 05 October 2014 Reviewed & Approved by: Dr. Ranjan Ganguly, Department of Biochemistry, Cellular & Molecular Biology. University of Tennessee, Knoxville, USA

SMYD2 is able to methylate numerous histone and non-histone proteins [8-11]. Crystallographic analysis of SMYD2 structures reveals that the broad substrate specificity of SMYD2 is achieved by multiple mechanisms such as distinct peptide binding modes and the intrinsic dynamics of peptide ligands [9]. SMYD2 is overexpressed in various human cancers and is of interest for its potential function as an oncogene [12-14]. Here we review the currently available information concerning the biological function of SMYD2 with an additional focus on its structural and functional relationships. Detailed understanding of SMYD2 structure and function could prove valuable in therapeutic intervention of human cancer.

Tissue and Cell Expression Profiles SMYD2 is preferentially expressed in heart and muscle in different animals. In mice, SMYD2 is expressed in heart, brain, liver, kidney, thymus, and ovary, as well as in the developing mouse embryo [6]. The highest expression occurs in mouse heart and brain and in the neonatal heart during cardiac development. SMYD2 is expressed in embryonic muscles of porcine during foetal muscle development [15]. SMYD2 expression in Xenopus laevis occurs in various muscles and is abundant in skeletal muscle and face region [16]. In zebrafish, SMYD2

Figure 1: Diagram structure of SMYD family proteins. S, S-sequence; MYND (Myeloid, Nervy, Deaf-1); SET-I, insertion SET (Su(Var)3-9, Enhancer-of-zeste, Trithorax) domain; (S)ET, core SET domain; Post-SET, SET C-terminal flanking domain; CTD, carboxy-terminal domain.

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

ISSN: 2325-4653

has two homologous genes: SMYD2a and SMYD2b. The expression of these genes is dependent on the developmental stage of the fish. SMYD2a is maternally expressed, reduced following fertilization, and induced again throughout gastrulation [3]. SMYD2b expression levels are low during early development and then drastically increase following gastrulation [3]. SMYD2a induction during gastrulation suggests that it might play a role in germ layer specification. SMYD2 shows cell-type specific expression in rat heart ventricles. Its expression occurs almost exclusively in rat cardiomyocytes (CM), as non-CM fractions do not show significant expression [1]. Additionally, SMYD2 is preferentially expressed in human somatic cells compared to pluripotent cells [3]. The expression levels of SMYD2 are low in embryonic stem cells but are strongly induced during differentiation [3]. Overexpressed SMYD2 localizes to the cytoplasm and nucleus in 293T cells [6]. Endogenous SMYD2 is expressed in both nuclear and cytosolic cell fractions of CM [1]. In cultured embryonic and neonatal vertebrate CM, SMYD2 shows primarily cytoplasmic localization [1].

Biological Function of SMYD2 SMYD2 plays an important role in heart and muscle development (Table 1). In zebrafish, knockdown of SMYD2 leads to developmental delays and severe cardiac defects [2]. The cardiac defects include malformation of the atria and ventricles, elongation and thinning of hearts, pericardial edema, and inflow tract edema. These defects are associated with reduced heart rate and impaired cardiac performance [2]. SMYD2 knockdown also affects skeletal muscle in zebrafish. Reduced SMYD2 expression causes defects in fish mobility and tail contraction. The knockdown fish shows an abnormal structure in skeletal muscle and disorganized sarcomere in the Z-disk and I-band regions [2]. However, data regarding the role of SMYD2 in cardiac development are currently conflicting. One recent study shows that SMYD2 knockdown in zebrafish has no cardiac phenotype [3]. This result is consistent with the observation that SMYD2 is dispensable for murine heart development. Cardiomyocyte-specific deletion of Table 1: List of SMYD2 functions. Function Heart development Skeletal muscle development Cardiomyocyte protection

Reference [2] [1-3] [4]

Stem cell differentiation

[3]

Cell cycle

[13]

Transcriptional regulation Tumorigenesis Protein–protein interaction

[3,4,6,7] [12-14] [1,2,6,7,18]

Post-translational modification Histones H3

[6,7,11,23]

H4

[11]

H2B

[11]

Non-histones p53

[20]

RB

[13,21]

Hsp90

[2,5]

ERα

[8]

PARP1

[10]

J Cytol Molecul Biol 1(2): 7 (2014)

SMYD2 has no effect on morphological and functional development of the murine heart [1]. This suggests that SMYD2 does not contribute non-redundantly to cardiac maintenance or development even though neonatal cardiomyocytes are the primary site of SMYD2 expression [1]. While the exact function of SMYD2 in cardiac development requires further clarification, additional studies show that SMYD2 has cardiac-protective functions [4]. SMYD2 is required to limit damage to the heart upon myocardial infarction. Knockdown of SMYD2 in cultured cardiomyocytes enhances cell apoptosis induced by the cobalt chloride treatment. SMYD2 overexpression reduces p53 accumulation in cardiomyocytes and partially restores the expression levels of Bcl2. Increased SMYD2 expression is associated with a marked increase in p53 methylation. These data indicate SMYD2 function as a cardioprotective protein by methylation of p53. SMYD2 plays a critical role in human embryonic stem cell (ES) differentiation [3]. Knockdown of SMYD2 in ES cells promotes the induction of endodermal markers during differentiation. The knockdown has no significant effect in the expression of ectodermal or mesodermal genes. SMYD2 overexpression in ES cells causes reduced induction of endodermal and mesodermal genes during differentiation. In contrast, the expression of the ectodermal marker PAX6 is increased in the SMYD2-overexpressing ES cells. This suggests that SMYD2 might act as a negative regulator of endodermal differentiation.

Molecular Mechanisms of SMYD2 Function Transcriptional regulation SMYD2 is involved in transcriptional regulation of cellular proliferation and development. Overexpression of SMYD2 in 293T cells has been associated with up-regulation of 37 genes and downregulation of 4 genes, most of which are involved in important processes such as the cell cycle, transcriptional regulation, and chromatin remodeling [7]. SMYD2 upregulates TACC2 expression by binding to the TACC2 promoter and methylating H3 at lysine 4 [7]. SMYD2 associates with the Sin3A histone deacetylase complex and represses transcription from an SV40-luciferase reporter [6]. In murine SMYD2-knockout heart, the majority of deregulated genes show reduced expression and are functionally associated with translation [1]. This indicates that SMYD2 might act as a transcriptional activator in the wild type murine heart. SMYD2a knockdown in zebrafish affects the expression of genes involved in the Nodal pathway, which is involved in gastrulation [3]. SMYD2a knockdown causes an induction of Nodal-target genes chd, gsc, bon, and cas during gastrulation events. bmp2a and β-catenin-1 are unaffected by SMYD2a knockdown. Increased expression of the Nodal-target genes in zebrafish promotes endoderm formation [17]. SMYD2 knockdown in zebrafish is also associated with low expression of ntl [3]. This is consistent with the role of SMYD2 in fish development and tail formation. Cardiac knockout of SMYD2 in vivo is associated with elevated expression of p53 and pro-apoptotic Bax, supporting its cardiac-protective role in mice [4].

Protein interaction partners Proteomic analysis shows that SMYD2 has 28 protein interactors in 293T cells [18]. SMYD2 associates with proteins involved in cellPage - 02

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

ISSN: 2325-4653

cycle regulation, DNA damage response, and chaperone machinery [7,18]. The association of SMYD2 with Hsp90 changes SMYD2 methylation preference from H3K36 to H3K4 and enhances its activity in vitro [7]. SMYD2 interacts with RNA polymerase II and RNA helicase HELZ [1]. This suggests that SMYD2 might facilitate target gene expression via the elongation of transcription. In mouse cardiomyocytes, SMYD2 associates with the sarcomeric I-band region at the titin N2A-domain [4]. Binding to N2A occurs via N-terminal and extreme C-terminal regions of SMYD2. SMYD2 also binds the protein titin in zebrafish and co-localizes with Hsp90 on myofibrils [2]. Similar interactions were observed in chick skeletal and cardiac myocytes, human heart myofibrils, and adult human diaphragm muscle [2]. SMYD2 associates with the Sin3A histone deacetylase complex and represses transcription [6]. SMYD2 interacts with DAL-1 via the MYND domain, which might link SMYD2 to the pathogenesis of meningioma [7,19].

Methylation targets SMYD2 is known to methylate histone-3 lysine-36 [6,7] as well as histone-3 lysine-4 [7], suggesting its role as a transcriptional activator in epigenetic gene regulation [6,7]. SMYD2 also methylates several non-histone proteins, demonstrating broad substrate specificity. These non-histone methylation targets include p53, retinoblastoma tumor suppressor (RB), cytoplasmic Hsp90, estrogen receptor α (ERα), and most recently Poly(ADP-ribose) polymerase-1 (PARP1) [2,5,8,10,13,18,20-22]. p53 is methylated by SMYD2 at lysine 370, attenuating p53-mediated transactivation activity [20]. SMYD2 mono-methylates RB at lysine 810 and 860, regulating RB activity during the cell cycle, differentiation, and in response to DNA damage [13,21]. SMYD2-mediated methylation of Hsp90 at lysine 615 regulates myofilament organization through the formation of a complex with the sarcomeric protein titin [2,5]. ERα methylation by SMYD2 at lysine 266 decreases ERα chromatin recruitment and inhibits ERα target gene activation under estrogen-depleted conditions [8]. SMYD2 mono-methylates PARP1 at lysine 528 and regulates poly(ADP-ribosyl)ation activity in response to oxidative DNA damage [10]. These findings suggest potential function of SMYD2 as an oncogene due to demonstrated methylation of tumor suppressors and inhibition of tumor suppressive functions.

Biochemical and Biophysical Characterization SMYD2 protein catalyzes lysine methylation using S-adenosylmethionine (AdoMet) as a methyl donor. Transfer of the methyl group to a target molecule converts AdoMet to S-adenosylhomocysteine (AdoHcy). Steady-state kinetic studies have shown that SMYD2 utilizes a rapid-equilibrium random Bi-Bi mechanism in catalysis [11]. Methylation of H3 or p53 is maximal at alkaline pH (pH 9.0–10.0) [11], while Hsp90 methylation peaks between pH 7.5 and 8.0 [18]. The effect of pH on SMYD2 catalytic efficiency is mainly a kcat-driven effect. The KMs of both AdoMet and the substrate remain the same at different pH conditions [11]. This suggests that solvent basicity may contribute to the deprotonation of the ε-amine group in target lysine. SMYD2 activity is sensitive to ionic strength, with optimal activity observed at low ionic strength. SMYD2 has maximal activity at ~32 °C, with activity rapidly dropping at temperatures above 37 °C [11]. The catalytic efficiency of SMYD2 differs significantly between J Cytol Molecul Biol 1(2): 7 (2014)

its substrates. SMYD2 shows more than 10-fold higher activity to p53 peptide than histone peptide substrates [23]. The activity of SMYD2 on p53 protein was approximately 3- to 6-fold higher than histone H3 or nucleosome substrates [23]. H4 and H2B are more efficient substrates with 3- to 5-fold higher activity compared to H3 [11]. Full-length H3 is an efficient substrate for SMYD2 compared to H3 peptides [11,23]. SMYD2 shows very weak activity to H3K36 peptides but possesses observable activity on H3K4 [7]. Differences in SMYD2 activity are correlated to substrate binding. ITC analysis shows that the binding of SMYD2 to p53 peptides (residues 361–380) has a dissociation constant (Kd) of ~20 µM, while binding to H3K4 (residues 1–20) peptides is about 35-fold weaker [23]. These data suggest that SMYD2 preferentially binds and methylates p53 in vitro. Hsp90 enhances SMYD2 methyltransferase activity [7]. This activity enhancement occurs on the substrate of H3K4 but not H3K36 [7]. Hsp90 also enhances H3K4 methylation activity of SMYD1 and SMYD3 [24,25]. However, the precise mechanism of the Hsp90induced activity enhancement is unknown. The activity of SMYD2 on Hsp90 is affected by some Hsp90 co-chaperones. AHA1 and p50 have no significant effect on Hsp90 methylation, but the presence of Hop causes a drastic decrease in methylation [18]. Hsp90 binds Hop via the C-terminal MEEVD motif. This region has been suggested to bind to the CTD domain of SMYD2 [9,26].

Structure and Function Relationships Overall SMYD2 structure Like all SMYD proteins, SMYD2 structure is bilobal, containing a N-terminal lobe and C-terminal lobe connected by a nonconserved sequence (Figure 2) [9,23,26-28]. The N-terminal-lobe is comprised of several domains, including the SET, SET-I, post-SET, and MYND domains, and the C-terminal-lobe is formed by the carboxy-terminal domain (CTD). SMYD2 has been co-crystallized with substrates ERα and p53 [9,23,28]. These complex structures show that the substratebinding site is located at the cleft between the N-terminal and C-terminal lobes, both of which are involved in substrate recognition (Figure 2A). In SMYD proteins, cofactor-binding modes are structurally conserved. The SET, SET-I, and post-SET domains form a surface pocket where L-shaped cofactors bind [9]. Lysine access channels of SMYD proteins have large ovular openings, a structural difference from other proteins with SET domains [9,26,29,30]. The complex structures also show that the target lysine inserts its side chain into the lysine access channel to reach the cofactor bound on the opposite side of the SET domain [9].

Catalytic SET domain SMYD2 has a “split” SET domain similar to other SMYD proteins [9,26,29,30]. The SET domain is split because the MYND domain separates it into two parts: the S-sequence and the core SET domain (Figure 2A). These two parts associate with each other in structure, creating a fold that is evolutionarily conserved in SET proteins. The methyltransferase activity of SMYD2 is dependent on the SET domain. Mutation of residues from the cofactor binding site, lysine access channel, or substrate binding site impairs the enzyme activity (Figure 2B). In structure, the SET domain is surrounded by the SET-I and post-SET domains. These two domains also contribute to the binding of cofactors and substrates [9,23,26,28]. The SET-I is a helix bundle Page - 03

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

ISSN: 2325-4653

B

A

CTD Substrate Binding

PXLXP Binding Site

CTD

E189K, E190K: reduce activity E187K: significantly

∆CTD significantly increases activity to histone substrate H3K4, but decreases activity to p53 Y374F&A: significantly decreases activity

decreases activity Y245F&A: significantly decreases activity D252R, R253Q: modest

MYND

effect on methylation

SET-I

AdoMet Bind ing

Post-SET

NHSC: sharp reduction of activity G18/20A: significantly decreases activity GEE: sharp reduction of activity

Target Lys Binding Y240F: catalytically inactive C264A: significantly reduces activity

Figure 2: SMYD2 crystal structure. The structure is shown in surface representation (A) and ribbon diagram (B). The S-sequence, MYND, SET-I, core SET, postSET, and CTD are depicted in light green, blue, pink, green, cyan, and red. AdoHcy (SAH) is represented by balls-and-sticks. Zinc ions are denoted by purple spheres. ERα and p53 peptides are shown in yellow and light blue in A. The locations of point mutations, deletions, and truncation in SMYD2 are indicated in the structure in B.

inserted in the SET domain. The post-SET, immediately downstream of the SET domain, is a small cysteine-rich domain folded with one zinc atom. Deletion of the post-SET domain abolishes SMYD2 methyltransferase activity [11]. Mutation of a zinc-chelating residue (C264S) within the post-SET domain results in a sharp reduction in SMYD2-mediated methylation [11]. This indicates that an intact post-SET domain is required for SMYD2 methyltransferase activity.

Zinc-finger MYND domain The MYND domain is a zinc-finger motif composed of two zinc atoms (Figure 2A). The MYND domain forms direct contact with the SET domain, but does not contribute residues to binding of cofactors and substrates. This is consistent with the findings that the MYND domain is dispensable for the histone methylation activity of SMYD2 [7]. MYND domains facilitate specific protein interaction by binding to a sequence rich in proline [31]. SMYD2 interacts with EBP41L3 via the MYND domain. EBP41L3 is a tumor suppressor containing a PXLXP motif [7]. The predicted EBP41L3 binding site is located in a shallow groove on the surface of the MYND domain [26]. The groove is fully exposed and readily accessible by potential binding proteins (Figure 2A). This suggests that the MYND domain may function primarily as a protein–protein interaction module, coordinating SMYD2 with other proteins to regulate tumor proliferation.

TPR-like Carboxy-terminal domain The CTD domain is comprised of seven antiparallel helices, forming a helix-turn-helix structure. This resembles the structure of tetratricopeptide repeats (TPR), though they lack significant similarities in sequence. This similarity in structure indicates that the CTD might function as a protein–protein interaction module, since TPR motifs mediate the assembly of multi-protein complexes [32]. Current evidence indicates that the CTD is indeed important for protein interaction. The CTD is responsible for interaction between J Cytol Molecul Biol 1(2): 7 (2014)

SMYD3 and HTLV-1 Tax in primary T cells [33]. Binding to the sarcomeric protein titin by SMYD2 is mediated by N-terminal and extreme C-terminal regions of SMYD2 [4]. In a SMYD2 structure, the CTD binds a PEG molecule, suggesting that an additional peptidebinding site might be present in the CTD, potentially allowing the binding of two different proteins [9]. The CTD is well conserved in the SMYD family of proteins, with the exception of SMYD5 [18]. However, the structural orientation of the CTD differs between SMYD proteins. SMYD1 has an “open” CTD conformation with the active site completely exposed in the structure [29]. The conformation of the CTD in SMYD3 is “closed”, resulting in a deep and narrow substrate-binding cleft [30]. SMYD2 appears to be a conformational intermediate between SMYD1 and SMYD3 [9,26]. In addition, the CTD in SMYD2 is able to adopt different conformations depending on the cofactor analogues binding to the protein [26]. These observations suggest the intra- and interdomain flexibility of CTD domains. The CTD in SMYD2 is vital to the creation of the substratebinding pocket, and has been found to stabilize interactions with p53 [23,28]. However, the effect of CTD on SMYD2 activity appears to be substrate-dependent. Deletion of the CTD dramatically reduced the methylation activity of SMYD2 on p53 proteins, while histone H3 methylation was unimpaired [23]. When peptides were used as substrates, CTD deletion showed increased activity on H3K4 but did not affect H3K36 or p53 methylation [23]. However, substitution of Tyr374 with alanine resulted in loss of p53 peptide methylation [23]. Tyr374 is located in the CTD and is involved in substrate binding (Figure 2B). This suggests that the CTD may play a role in substrate recognition, though further clarification is still needed.

Molecular basis of broad substrate specificity Comparison of the SMYD2–ERα and SMYD2–p53 structures has Page - 04

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

ISSN: 2325-4653

Ascending U-arm

Descending U-arm

R+3

R+5

Q+5

B

G+4

H-2

K+3

M-2 H+1/S+1

K+2/K+2

A

C

L-1/L-1

U-base KO/KO

Figure 3: Structural comparison of SMYD2­–ERα and SMYD2–p53. (A) Superposition of the structures of the ERα peptide (green) and p53 peptide (light blue; PDB code: 3TG5). Position 0 refers to the target lysine. (B) and (C) Structural and binding differences of ER and p53 at position +3 and +5. SMYD2 is colored according to domains while the ERα and p53 peptides are shown in green and light blue.

provided insight into the broad substrate specificity of SMYD2 [9]. ERα and p53 peptides bind SMYD2 in similar U-shaped conformations in structure (Figure 2A). Significant structural differences occur at residues C-terminal to the target lysine (Figure 3A). For example, in SMYD2–ERα Arg+3 (R269) binds in the β8–β9 region of the SET domain, whereas in SMYD2–p53 the side chain of Lys+3 (K273) is stabilized by interaction with Tyr370 and Tyr374 from the CTD and Asp242 from the loop preceding the post-SET domain (Figure 3B). Similarly, Arg+5 (R271) in SMYD2–ERα interacts with the β8–β9 hairpin, whereas Gln+5 (Q375) in SMYD2–p53 inserts its side chain into a deep pocket formed by His341, Tyr344, Gln345, and Tyr370 from the CTD and Leu244 and Tyr245 from the post-SET-preceding loop (Figure 3C). Analysis of these differences has suggested that the SMYD2 targeting diversity is achieved by multiple mechanisms including various distinct binding sites, conformational plasticity of the substrate-binding pocket, intrinsic dynamics of peptide ligands, and substrate-specific intrapeptide interaction [9]. The interplay of these mechanisms would create sets of complex-specific states that may underlie the ability of SMYD2 to methylate various substrates [9]. Therefore, structure determination of additional SMYD2–substrate complexes will allow further identification of diversity determinants responsible for substrate discrimination.

Disease Relevance SMYD2 is overexpressed in various human cancers and is of interest for its potential function as an oncogene. Overexpression of SMYD2 is associated with tumorigenesis in esophageal squamous cell carcinoma (ESCC) [12]. SMYD2 overexpressing tumors are correlated to worse survival rate of ESCC patients in comparison to those with non-SMYD2 expressing tumors. SMYD2 knockdown inhibits growth of ESCC cells, while overexpression promotes cell proliferation [12]. SMYD2 expression is also altered in bone marrow samples in patients with pediatric acute lymphoblastic leukemia (ALL) [14]. High expression of SMYD2 in ALL patients is correlated J Cytol Molecul Biol 1(2): 7 (2014)

with poor prognosis. In patients that respond to chemotherapy, SMYD2 expression levels decrease significantly with treatment [14]. In bladder carcinoma, expression levels of SMYD2 are significantly higher in comparison to non-neoplastic bladder tissues [13]. The expression levels of SMYD2 are significantly higher in tumor tissues than that of any normal tissue. Knockdown of SMYD2 leads to suppression of cancer cell growth. Analysis of the cell cycle suggests a potential role of SMYD2 in the transition from G1 to S phase [13]. Due to its close association with various human cancers, SMYD2 is of interest for its potential as a therapeutic target in human cancer. The clear role of SMYD2 in tumorigenesis has not yet been identified, but the ability of SMYD2 to methylate proteins involved in cancer progression provide potential mechanisms for its putative tumorigenic function. Repression of p53 activity by SMYD2mediated methylation suggests that SMYD2 may function as an oncogene by repressing p53 tumor suppressive function and interfering with apoptotic pathways [20]. The identification of RB as a target of SMYD2 indicates that SMYD2 may be involved in malignant cell cycle regulation during tumorigenesis [21]. RB is a central cell cycle regulator and tumor suppressor, and mutation of this protein has been found in a large spectrum of human cancers [34,35]. Although the consequence of Hsp90 methylation by SMYD2 has not yet been determined, any effect on Hsp90 chaperone activity would link SMYD2 to Hsp90-mediated signaling transduction, angiogenesis, and apoptosis [2,36,37]. Hsp90 is overexpressed in many cancers and acts as a protector to stabilize mutant proteins such as v-Src, the fusion oncogene Bcr/Abl, and mutant forms of p53 during cell transformation [38]. In addition, SMYD2 methylating PARP1 suggests that SMYD2 may function in tumorigenesis through regulating PARP1-mediated DNA repair and maintaining genomic stability of cancerous cells [10,39,40]. While SMYD2 has not yet been associated with breast cancer, ERα methylation by SMYD2 implicates its potential role in breast cancer development. SMYD2 Page - 05

Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

ISSN: 2325-4653

inhibits ERα function in an estrogen-depleted manner [8]. This suggests that SMYD2 might be actually associated with the highly malignant ERα-negative breast cancer. The exact role of SMYD2 in tumorigenesis requires additional investigation. Studying the mechanism controlling the broad substrate specificity of SMYD2 could prove valuable in selective drug design and enables targeting of specific pathways driving tumorigenesis. References 1. Diehl F, Brown MA, van Amerongen MJ, Novoyatleva T, Wietelmann A, et al. (2010) Cardiac deletion of Smyd2 is dispensable for mouse heart development. PLoS One 5: e9748. 2. Donlin LT, Andresen C, Just S, Rudensky E, Pappas CT, et al. (2012) Smyd2 controls cytoplasmic lysine methylation of Hsp90 and myofilament organization. Genes Dev 26: 114-119. 3. Sese B, Barrero MJ, Fabregat MC, Sander V, Izpisua Belmonte JC (2013) SMYD2 is induced during cell differentiation and participates in early development. Int J Dev Biol 57: 357-364. 4. Sajjad A, Novoyatleva T, Vergarajauregui S, Troidl C, Schermuly RT, et al. (2014) Lysine methyltransferase Smyd2 suppresses p53-dependent cardiomyocyte apoptosis. Biochim Biophys Acta 1843: 2556-2562. 5. Voelkel T, Andresen C, Unger A, Just S, Rottbauer W, et al. (2013) Lysine methyltransferase Smyd2 regulates Hsp90-mediated protection of the sarcomeric titin springs and cardiac function. Biochim Biophys Acta 1833: 812-822. 6. Brown MA, Sims RJ 3rd, Gottlieb PD, Tucker PW (2006) Identification and characterization of Smyd2: a split SET/MYND domain-containing histone H3 lysine 36-specific methyltransferase that interacts with the Sin3 histone deacetylase complex. Mol Cancer 5: 26.

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Citation: Rice M, Jiang Y, Holcomb J, Trescott L, Spellmon N, et al. SMYD2 Structure and Function: A Multispecificity Protein Lysine Methyltransferase. J Cytol Molecul Biol. 2014;1(2): 7.

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Acknowledgements This study was supported by the Aplastic Anemia & MDS International Foundation and Leukemia Research Foundation (to ZY).

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