Expression Pattern, Regulation, and Functions of Methionine Adenosyltransferase 2β Splicing Variants in Hepatoma Cells

Yang, Heping; Iglesias–Ara, Ainhoa; Magilnick, Nathaniel; Xia, Meng; Ramani, Komal; Chen, Hui; Lee, Taunia D.; Mato, José M.; Lu, Shelly C.

doi:10.1053/j.gastro.2007.10.027

Cited by 65 publications

(108 citation statements)

References 24 publications

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“…The magnitude of increase in MAT2␤ protein level is ϳ150-fold in V1-or V2-overexpressing HuH-7 cells, because the baseline expression is absent. Likewise, MAT2␤ is not expressed in normal liver (2,4), and the level of MAT2␤ protein in HCC specimens is comparable to HuH-7 cells overexpressing V1 or V2. There is a faint MAT2␤ band seen in NL1, which may be due to the fact that MAT2␤ is induced in cirrhosis, which is almost always present in the setting of HCC (2).…”

Section: Resultsmentioning

confidence: 94%

“…V1 exerts a stronger effect than V2 on cyclin expression and growth and may be related to the fact that it also exerts a stronger effect in modulating cytoplasmic HuR content. Interestingly, although both variants increase growth, only V1 regulates apoptosis and c-Jun N-terminal kinase signaling (4). There are also differences in the tissue distribution of these two variants, with some tissues expressing predominantly V1, whereas others express V2 (4).…”

Section: Resultsmentioning

confidence: 99%

“…Cell Transfection and Gene Expression Analysis-The expression plasmids pcDNA3.1D/V5-His/MAT2␤ V1 (V1 expression vector) or pcDNA3.1D/V5-His/MAT2␤ V2 (V2 expression vector) have been described previously (4). To overexpress MAT2␤ variant proteins in HuH-7 cells, 90% confluent HuH-7 cells were transiently transfected with the V1 or V2 plasmids using Lipofectamine 2000 (Invitrogen) following the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

“…V1 encodes a 334-amino acid protein beginning MVGREKELSIHFVPGSCRLVE…. The alternatively spliced V2 utilizes a different first exon lying further upstream in the genomic sequence to encode a hypothetical 323-amino acid isoform beginning MPEMPEDMEQ… (4). The reading frame for both variants converge after this point and are identical.…”

Section: Methionine Adenosyltransferase (Mat)mentioning

confidence: 99%

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Novel Function and Intracellular Localization of Methionine Adenosyltransferase 2β Splicing Variants

Xia

Chen

Wang

et al. 2010

Journal of Biological Chemistry

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Human methionine adenosyltransferase 2␤ (MAT2␤) encodes for two major splicing variants, V1 and V2, which are differentially expressed in normal tissues. Both variants are induced in human liver cancer and positively regulate growth. The aim of this work was to identify interacting proteins of V1 and V2. His-tagged V1 and V2 were overexpressed in Rosetta pLysS cells, purified, and used in a pulldown assay to identify interacting proteins from human colon cancer cell line RKO cell lysates. The eluted lysates were subjected to Western blot and in solution proteomic analyses. HuR, an mRNA-binding protein known to stabilize the mRNA of several cyclins, was identified to interact with V1 and V2. Immunoprecipitation and Western blotting confirmed their interaction in both liver and colon cancer cells. These variant proteins are located in both nucleus and cytoplasm in liver and colon cancer cells and, when overexpressed, increased the cytoplasmic HuR content. This led to increased expression of cyclin D1 and cyclin A, known targets of HuR. When endogenous expression of V1 or V2 is reduced by small interference RNA, cytoplasmic HuR content fell and the expression of these HuR target genes also decreased. Knockdown of cyclin D1 or cyclin A blunted, whereas knockdown of HuR largely prevented, the ability of V1 or V2 overexpression to induce growth. In conclusion, MAT2␤ variants reside mostly in the nucleus and regulate HuR subcellular content to affect cell proliferation. Methionine adenosyltransferase (MAT)2 is essential to life, because it is the only enzyme that catalyzes the formation of S-adenosylmethionine, the principal biological methyl donor (1). In mammals, two different genes, MAT1A and MAT2A, encode for two homologous MAT catalytic subunits, ␣1 and ␣2. MAT1A is expressed mostly in the liver while MAT2A is widely distributed. In adult liver, increased expression of MAT2A is associated with rapid growth or de-differentiation. Up until recently, the MAT2␤ gene was thought to encode for the regulatory subunit (␤) that is associated only with MAT2A-encoded enzyme (MATII) to lower its K m and K i for methionine and S-adenosylmethionine, respectively. MAT2␤ expression is induced in cirrhosis and hepatocellular carcinoma (HCC). Importantly, increased MAT2A and MAT2␤ expression offer liver cancer cells a growth advantage (2, 3).In a recent publication (4), we described novel functions of MAT2␤ that greatly increased its importance in biology. To study transcriptional regulation of MAT2␤, we cloned and characterized its 5Ј-flanking region and uncovered multiple alternate splicing variants and termed the two major variants V1 and V2. V1 encodes a 334-amino acid protein beginning MVGREKELSIHFVPGSCRLVE…. The alternatively spliced V2 utilizes a different first exon lying further upstream in the genomic sequence to encode a hypothetical 323-amino acid isoform beginning MPEMPEDMEQ… (4). The reading frame for both variants converge after this point and are identical. We examined their expression pattern in human tissues and H...

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Section: Resultsmentioning

confidence: 94%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methionine Adenosyltransferase (Mat)mentioning

confidence: 99%

See 2 more Smart Citations

Novel Function and Intracellular Localization of Methionine Adenosyltransferase 2β Splicing Variants

Xia

Chen

Wang

et al. 2010

Journal of Biological Chemistry

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“…MATα and MATβ subunits can form several MATαβ complexes (3,4). There are two major splicing forms of the MAT2B gene that encode for MATβV1 and MATβV2, which share very high sequence similarity and differ by only 20 residues in the N terminus (5). We recently reported that MATβ isoforms interact through their C terminus with MATα2 to form the MATαβ complexes MATαβV1 and MATαβV2 (4).…”

mentioning

confidence: 99%

Crystallography captures catalytic steps in human methionine adenosyltransferase enzymes

Murray

Antonyuk

Marina

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The principal methyl donor of the cell, S-adenosylmethionine (SAMe), is produced by the highly conserved family of methionine adenosyltranferases (MATs) via an ATP-driven process. These enzymes play an important role in the preservation of life, and their dysregulation has been tightly linked to liver and colon cancers. We present crystal structures of human MATα2 containing various bound ligands, providing a "structural movie" of the catalytic steps. High-to atomic-resolution structures reveal the structural elements of the enzyme involved in utilization of the substrates methionine and adenosine and in formation of the product SAMe. MAT enzymes are also able to produce S-adenosylethionine (SAE) from substrate ethionine. Ethionine, an S-ethyl analog of the amino acid methionine, is known to induce steatosis and pancreatitis. We show that SAE occupies the active site in a manner similar to SAMe, confirming that ethionine also uses the same catalytic site to form the product SAE.methionine adenosyltransferase | cell growth | liver cancer | X-ray crystallography | methylation

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Switch from Mnt‐Max to Myc‐Max induces p53 and cyclin D1 expression and apoptosis during cholestasis in mouse and human hepatocytes†

Yang

et al. 2009

Hepatology

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Background and rationale Toxic bile acids induce hepatocyte apoptosis for which p53 and cyclin D1 have been implicated as underlying mediators. Both p53 and cyclin D1 are targets of c-Myc, which is also up-regulated in cholestasis. Myc and Mnt use Max as a cofactor for DNA binding. Myc-Max typically activates transcription via E-box binding. Mnt-Max also binds E-box sequence but serves as a repressor and inhibits the enhancer activity of Myc-Max. The current work tested the hypothesis that the switch from Mnt-Max to Myc-Max is responsible for p53 and cyclin D1 up-regulation and apoptosis during cholestasis. Results Following common bile duct ligation or left hepatic bile duct ligation, the expression of p53, c-Myc and cyclin D1 increased markedly while Mnt expression decreased. Nuclear binding activity of Myc to E-box element of p53 and cyclin D1 increased, while that of Mnt decreased in a time-dependent fashion. Lithocolic acid (LCA) treatment of primary human hepatocytes and HuH-7 cells induced a similar switch from Mnt to Myc, increased p53 and cyclin D1 promoter activity, endogenous p53 and cyclin D1 expression and apoptosis. Blocking c-Myc induction in HuH-7 cells prevented the LCA-mediated increase in p53 and cyclin D1 expression and reduced apoptosis. Lowering Mnt expression further enhanced LCA’s inductive effect on p53 and cyclin D1. Bile duct ligated mice treated with lentivirus harboring c-myc siRNA were protected from hepatic induction of p53 and cyclin D1, switch in Mnt to Myc nuclear binding to E-box, and hepatocyte apoptosis. Conclusions The switch from Mnt to Myc during BDL and in hepatocytes treated with LCA is responsible for the induction in p53 and cyclin D1 expression and contributes to apoptosis.

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Expression Pattern, Regulation, and Functions of Methionine Adenosyltransferase 2β Splicing Variants in Hepatoma Cells

Cited by 65 publications

References 24 publications

Novel Function and Intracellular Localization of Methionine Adenosyltransferase 2β Splicing Variants

Novel Function and Intracellular Localization of Methionine Adenosyltransferase 2β Splicing Variants

Crystallography captures catalytic steps in human methionine adenosyltransferase enzymes

Switch from Mnt‐Max to Myc‐Max induces p53 and cyclin D1 expression and apoptosis during cholestasis in mouse and human hepatocytes†

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