DUT (gene)

Last updated

DUT
Protein DUT PDB 1q5h.png
Available structures
PDB Ortholog search: PDBe RCSB
Identifiers
Aliases DUT , dUTPase, deoxyuridine triphosphatase
External IDs OMIM: 601266; MGI: 1346051; HomoloGene: 31475; GeneCards: DUT; OMA:DUT - orthologs
Orthologs
SpeciesHumanMouse
Entrez

1854

110074

Ensembl

ENSG00000128951

ENSMUSG00000027203

UniProt

P33316

Q9CQ43

RefSeq (mRNA)

NM_001025248
NM_001025249
NM_001948
NM_001330286

NM_001159646
NM_023595

RefSeq (protein)

NP_001020419
NP_001020420
NP_001317215
NP_001939

NP_001153118
NP_076084

Location (UCSC) Chr 15: 48.33 – 48.34 Mb Chr 2: 125.09 – 125.1 Mb
PubMed search [3] [4]
Wikidata
View/Edit Human View/Edit Mouse

DUTP pyrophosphatase, also known as DUT, is an enzyme which in humans is encoded by the DUT gene on chromosome 15. [5]

Contents

This gene encodes an essential enzyme of nucleotide metabolism. The encoded protein forms a ubiquitous, homotrimeric enzyme that hydrolyzes dUTP to dUMP and pyrophosphate. This reaction serves two cellular purposes: providing a precursor (dUMP) for the synthesis of thymine nucleotides needed for DNA replication, and limiting intracellular pools of dUTP. Elevated levels of dUTP lead to increased incorporation of uracil into DNA, which induces extensive excision repair mediated by uracil glycosylase. This repair process, resulting in the removal and reincorporation of dUTP, is self-defeating and leads to DNA fragmentation and cell death. Alternative splicing of this gene leads to different isoforms that localize to either the mitochondrion or nucleus. A related pseudogene is located on chromosome 19. [5]

Structure

In humans, this gene encodes a homotrimeric enzyme with two isoforms characterized by their distinct subcellular localizations: the nuclear isoform (DUT-N) and mitochondrial isoform (DUT-M). [6] [7] [8]

Gene

Northern blot analysis reveals distinct mRNA transcripts for DUT-N (1.1 kb) and DUT-M (1.4 kb). [7] The isoforms are produced from alternative splicing at different 5' exons, with the first exon of DUT-N occurring 767 base pairs downstream of the first exon in DUT-M. [7] [8] Regulation at different promoters has been proposed to account for the differential expression of these isoforms. [7]

Protein

The mature forms of DUT-N (22 kDa) and DUT-M (23 kDa) are nearly identical except for a short N-terminal region present in DUT-M. The DUT-M precursor (31 kDa) contains an arginine-rich, 69-residue mitochondrial targeting sequence which undergoes post-translational cleavage to effect mitochondrial import. [6] [7] [8] Meanwhile, the monopartite NLS sequence is critical for the function and nuclear localization of DUT-N, which would otherwise accumulate in the cytoplasm. [6] [8] Though both isoforms contain the NLS, the sequence in DUT-M is sequestered away from cognate karyopherins. [6] The isoelectric points of DUT-N (6.0) and DUT-M (8.1) correspond to the pH of their respective subcellular compartments. [7]

DUT is a homotrimer with three active sites formed by each of its three subunits. [8] Typically, each subunit forms an eight-stranded barrel that swaps C-terminal β-strands with the other subunits to assemble into the trimer structure. In addition to the β-strand swapping, these subunits interact via extended bimolecular interfaces and three-fold central channel. [9] As a member of the dUTPase family, DUT requires the presence of a divalent metal ion such as Mg2+ for their enzymatic function. [10] DUT-N also contains a consensus cyclin-dependent kinase phosphorylation site that is phosphorylated at the serine as part of its cell cycle regulation. [7]

Function

DUT is a member of the dUTPase family, which is known for catalyzing the pyrophosphoralysis of dUTP into dUMP and inorganic pyrophosphate. This function contributes to DNA replication and repair via de novo thymidylate biosynthesis, as the dUMP product is methylated by thymidylate synthase (TS) to form dTMP, which is then phosphorylated to dTTP. [6] [7] [8] [11] DUT is also crucial for maintaining genome integrity by reducing cellular dUTP levels, thereby preventing the repeated cycles of uracil misincorporation into DNA and DNA repair-mediated strand breaks that would lead to cell death. [6] [7] [8] [10] [11]

In addition to their different localizations, the two DUT isoforms display different expression patterns: while DUT-M is constitutively expressed, DUT-N is under cell cycle control and notably upregulated during S phase. [6] [7] These expression patterns correspond with their roles in the DNA replication cycle of their respective genomes, and thus indicate different regulatory mechanisms affecting each isoform. [7]

Mechanism

The dUTP hydrolysis cycle can be outlined in the following four enzymatic steps: (i) fast substrate binding, (ii) isomerization of the enzyme-substrate complex into the catalytically competent conformation, (iii) hydrolysis of the substrate, and (iv) rapid, non-ordered release of the products. [12]

Clinical significance

Since many chemotherapeutic agents such as 5-fluorouracil treat neoplastic diseases, including head and neck cancer, breast cancer, and gastrointestinal cancer, by targeting TS in thymidylate metabolism, DUT may protect against the cytotoxic side effects by countering dUTP accumulation. [7] [8] [11] [13] [14] At the same time, high levels of DUT-N have been associated with chemoresistance and faster tumor progression, and thus, could also serve as a prognostic marker for overall survival and response to chemotherapy. [7] [8] [11] [12] [13] Similarly, DUT is significantly overexpressed in hepatocellular carcinoma and may serve as a prognostic marker for the cancer. [15] Notably, DUT expression is regulated by the tumor suppressor gene p53 in order to promote apoptosis of tumor cells.<pmid19015155/> Abnormal DUT expression and localization has been speculated to promote cancer transformation. [8]

Interactions

DUT interacts with dUTP to catalyze its hydrolysis into dUMP and pyrophosphate. [5] E2F and Sp1 enhance DUT expression by binding its promoter, while p53 inhibits DUT transcription by binding its promoter. A putative NF-κB binding site was also identified in the DUT promoter. [14]

References

  1. 1 2 3 GRCh38: Ensembl release 89: ENSG00000128951 – Ensembl, May 2017
  2. 1 2 3 GRCm38: Ensembl release 89: ENSMUSG00000027203 – Ensembl, May 2017
  3. ↑ "Human PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
  4. ↑ "Mouse PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
  5. 1 2 3 "Entrez Gene: DUT dUTP pyrophosphatase".
  6. 1 2 3 4 5 6 7 Róna G, Marfori M, Borsos M, Scheer I, Takács E, Tóth J, Babos F, Magyar A, Erdei A, Bozóky Z, Buday L, Kobe B, Vértessy BG (Dec 2013). "Phosphorylation adjacent to the nuclear localization signal of human dUTPase abolishes nuclear import: structural and mechanistic insights" (PDF). Acta Crystallographica Section D. 69 (Pt 12): 2495–505. doi:10.1107/S0907444913023354. PMID   24311590.
  7. 1 2 3 4 5 6 7 8 9 10 11 12 13 Ladner RD, Caradonna SJ (Jul 1997). "The human dUTPase gene encodes both nuclear and mitochondrial isoforms. Differential expression of the isoforms and characterization of a cDNA encoding the mitochondrial species". The Journal of Biological Chemistry. 272 (30): 19072–80. doi: 10.1074/jbc.272.30.19072 . PMID   9228092.
  8. 1 2 3 4 5 6 7 8 9 10 Tinkelenberg BA, Fazzone W, Lynch FJ, Ladner RD (Jul 2003). "Identification of sequence determinants of human nuclear dUTPase isoform localization". Experimental Cell Research. 287 (1): 39–46. doi:10.1016/s0014-4827(03)00048-x. PMID   12799180.
  9. ↑ Takács E, Barabás O, Petoukhov MV, Svergun DI, Vértessy BG (Mar 2009). "Molecular shape and prominent role of beta-strand swapping in organization of dUTPase oligomers". FEBS Letters. 583 (5): 865–71. doi: 10.1016/j.febslet.2009.02.011 . PMID   19302784.
  10. 1 2 Persson R, Cedergren-Zeppezauer ES, Wilson KS (Dec 2001). "Homotrimeric dUTPases; structural solutions for specific recognition and hydrolysis of dUTP". Current Protein & Peptide Science. 2 (4): 287–300. doi:10.2174/1389203013381035. PMID   12369926.
  11. 1 2 3 4 Ladner RD (Dec 2001). "The role of dUTPase and uracil-DNA repair in cancer chemotherapy". Current Protein & Peptide Science. 2 (4): 361–70. doi:10.2174/1389203013380991. PMID   12374095.
  12. 1 2 Tóth J, Varga B, Kovács M, Málnási-Csizmadia A, Vértessy BG (Nov 2007). "Kinetic mechanism of human dUTPase, an essential nucleotide pyrophosphatase enzyme". The Journal of Biological Chemistry. 282 (46): 33572–82. doi: 10.1074/jbc.M706230200 . hdl: 10831/92830 . PMID   17848562.
  13. 1 2 Ladner RD, Lynch FJ, Groshen S, Xiong YP, Sherrod A, Caradonna SJ, Stoehlmacher J, Lenz HJ (Jul 2000). "dUTP nucleotidohydrolase isoform expression in normal and neoplastic tissues: association with survival and response to 5-fluorouracil in colorectal cancer". Cancer Research. 60 (13): 3493–503. PMID   10910061.
  14. 1 2 Wilson PM, Fazzone W, LaBonte MJ, Lenz HJ, Ladner RD (Jan 2009). "Regulation of human dUTPase gene expression and p53-mediated transcriptional repression in response to oxaliplatin-induced DNA damage". Nucleic Acids Research. 37 (1): 78–95. doi:10.1093/nar/gkn910. PMC   2615606 . PMID   19015155.
  15. ↑ Takatori H, Yamashita T, Honda M, Nishino R, Arai K, Yamashita T, Takamura H, Ohta T, Zen Y, Kaneko S (Mar 2010). "dUTP pyrophosphatase expression correlates with a poor prognosis in hepatocellular carcinoma". Liver International. 30 (3): 438–46. doi:10.1111/j.1478-3231.2009.02177.x. PMID   19968781. S2CID   26272889.

Further reading

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