“Terminal deoxynucleotidyl transferase”的意思、由来-开放百科全书

TdT is absent in fetal liver HSCs, significantly impairing junctional diversity in B-cells during the fetal period.^[5]

Function and regulation

TdT catalyses the addition of nucleotides to the 3' terminus of a DNA molecule. Unlike most DNA polymerases, it does not require a template. The preferred substrate of this enzyme is a 3'-overhang, but it can also add nucleotides to blunt or recessed 3' ends. Cobalt is a necessary cofactor, however the enzyme catalyzes reaction upon Mg and Mn administration in vitro.

TdT is expressed mostly in the primary lymphoid organs, like the thymus and bone marrow. Regulation of its expression occurs via multiple pathways. These include protein-protein interactions, like those with TdIF1. TdIF1 is another protein that interacts with TdT to inhibit its function by masking the DNA binding region of the TdT polymerase. The regulation of TdT expression also exists at the transcriptional level, with regulation influenced by stage-specific factors, and occurs in a developmentally restrictive manner.^[3]^[6]^[7] Although expression is typically found to be in the primary lymphoid organs, recent work has suggested that stimulation via antigen can result in secondary TdT expression along with other enzymes needed for gene rearrangement outside of the thymus for T-cells.^[8]

Mechanism

Upon the action of the RAG 1/2 enzymes, the cleaved double stranded DNA is left with hairpin structures at the end of each DNA segment created by the cleavage event. The hairpins are both opened by the Artemis complex, which has endonuclease activity when phosphorylated, providing the free 3' OH ends for TdT to act upon. Once the Artemis complex has done its job and added palindromic nucleotides (P-nucleotides) to the newly opened DNA hairpins, the stage is set for TdT to do its job. TdT is now able to come in and add N-nucleotides to the existing P-nucleotides in a 5' to 3' direction that polymerases are known to function. On average 2-5 random base pairs are added to each 3' end generated after the action of the Artemis complex. The number of bases added is enough for the two newly synthesized ssDNA segments to undergo micro homology alignment during non-homologous end joining according to the normal Watson-Crick base pairing patterns (A-T, C-G). From there unpaired nucleotides are excised by an exonuclease, like the Artemis Complex (which has exonuclease activity in addition to endonuclease activity), and then template dependent polymerases are able to fill the gaps, finally creating the new coding joint with the action of ligase to combine the segments. Although TdT does not discriminate among the four base pairs when adding them to the N-nucleotide segments, it has shown a bias for guanine and cytosine base pairs.^[3]

Polymerase λ has also been found to exhibit similar template independent synthetic activity. Along with activity as a terminal transferase, it is known to also work in a template dependent fashion. This is interesting because TdT works exclusively via a template independent pathway for the addition of nucleotides.^[9]

Uses

Terminal transferase has applications in molecular biology. It can be used in RACE to add nucleotides that can then be used as a template for a primer in subsequent PCR. It can also be used to add nucleotides labeled with radioactive isotopes, for example in the TUNEL assay (Terminal deoxynucleotidyl transferase dUTP Nick End Labeling) for the demonstration of apoptosis (which is marked, in part, by fragmented DNA). It is also used in the immunofluorescence assay for the diagnosis of acute lymphoblastic leukemia.^[10]

In immunohistochemistry, antibodies to TdT can be used to demonstrate the presence of immature T and B cells and pluripotent hematopoietic stem cells, which possess the antigen, while mature lymphoid cells are always TdT-negative. While TdT-positive cells are found in small numbers in healthy lymph nodes and tonsils, the malignant cells of acute lymphoblastic leukaemia are also TdT-positive, and the antibody can, therefore, be used as part of a panel to diagnose this disease and to distinguish it from, for example, small cell tumours of childhood.^[11]

TdT has also seen recent application in the De Novo synthesis of oligonucleotides, with TdT-dNTP tethered analogues capable of primer extension by 1 nt at a time. ^[12] In other words, the enzyme TdT has demonstrated the capability of making synthetic DNA by adding one letter at a time to a primer sequence.

See also

References

1. ^{{cite journal | vauthors = Isobe M, Huebner K, Erikson J, Peterson RC, Bollum FJ, Chang LM, Croce CM | title = Chromosome localization of the gene for human terminal deoxynucleotidyltransferase to region 10q23-q25 | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 82 | issue = 17 | pages = 5836–40 | date = Sep 1985 | pmid = 3862101 | pmc = 390648 | doi = 10.1073/pnas.82.17.5836 }}
2. ^{{cite journal | vauthors = Yang-Feng TL, Landau NR, Baltimore D, Francke U | title = The terminal deoxynucleotidyltransferase gene is located on human chromosome 10 (10q23----q24) and on mouse chromosome 19 | journal = Cytogenetics and Cell Genetics | volume = 43 | issue = 3-4 | pages = 121–6 | year = 1986 | pmid = 3467897 | doi = 10.1159/000132309 }}
3. ^¹²{{cite journal | vauthors = Motea EA, Berdis AJ | title = Terminal deoxynucleotidyl transferase: the story of a misguided DNA polymerase | journal = Biochimica et Biophysica Acta | volume = 1804 | issue = 5 | pages = 1151–66 | date = May 2010 | pmid = 19596089 | pmc = 2846215 | doi = 10.1016/j.bbapap.2009.06.030 }}
4. ^{{cite journal | vauthors = Haeryfar SM, Hickman HD, Irvine KR, Tscharke DC, Bennink JR, Yewdell JW | title = Terminal deoxynucleotidyl transferase establishes and broadens antiviral CD8+ T cell immunodominance hierarchies | language = en | journal = Journal of Immunology | volume = 181 | issue = 1 | pages = 649–59 | date = Jul 2008 | pmid = 18566432 | doi = 10.4049/jimmunol.181.1.649 | pmc=2587314}}
5. ^{{cite book |last1=Hardy |first1=Richard |editor1-first=William |editor1-last=Paul | name-list-format = vanc | title=Fundamental Immunology |type=Book |edition=6th |year=2008 |publisher=Lippincott Williams & Wilkins |location=Philadelphia |isbn=0-7817-6519-6 |pages=237–269 |chapter=Chapter 7: B Lymphocyte Development and Biology}}
6. ^{{cite journal | vauthors = Cherrier M, D'Andon MF, Rougeon F, Doyen N | title = Identification of a new cis-regulatory element of the terminal deoxynucleotidyl transferase gene in the 5' region of the murine locus | journal = Molecular Immunology | volume = 45 | issue = 4 | pages = 1009–17 | date = Feb 2008 | pmid = 17854898 | doi = 10.1016/j.molimm.2007.07.027 }}
7. ^{{cite journal | vauthors = Kubota T, Maezawa S, Koiwai K, Hayano T, Koiwai O | title = Identification of functional domains in TdIF1 and its inhibitory mechanism for TdT activity | language = en | journal = Genes to Cells | volume = 12 | issue = 8 | pages = 941–59 | date = Aug 2007 | pmid = 17663723 | doi = 10.1111/j.1365-2443.2007.01105.x }}
8. ^{{cite journal | vauthors = Zhang Y, Shi M, Wen Q, Luo W, Yang Z, Zhou M, Ma L | title = Antigenic stimulation induces recombination activating gene 1 and terminal deoxynucleotidyl transferase expression in a murine T-cell hybridoma | journal = Cellular Immunology | volume = 274 | issue = 1-2 | pages = 19–25 | date = 2012-01-01 | pmid = 22464913 | doi = 10.1016/j.cellimm.2012.02.008 }}
9. ^{{cite journal | vauthors = Maga G, Ramadan K, Locatelli GA, Shevelev I, Spadari S, Hübscher U | title = DNA elongation by the human DNA polymerase lambda polymerase and terminal transferase activities are differentially coordinated by proliferating cell nuclear antigen and replication protein A | language = en | journal = The Journal of Biological Chemistry | volume = 280 | issue = 3 | pages = 1971–81 | date = Jan 2005 | pmid = 15537631 | doi = 10.1074/jbc.M411650200 }}
10. ^{{cite journal | vauthors = Faber J, Kantarjian H, Roberts MW, Keating M, Freireich E, Albitar M | title = Terminal deoxynucleotidyl transferase-negative acute lymphoblastic leukemia | journal = Archives of Pathology & Laboratory Medicine | volume = 124 | issue = 1 | pages = 92–7 | date = Jan 2000 | pmid = 10629138 | doi = 10.1043/0003-9985(2000)124<0092:TDTNAL>2.0.CO;2 }}
11. ^{{cite book|author=Leong, Anthony S-Y|author2=Cooper, Kumarason|author3=Leong, F Joel W-M|year=2003|title=Manual of Diagnostic Cytology|edition=2|publisher=Greenwich Medical Media, Ltd.|pages=413–414|isbn=1-84110-100-1}}
12. ^Palluk, S. et al. Nature Biotechnology 2018, 36, 645-650.

Further reading

{{cite journal | vauthors = O'Malley DP, Orazi A | title = Terminal deoxynucleotidyl transferase-positive cells in spleen, appendix and branchial cleft cysts in pediatric patients | journal = Haematologica | volume = 91 | issue = 8 | pages = 1139–40 | date = Aug 2006 | pmid = 16885057 | doi = }}

{{cite journal | vauthors = Yamashita N, Shimazaki N, Ibe S, Kaneko R, Tanabe A, Toyomoto T, Fujita K, Hasegawa T, Toji S, Tamai K, Yamamoto H, Koiwai O | title = Terminal deoxynucleotidyltransferase directly interacts with a novel nuclear protein that is homologous to p65 | journal = Genes to Cells | volume = 6 | issue = 7 | pages = 641–52 | date = Jul 2001 | pmid = 11473582 | doi = 10.1046/j.1365-2443.2001.00449.x }}

{{cite journal | vauthors = Chang LM, Bollum FJ | title = Molecular biology of terminal transferase | journal = CRC Critical Reviews in Biochemistry | volume = 21 | issue = 1 | pages = 27–52 | year = 1986 | pmid = 3524991 | doi = 10.3109/10409238609113608 }}

{{cite journal | vauthors = Maezawa S, Hayano T, Koiwai K, Fukushima R, Kouda K, Kubota T, Koiwai O | title = Bood POZ containing gene type 2 is a human counterpart of yeast Btb3p and promotes the degradation of terminal deoxynucleotidyltransferase | journal = Genes to Cells | volume = 13 | issue = 5 | pages = 439–57 | date = May 2008 | pmid = 18429817 | doi = 10.1111/j.1365-2443.2008.01179.x }}

{{cite journal | vauthors = Taplin ME, Frantz ME, Canning C, Ritz J, Blumberg RS, Balk SP | title = Evidence against T-cell development in the adult human intestinal mucosa based upon lack of terminal deoxynucleotidyltransferase expression | journal = Immunology | volume = 87 | issue = 3 | pages = 402–7 | date = Mar 1996 | pmid = 8778025 | pmc = 1384108 | doi = 10.1046/j.1365-2567.1996.496571.x }}

{{cite journal | vauthors = Grupe A, Li Y, Rowland C, Nowotny P, Hinrichs AL, Smemo S, Kauwe JS, Maxwell TJ, Cherny S, Doil L, Tacey K, van Luchene R, Myers A, Wavrant-De Vrièze F, Kaleem M, Hollingworth P, Jehu L, Foy C, Archer N, Hamilton G, Holmans P, Morris CM, Catanese J, Sninsky J, White TJ, Powell J, Hardy J, O'Donovan M, Lovestone S, Jones L, Morris JC, Thal L, Owen M, Williams J, Goate A | title = A scan of chromosome 10 identifies a novel locus showing strong association with late-onset Alzheimer disease | journal = American Journal of Human Genetics | volume = 78 | issue = 1 | pages = 78–88 | date = Jan 2006 | pmid = 16385451 | pmc = 1380225 | doi = 10.1086/498851 }}

{{cite journal | vauthors = Dworzak MN, Fritsch G, Fröschl G, Printz D, Gadner H | title = Four-color flow cytometric investigation of terminal deoxynucleotidyl transferase-positive lymphoid precursors in pediatric bone marrow: CD79a expression precedes CD19 in early B-cell ontogeny | journal = Blood | volume = 92 | issue = 9 | pages = 3203–9 | date = Nov 1998 | pmid = 9787156 | doi = }}

{{cite journal | vauthors = Fujita K, Shimazaki N, Ohta Y, Kubota T, Ibe S, Toji S, Tamai K, Fujisaki S, Hayano T, Koiwai O | title = Terminal deoxynucleotidyltransferase forms a ternary complex with a novel chromatin remodeling protein with 82 kDa and core histone | journal = Genes to Cells | volume = 8 | issue = 6 | pages = 559–71 | date = Jun 2003 | pmid = 12786946 | doi = 10.1046/j.1365-2443.2003.00656.x }}

{{cite journal | vauthors = Kubota T, Maezawa S, Koiwai K, Hayano T, Koiwai O | title = Identification of functional domains in TdIF1 and its inhibitory mechanism for TdT activity | journal = Genes to Cells | volume = 12 | issue = 8 | pages = 941–59 | date = Aug 2007 | pmid = 17663723 | doi = 10.1111/j.1365-2443.2007.01105.x }}

{{cite journal | vauthors = Suzuki Y, Yoshitomo-Nakagawa K, Maruyama K, Suyama A, Sugano S | title = Construction and characterization of a full length-enriched and a 5'-end-enriched cDNA library | journal = Gene | volume = 200 | issue = 1-2 | pages = 149–56 | date = Oct 1997 | pmid = 9373149 | doi = 10.1016/S0378-1119(97)00411-3 }}

{{cite journal | vauthors = Bridges SL | title = Frequent N addition and clonal relatedness among immunoglobulin lambda light chains expressed in rheumatoid arthritis synovia and PBL, and the influence of V lambda gene segment utilization on CDR3 length | journal = Molecular Medicine | volume = 4 | issue = 8 | pages = 525–53 | date = Aug 1998 | pmid = 9742508 | pmc = 2230400 | doi = }}

{{cite journal | vauthors = Liu L, McGavran L, Lovell MA, Wei Q, Jamieson BA, Williams SA, Dirks NN, Danielson MS, Dubie LM, Liang X | title = Nonpositive terminal deoxynucleotidyl transferase in pediatric precursor B-lymphoblastic leukemia | journal = American Journal of Clinical Pathology | volume = 121 | issue = 6 | pages = 810–5 | date = Jun 2004 | pmid = 15198352 | doi = 10.1309/QD18-PPV1-NH3T-EUTF }}

{{cite journal | vauthors = Yang B, Gathy KN, Coleman MS | title = Mutational analysis of residues in the nucleotide binding domain of human terminal deoxynucleotidyl transferase | journal = The Journal of Biological Chemistry | volume = 269 | issue = 16 | pages = 11859–68 | date = Apr 1994 | pmid = 8163485 | doi = }}

{{cite journal | vauthors = Thai TH, Kearney JF | title = Distinct and opposite activities of human terminal deoxynucleotidyltransferase splice variants | journal = Journal of Immunology | volume = 173 | issue = 6 | pages = 4009–19 | date = Sep 2004 | pmid = 15356150 | doi = 10.4049/jimmunol.173.6.4009 }}

{{cite journal | vauthors = Shimazaki N, Fujita K, Koiwai O | title = [Expression and function of terminal deoxynucleotidyl-transferase and discovery of novel DNA polymerase mu] | journal = Seikagaku. The Journal of Japanese Biochemical Society | volume = 74 | issue = 3 | pages = 227–32 | date = Mar 2002 | pmid = 11974916 | doi = }}

{{cite journal | vauthors = Mahajan KN, Mitchell BS | title = Role of human Pso4 in mammalian DNA repair and association with terminal deoxynucleotidyl transferase | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 100 | issue = 19 | pages = 10746–51 | date = Sep 2003 | pmid = 12960389 | pmc = 196874 | doi = 10.1073/pnas.1631060100 }}

{{cite journal | vauthors = Mahajan KN, Gangi-Peterson L, Sorscher DH, Wang J, Gathy KN, Mahajan NP, Reeves WH, Mitchell BS | title = Association of terminal deoxynucleotidyl transferase with Ku | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 96 | issue = 24 | pages = 13926–31 | date = Nov 1999 | pmid = 10570175 | pmc = 24167 | doi = 10.1073/pnas.96.24.13926 }}