Disease relevance of EXO1

• Germline mutations of EXO1 gene in patients with hereditary nonpolyposis colorectal cancer (HNPCC) and atypical HNPCC forms [1].
• We investigated these families for any indication of predisposition to colorectal cancer or other HNPCC spectrum cancers by means of detailed questionnaires, interviews, and examination of EXO1-null skin leiomyomata for microsatellite instability (MSI) [2].
• In one recent study, germline variants of EXO1 were reported to be associated with predisposition to colorectal cancer in families with phenotypes similar to hereditary nonpolyposis colon cancer (HNPCC) [2].
• We recently identified nine individuals from two British families with multiple cutaneous and uterine leiomyomatosis with independently arising heterozygous germline deletions of 1q42.3 approximately q43 encompassing not only FH, the multiple leiomyomatosis-associated gene, but also several flanking genes, including EXO1 [2].
• We investigated the relationship of single nucleotide polymorphisms (SNPs) at exon 10 (T439M) and exon 13 (P757L) of the EXO1 gene with development, progression and metastasis of colorectal cancer [3].

High impact information on EXO1

• Cleaved single-strand tails will be processed by error-prone DNA polymerase-mediated gap-filling or exonuclease-mediated resection [4].
• A mismatch-containing DNA segment spanned by two strand breaks is removed by the 5'-to-3' activity of MutSalpha-activated exonuclease I. The probable endonuclease active site has been localized to a PMS2 DQHA(X)(2)E(X)(4)E motif [5].
• The upstream cleavage product corresponds to the mature histone mRNA, while the downstream product is degraded by a 5'-3' exonuclease, also dependent on the U7 snRNP [6].
• The purified Artemis protein alone possesses single-strand-specific 5' to 3' exonuclease activity [7].
• These structures unify Mre11's multiple nuclease activities in a single endo/exonuclease mechanism and reveal eukaryotic macromolecular interaction sites by mapping human and yeast Mre11 mutations [8].

Chemical compound and disease context of EXO1

• Deletions into each end of the adenovirus inverted terminal repeat (ITR) were generated with Bal31 exonuclease and the resulting molecules constructed into plasmids which contained two inverted copies of the deleted ITR separated by the bacterial neomycin phosphotransferase gene [9].
• We have investigated the mechanistic basis for the differential toxicity of these three cytosine analogs by comparing the effects of dideoxy-CTP), (+)3TC-triphosphate (TP), and (-)3TC-TP on the polymerase and exonuclease activities of recombinant human Pol gamma [10].
• The method was verified with the ternary 30 kDa complex between the lanthanide-labeled N-terminal domain of the epsilon exonuclease subunit from the Escherichia coli DNA polymerase III, the subunit theta, and thymidine [11].
• Using an in vivo Exonuclease III footprinting assay, we found that estrogen stimulation of MCF-7 cells induced loading of a transcription factor(s) to a portion of the promoter (-124 to -104) that is homologous to the adenovirus major late promoter element [12].
• The requirements for expression of genes under the control of early (alkaline exonuclease) and late (VP5) herpes simplex virus type 1 (HSV-1) gene promoters were examined in a transient expression assay, using the bacterial chloramphenicol acetyltransferase gene as an expression marker [13].

Biological context of EXO1

• METHODS: All 14 exons of EXO1 were scanned for mutations in index patients from 33 families with HNPCC fulfilling the Amsterdam criteria and in 225 index patients suspected of HNPCC [1].
• To determine its role in HNPCC, EXO1 was scanned for germline mutations [1].
• RESULTS: Germline variants of EXO1 were detected in 14 patients, including one splice-site mutation in a family with HNPCC and 13 missense mutations in patients with atypical HNPCC [1].
• Heterozygosity analysis showed one case without EXO1 allelic loss and 12 tumors with loss of the mutant allele and retention of the normal one [1].
• Human exonuclease 1 (hExo1) possesses both 5'exonuclease and flap endonuclease activities and plays a role in DNA repair, recombination, and replication [14].

Anatomical context of EXO1

• The hExoI cDNA comprises 2541 bp, which code for a Mr 94,000 protein that appears to be highly expressed in testis tissue and at very low levels in other tissues [15].
• Northern blot analysis demonstrates that hEXO1 is expressed in high levels in testis; elevated expression was also observed in thymus and colon and to a lesser extent in small intestine, placenta, spleen, and ovary [16].
• Northern blot analysis also revealed that hEXO1/HEX1 is highly expressed in several liver cancer cell lines as well as in colon and pancreas adenocarcinomas, but not in the corresponding non-neoplastic tissue [17].
• Northern blot analysis revealed that HEX1 expression is highest in fetal liver and adult bone marrow, suggesting that the encoded protein may operate prominently in processes specific to hemopoietic stem cell development [18].
• WRN, which functions in cellular recombination pathways via its helicase and exonuclease activities, is not absolutely required for viral replication, as viral yields are only very slightly, if at all, decreased in WRN-deficient human primary fibroblasts compared to control cells [19].

Associations of EXO1 with chemical compounds

• Both the 5' to 3' exonuclease and flap endonuclease activities require a divalent metal cofactor, with Mg(2+) being the preferred metal ion [20].
• Concerted Action of Exonuclease and Gap-dependent Endonuclease Activities of FEN-1 Contributes to the Resolution of Triplet Repeat Sequences (CTG)n- and (GAA)n-derived Secondary Structures Formed during Maturation of Okazaki Fragments [21].
• Furthermore, anti-WRN immunoprecipitates from CML cells treated with STI-571 show increased 3'-->5' exonuclease activity [22].
• Other DNA modifications, including uracil, hypoxanthine and ethenoadenine, did not inhibit AtWRNexo-p. A mutation of a conserved residue within the exonuclease domain (E135A) completely abolished the exonucleolytic activity [23].
• Zn(2+) strongly stimulated the exonuclease activity of a WRN exonuclease domain fragment, suggesting a Zn(2+) binding site in the WRN exonuclease domain [24].

Physical interactions of EXO1

• The EXO1 gene was identified in Saccharomyces cerevisiae as a gene encoding an exonuclease that interacts with MSH2 and functions in mismatch repair and genetic recombination [16].
• In addition, hExoI forms an immunoprecipitable complex with hMLH1/hPMS2 in vivo [25].
• Second, the exonuclease co-purifies with the 160-kDa WRN protein and its associated DNA helicase and ATPase activities through successive steps of ion exchange and affinity chromatography, suggesting that all three activities are physically associated [26].
• Mutation of either the CREB-1 or Sp1/Sp3 binding sites dramatically reduces HEX1/hExo1 promoter activity and elimination of both elements abolishes promoter activity [27].
• EXOI interacts with PCNA [28].

Enzymatic interactions of EXO1

• In addition to its flap endonuclease (FEN) and nick exonuclease (EXO) activities, a new gap endonuclease (GEN) activity has been characterized [29].
• Exogenous expression of exonuclease domain-deleted WRN interferes with the repair of radiation-induced DNA damages [30].
• Optimal reaction conditions for the 3 -5 exonuclease hydrolysis reaction catalyzed by APE1 in vitro have been established, and conditions when photoreactive residues are not removed by APE1 have been chosen [31].
• The 3'-5' exonuclease activity is considered important in proofreading of DNA synthesis catalyzed by DNA polymerase beta [32].
• Moreover, the DNA polymerase I usually applied in the in situ nick translation techniques shows both 5' to 3' and 3' to 5' exonuclease activities [33].

Regulatory relationships of EXO1

• The exonucleolytic and endonucleolytic cleavage activities of human exonuclease 1 are stimulated by an interaction with the carboxyl-terminal region of the Werner syndrome protein [34].
• The 3'-5' exonuclease activity was stimulated 8-10 fold by the addition of HSSB, and this stimulatory effect was preferential to HSSB since other SSBs from E. coli, T4 or adenovirus, had a little or no effect [35].
• Functionally, BLM inhibited the exonuclease activity of WRN [36].
• HNPCC mutations in the human DNA mismatch repair gene hMLH1 influence assembly of hMutLalpha and hMLH1-hEXO1 complexes [37].
• Consistent with recent evidence, we have assembled a novel cell-cycle-dependent model in which DNA-PK inhibits RPA in S-phase of the cell cycle, while BRCA1 inhibits the exonuclease processivity of the MRE11/RAD50/NBS1 (MRN) complex and facilitates the removal of RPA in S and G2 phase [38].

Other interactions of EXO1

• Thus, the genomic instability observed in WRN-/- cells may be at least partially attributed to the lack of interactions between the WRN protein and human nucleases including EXO-1 [34].
• Although the role of EXO-1 is not well understood, its 5' to 3'-exonuclease and flap endonuclease activities may cleave intermediates that arise during DNA metabolism [34].
• Characterization of human exonuclease 1 in complex with mismatch repair proteins, subcellular localization and association with PCNA [39].
• We also show that both hMLH1-hPMS2 (MutLalpha) and hMLH1-hEXO1 complexes are formed in a reaction mixture containing all three proteins [39].
• We report here a novel interaction of the BLM protein with the human 5'-flap endonuclease/5'-3' exonuclease (FEN-1), a genome stability factor involved in Okazaki fragment processing and DNA repair [40].

Analytical, diagnostic and therapeutic context of EXO1

• Fen1 or maturation factor 1 is a 5'-3' exonuclease essential for the degradation of the RNA primer-DNA junctions at the 5' ends of immature Okazaki fragments prior to their ligation into a continuous DNA strand [41].
• The hExoI gene has 14 exons and is located on chromosome 1q43, as determined by fluorescence in situ hybridization and radiation hybrid mapping. hExoI was found to interact strongly with the human MMR protein hMSH2, suggesting its involvement in the MMR process and/or DNA recombination [15].
• The new polymerase species appears to have ATPase and 3'-5' exonuclease activities as shown by the chromatographies [42].
• Site-specific mutagenesis studies demonstrate that amino acids D78, D173 and D225 are critical for Exo1 nuclease function [43].
• The gamma-beta interaction via the initiator results in a DNA loop, as revealed by the novel technique of cyclization enhancement and as confirmed by exonuclease III protection, electron microscopy, and chemical footprinting [44].

References