Switching to tenofovir vs continuing entecavir for hepatitis B virus with partial virologic response to entecavir: a randomized controlled trial.

Entecavir 0.5 mg (ETV) is widely used among treatment-naïve chronic hepatitis B (CHB) patients. However, 10%-30% of patients show partial virologic response (PVR) to the drug. If the hepatitis B virus (HBV) continues to replicate, the underlying liver disease may progress. Herein, we compared the efficacy of switching to tenofovir disoproxil fumarate (TDF) with that of continuing ETV in CHB patients with PVR to ETV. This was an open-label randomized controlled trial including CHB patients who had been receiving 0.5 mg of ETV for >12 months, but who still had detectable HBV DNA levels of >60 IU/mL without known resistance to ETV. Sixty patients were enrolled and 45 qualified for the study: Twenty-two patients were randomly assigned into the TDF group and 23 into the ETV group. After 12 months of treatment, the virologic response rate (HBV DNA <20 IU/mL) was significantly higher in the TDF group than in the ETV group, as measured using per-protocol analysis (55% vs 20%; P = .022) and intention-to-treat analysis (50% vs 17.4%; P = .020). The reduction in HBV DNA was greater (-1.13 vs -0.67 log10 IU/mL; P = .024), and the mean HBV DNA level was lower (1.54 vs 2.01 log10 IU/mL; P = .011) in the TDF group than in the ETV group. In conclusion, to achieve optimal response in CHB patients with PVR to ETV, switching to TDF would be a better strategy than continuing ETV. Appropriate modification of therapy would further improve the outcome of chronic HBV infection.

Abdominal adiposity intensifies the negative effects of ambient air pollution on lung function in Korean men.

BACKGROUND: Some studies have provided the possibility that adipose tissue may mediate air pollution-induced lung dysfunction. Studies using quantified fat mass data are needed to understand the biological mechanisms between adipocyte and air pollution in lung function. We aimed to investigate whether abdominal adiposity measured by computed tomography (CT) modifies the effects of air pollution on lung function in Korean men. METHODS: A total of 1876 men who visited one of two health checkup centers were recruited for this study. Adiposity traits such as visceral adipose tissue (VAT), subcutaneous adipose tissue (SAT) and total adipose tissue (TAT) areas were measured by CT. We used the annual mean concentrations of ambient air pollutants including nitrogen dioxide (NO2) and particulate matter with an aerodynamic diameter ⩽10 µm (PM10). RESULTS: Interquartile range (IQR) increase in annual mean concentration of NO2 was significantly associated with a 2.5% lower forced expiratory volume in 1 s (FEV1) and 2.9% lower forced vital capacity (FVC) (both P<0.05). The decrease in lung function was more strongly associated with adiposity traits than with body mass index. In a stratified analysis of adiposity, compared with subjects with low-VAT area (VAT⩽200 cm2), those with high-VAT area (VAT>200 cm2) showed a rapid decrease in FEV1 with each IQR increase in PM10 (ß=-0.0812; 95% confidence interval (CI) =-0.1590, -0.0035) and NO2 (ß=-0.0979; 95% CI=-0.1611, -0.0346). In the high-VAT group, each IQR increase in NO2 content was significantly associated with a 10.6% decrease (ß=-0.1056; 95% CI=-0.1770, -0.0343) in FVC. SAT and TAT areas showed similar patterns. CONCLUSIONS: We report the first finding that abdominal adiposity intensifies the inverse relationship between air pollution and lung function.

Curcumin decreases oleic acid-induced lipid accumulation via AMPK phosphorylation in hepatocarcinoma cells.

BACKGROUND AND OBJECTIVES: Non-alcoholic fatty liver disease (NAFLD) is one of the most common metabolic syndromes and is characterized by the accumulation of hepatic triglycerides (TG), which result from an imbalance between uptake, synthesis, export, and oxidation of fatty acids. Curcumin is a polyphenol derived from the herbal remedy and dietary spice turmeric, was found to prevent obesity and diabetes in mouse models. However, a hypolipidemic effect of curcumin in oleic acid- induced hepatocarcinoma cells has not been reported. In this study, we examined the effect of curcumin on reducing lipid accumulation in hepatic cells. MATERIALS AND METHODS: Hepatocytes were treated with oleic acid (OA) containing with or without curcumin to observe the lipid accumulation by Oil Red O stain. We also tested the effects of curcumin on triglycerides (TG) and total cholesterol (TC) in HepG2 cells. Western blot and reverse transcription polymerase chain reaction (RT-PCR) was used to measure sterol regulatory element binding proteins-1 (SREBP-1), fatty acid synthase (FAS), peroxisome proliferator-activated receptor (PPAR)-α, and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) expression. RESULTS: Curcumin suppressed OA-induced lipid accumulation and TG and TC levels. Also, curcumin decreased hepatic lipogenesis such as SREBP-1, and FAS. Besides, we also found out the antioxidative effect of curcumin by increasing the expression of PPARα. Curcumin increased AMPK phosphorylation in hepatocytes. CONCLUSIONS: These results indicated that curcumin has the same ability to activate AMPK and then reduce SREBP-1, and FAS expression, finally leading to inhibit hepatic lipogenesis and hepatic antioxidative ability. In this report, we found curcumin exerted a regulatory effect on lipid accumulation by decreasing lipogenesis in hepatocyte. Therefore, curcumin extract may be active in the prevention of fatty liver.

Anti-inflammatory effects of Hylomecon hylomeconoides in RAW 264.7 cells.

BACKGROUND AND OBJECTIVES: Papaveraceae serve as a rich source of various alkaloids which have anti-inflammatory effect. MATERIALS AND METHODS: In this study, we investigated the effect of Hylomecon hylomeconoides ethanol extract (HHE) on lipopolysaccharide (LPS)-induced NO and interleukin-6 (IL-6) production in RAW 264.7 cells. RESULTS: HHE inhibited LPS-induced NO and IL-6 production. Moreover, HHE suppressed the phosphorylation of ERK1/2 and p38 in LPS-induced RAW 264.7 in a dose-dependent manner. Furthermore, major constituents, dihydrosanguinarine and 6-methoxydihydrosanguinarine, of the chloroform-soluble extract were analyzed. CONCLUSIONS: Taken together, the results of this study indicate that the anti-inflammatory effects of HHE may occur via the inhibition of NO and IL-6 expression through the down-regulation of MAP kinase (ERK1/2, p38) phosphorylation in RAW 264.7 cells.

Virologic and biochemical responses to clevudine in patients with chronic HBV infection-associated cirrhosis: data at week 48.

Clevudine shows high rates of virologic and biochemical responses in patients with chronic hepatitis B. However, the efficacy and safety of clevudine in patients with cirrhosis are unknown. The aims of this study were to evaluate the safety and to assess the virologic and the biochemical responses to clevudine in patients with cirrhosis with chronic hepatitis B virus (HBV) infection. We reviewed data from treatment-naïve patients with chronic hepatitis B with and without cirrhosis who started clevudine between April 2007 and March 2008 (n = 52, hepatitis B without cirrhosis n = 21 and chronic hepatitis B with cirrhosis n = 31) at Korea University Ansan/Guro Hospital. All of the patients were treated for more than 48 weeks. The mean age was older in the patients with cirrhosis. Baseline HBV DNA levels were 6.9 and 7.78 log copies/mL (P = 0.042), and alanine aminotransferase (ALT) levels were 104.9 and 147.4 IU/L (P = 0.204), for those with and without cirrhosis, respectively. Virologic response (HBV DNA <1000 copies/mL) (87.1%vs 71.4%, P = 0.24) and biochemical response (83.9%vs 80.9%, P = 0.99) at week 48 were not significantly different between the two groups. Early virologic response at week 12 was even higher in the patients with cirrhosis (61.3%vs 28.6%, P = 0.026). Neither ALT flare nor newly onset hepatic decompensation was found in the patients with cirrhosis, whereas ALT flare was transiently observed in 14.3% of the chronic hepatitis group. In conclusion, although clevudine may produce a transient elevation of ALT during the early treatment period, such findings were not observed in patients with cirrhosis and the virologic and biochemical responses of the groups were comparable.

The effect of the respiratory cycle on liver stiffness values as measured by transient elastography.

The findings of several studies suggest that liver stiffness values can be affected by the degree of intrahepatic congestion respiration influence intrahepatic blood volume and may affect liver stiffness. We evaluated the influence of respiration on liver stiffness. Transient elastography (TE) was performed at the end of inspiration and at the end of expiration in patients with chronic liver disease. The median values obtained during the inspiration set and during the expiration set were defined as inspiratory and expiratory liver stiffness, respectively. A total of 123 patients with chronic liver disease were enrolled (mean age 49years; 64.2% men). Liver cirrhosis coexisted in 29 patients (23.6%). Expiratory liver stiffness was significantly higher than inspiratory liver stiffness (8.7 vs 7.9kPa, P=0.001), while the expiratory interquartile range/median ratio (IQR ratio) did not differ from the inspiratory IQR ratio. Expiratory liver stiffness was significantly higher than inspiratory liver stiffness in 49 (39.8%) patients (HE group), expiratory liver stiffness was significantly lower than inspiratory stiffness in 15 (12.2%) patients, and there was no difference in 59 (48.0%) patients. Liver cirrhosis was more frequent in those who had a lower liver stiffness reading in expiration, and only the absence of liver cirrhosis was significantly associated with a higher reading in expiration in multivariate analysis. In conclusion, liver stiffness was significantly elevated during expiration especially in patients without liver cirrhosis. The effect of respiration should be kept in mind during TE readings.

Epidemiology of group B streptococcus in Korean pregnant women.

Between January 2006 and May 2008, 2624 pregnant S. Korean women between 35-37 weeks gestation were screened for group B streptococcus (GBS). Resistance to antimicrobials was tested by disk diffusion and serotype determined using co-agglutination assays and microarray methods. Overall, 8% of pregnant women were colonized. Serotype III was the predominant serotype (43.8%), followed by serotypes V (20.3%), Ia (12.1%), and Ib (9.5%). GBS was frequently resistant to clindamycin (54.0%) and erythromycin (25.6%); 3.7% were resistant to cefazolin. More than three-quarters of serotype V were resistant to clindamycin or erythromycin or both, and 71% of serotype III were resistant to clindamycin but only 12% were resistant to erythromycin. GBS prevalence exceeded earlier reports by one-third. This is the first report of cefazolin resistance in Korea. These results underscore the need to establish screening measures and chemoprophylaxis guidelines regarding GBS infections in Korea.

Diagnosis of Pierce's disease using biomarkers specific to Xylella fastidiosa rRNA and Vitis vinifera gene expression.

Pierce's disease (PD), caused by Xylella fastidiosa, represents one of the most damaging diseases of cultivated grape. Management of PD in the vineyard often relies on the removal of infected individuals, which otherwise serve as a source of inoculum for nearby healthy vines. Effective implementation of such control measures requires early diagnosis, which is complicated by the fact that infected vines often harbor high titers of the pathogen in advance of visual symptom development. Here, we report a biomarker system that simultaneously monitors Xylella-induced plant transcripts as well as Xylella ribosomal (r)RNA. Plant biomarker genes were derived from a combination of in silico analysis of grape expressed sequence tags and validation by means of reverse-transcriptase polymerase chain reaction (RT-PCR). Four genes upregulated upon PD infection were individually multiplexed with an X. fastidiosa marker rRNA and scored using either real-time RT-PCR or gel-based conventional RT-PCR techniques. The system was sufficiently sensitive to detect both host gene transcript and pathogen rRNA in asymptomatic infected plants. Moreover, these plant biomarker genes were not induced by water deficit, which is a component of PD development. Such biomarker genes could have utility for disease control by aiding early detection and as a screening tool in breeding programs.

Quantitative Resistance to Bean dwarf mosaic virus in Common Bean Is Associated with the Bct Gene for Resistance to Beet curly top virus.

The dominant resistance gene, Bct, in common bean (Phaseolus vulgaris) confers qualitative resistance to Beet curly top virus, a leafhopper-transmitted geminivirus in the genus Curtovirus. To determine whether this gene confers resistance to other geminiviruses, bean plants of a recombinant inbred population were sap-inoculated with Bean dwarf mosaic virus (BDMV), a whitefly-transmitted bipartite begomovirus in the genus Begomovirus. Results indicated that Bct (or tightly linked gene) is associated with quantitative resistance to BDMV; thus, the Bct locus is associated with resistance to a bean-infecting begomovirus and curtovirus. The difference in the nature of the resistance to these geminiviruses may indicate a role for minor genes in begomovirus resistance or differences in the virus-host interaction. The Bct locus, whether it acts alone or represents a cluster of tightly linked genes, will be useful in breeding for broad-spectrum begomovirus resistance in common bean.

Risk Factors for Dropout From the Liver Transplant Waiting List of Hepatocellular Carcinoma Patients Under Locoregional Treatment.

BACKGROUND: In new organ allocation policy, patients with hepatocellular carcinoma (HCC) experience a 6-month delay in being granted Model for End-Stage Liver Disease exception points. However, it may not be fair for patients at risk of early progression of HCC. METHODS: All patients who were diagnosed as United Network for Organ Sharing (UNOS) stage 1 or 2 of HCC between January 2004 and December 2012 were included. Patients who received surgical resection or liver transplant (LT) as a primary treatment and who did not receive any treatment for HCC were excluded. Patients with baseline Model for End-Stage Liver Disease score ≥22 were also excluded because they have a higher chance of receiving LT. Patients who developed extrahepatic progression within 1 year were considered as high-risk for early recurrence after LT. RESULTS: A total of 586 patients were included. Mean (SD) age was 59.9 (10.3) years and 409 patients (69.8%) were men. The cumulative incidence of estimated dropout was 8.9% at 6 months; size of the maximum nodule (≥3 cm) and nonachievement of complete response were independent factors. Extrahepatic progression developed in 16 patients (2.7%) within 1 year; size of the maximum nodule (4 cm) and alpha-fetoprotein level (>100 ng/mL) were independent predictors. CONCLUSIONS: The estimated dropout rate from the waiting list within 6 months was 8.9%. Advantage points might be needed for patients with maximum nodule size ≥3 cm or those with noncomplete response. However, in patients with maximum nodule size ≥4 cm or alpha-fetoprotein level >100 ng/mL, caution is needed.

Initiation of simian virus 40 DNA synthesis in vitro.

Simian virus 40 (SV40) T antigen can efficiently initiate SV40 origin-dependent DNA synthesis in crude extracts of HeLa cells. Therefore, initiation of SV40 DNA synthesis can be analyzed in detail. We present evidence that antibodies which neutralize proliferating cell nuclear antigen (PCNA) inhibit but do not abolish pulse-labeling of nascent DNA. The lengths of DNA products formed after a 5-s pulse in the absence and presence of anti-PCNA serum averaged 150 and 34 nucleotides, respectively. The small DNAs formed in the presence of anti-PCNA serum underwent little or no increase in size during further incubation periods. The addition of PCNA to reaction mixtures inhibited with anti-PCNA serum largely reversed the inhibitory effect of the antiserum. The small nascent DNAs formed in the presence or absence of anti-PCNA serum products arose from the replication of lagging strands. These results suggest that a PCNA-dependent elongation reaction participates in the synthesis of lagging strands as well as leading strands. We also present evidence that in crude extracts of HeLa cells, DNA synthesis generally does not initiate within the core origin. Initiation of DNA synthesis outside of a genetically defined origin region has not been previously described in a eukaryotic replication system but appears to be a common feature of initiation events in many prokaryotic organisms. Additional results presented indicate that in the absence of nucleoside triphosphates other than ATP, the preinitiation complex remains within or close to the SV40 origin.

Resistance to adefovir dipivoxil in lamivudine resistant chronic hepatitis B patients treated with adefovir dipivoxil.

BACKGROUND: Adefovir dipivoxil (ADV) is a potent nucleotide analogue against both the wild-type and lamivudine (LMV) resistant hepatitis B virus (HBV). The cumulative incidence of ADV resistant mutations in the nucleoside/-tide treatment naive chronic hepatitis B patient (CHB) at weeks 48, 96, and 144 was 0, 0.8-3%, and approximately 5.9%, respectively. AIMS: The aim of this study was to characterise the genotypic and phenotypic mutation profiles to ADV in 67 LMV resistant CHB patients who were treated with ADV. METHODS: Serum HBV DNA was quantified by real time polymerase chain reaction. The ADV mutant was detected using matrix assisted laser desorption/ionisation time of flight mass spectrometry based genotyping assays, termed restriction fragment mass polymorphism (RFMP). RESULTS: RFMP analysis revealed that a total of 11 amino acid substitutions developed in the rt domain of the HBV polymerase in nine patients. The cumulative incidence of genotypic ADV resistance at months 12 and 24 was 6.4% and 25.4%, respectively. The rtA181V, rtN236T, and rtA181T mutations were detected in five, four, and two of the 67 patients at treatment months 12-17, 3-19, and 7-20, respectively. Serial quantification of serum HBV DNA revealed that two patients with the rtA181V mutation, with or without the rtN236T mutation, and one patient with the rtA181T mutation displayed HBV DNA rebound. CONCLUSION: Emergence of the ADV mutation in LMV resistant patients who are treated with ADV appeared to present earlier and more frequently than was reported in previous studies on nucleoside/-tide treatment naive patients.

The endonuclease activity of the yeast Dna2 enzyme is essential in vivo.

Dna2 is a multifunctional enzyme in yeast that possesses endonuclease activity well suited to remove RNA-DNA primers of Okazaki fragments, raising the question of whether endonuclease activity is essential for in vivo Dna2 function. Systematic site-directed mutations of amino acid residues in Saccharomyces cerevisiae DNA2 conserved in the central region of many eukaryotic DNA2 homologs allowed us to identify mutant dna2 alleles that were divided into three groups based on the viability of the mutant cells: (i) viable; (ii) inviable only when expression was repressed; (iii) inviable. Biochemical analyses of recombinant mutant Dna2 proteins isolated from the latter two groups revealed that they possessed normal ATPase/helicase activity, but were impaired in their endonuclease activity. Cells expressing mutant Dna2 enzymes partially impaired in endonuclease activity were viable, but were unable to grow when expression of their mutant Dna2 enzymes was further reduced. Their growth was restored when the mutant Dna2 proteins decreased in nuclease activity were induced to overexpress. In contrast, mutant Dna2 proteins lacking endonuclease activity did not allow cells to grow under any conditions tested. These in vivo and in vitro results demonstrate that the endonuclease activity of Dna2 is essential for Okazaki fragment processing.

Tripartite structure of Saccharomyces cerevisiae Dna2 helicase/endonuclease.

In order to gain insights into the structural basis of the multifunctional Dna2 enzyme involved in Okazaki fragment processing, we performed biochemical, biophysical and genetic studies to dissect the domain structure of Dna2. Proteolytic digestion of Dna2 using subtilisin produced a 127 kDa polypeptide that lacked the 45 kDa N-terminal region of Dna2. Further digestion generated two subtilisin-resistant core fragments of approximately equal size, 58 and 60 kDa. Surprisingly, digestion resulted in a significant (3- to 8-fold) increase in both ATPase and endonuclease activities compared to the intact enzyme. However, cells with a mutant DNA2 allele lacking the corresponding N-terminal region were severely impaired in growth, being unable to grow at 37 degrees C, indicating that the N-terminal region contains a domain critical for a cellular function(s) of Dna2. Analyses of the hydrodynamic properties of and in vivo complex formation by wild-type and/or mutant Dna2 lacking the N-terminal 45 kDa domain revealed that Dna2 is active as the monomer and thus the defect in the mutant Dna2 protein is not due to its inability to multimerize. In addition, we found that the N-terminal 45 kDa domain interacts physically with a central region located between the two catalytic domains. Our results suggest that the N-terminal 45 kDa domain of Dna2 plays a critical role in regulation of the enzymatic activities of Dna2 by serving as a site for intra- and intermolecular interactions essential for optimal function of Dna2 in Okazaki fragment processing. The possible mode of regulation of Dna2 is discussed based upon our recent finding that replication protein A interacts functionally and physically with Dna2 during Okazaki fragment processing.

Evaluation of Cotton Germ Plasm for Resistance to the Whitefly and Cotton Leaf Crumple (CLCr) Disease and Etiology of CLCr in California's Imperial Valley.

Cotton (Gossypium hirsutum) entries were evaluated for resistance to the whitefly (Bemisia tabaci biotype B) and cotton leaf crumple (CLCr) disease during the 1999 to 2001 growing seasons in the Imperial Valley of California. Entries were evaluated for densities of whitefly adults and nymphs, and for CLCr, by visual rating and squash/dot blot hybridization analyses. Differences in whitefly densities were detected among entries, but none were highly resistant, nor was there any correlation with CLCr disease severity. Entries AP 4103 and AP 6101 had relatively low whitefly densities and were highly susceptible (high CLCr disease severity ratings and viral titers), whereas NK 2387C and DPX 1883 also had low whitefly densities but were highly resistant (no symptoms or detectable viral titers). Other entries showed moderate CLCr resistance, which was independent of whitefly density. Geminivirus DNA-A and DNA-B components were consistently detected in cotton leaves with CLCr symptoms by polymerase chain reaction (PCR) with degenerate begomovirus primers, and full-length DNA-A and DNA-B clones were obtained. Cotton seedlings inoculated with these cloned DNAs by particle bombardment developed CLCr symptoms, and progeny virus was whitefly-transmissible. Sequence analysis revealed that these clones comprised the genome of a California isolate of the bipartite begomovirus Cotton leaf crumple virus (CLCrV-CA). Thus, CLCr disease in the Imperial Valley is caused by CLCrV-CA, and cotton entries with high levels of resistance were identified.

Structure and stress-related expression of two cDNAs encoding proteinase inhibitor II of Nicotiana glutinosa L.

Two cDNAs, pNGPI-1 and pNGPI-2, encoding Nicotiana glutinosa proteinase inhibitor II (PI-II) have been cloned, sequenced and identified. The deduced amino acid sequences are 54-82% identical to those of other plant PI-II. The NGPI-1 protein is composed of eight repeated domains, while NGPI-2 contains six repeated regions, each with a putative reactive site. The expression of NGPI-1 is highly regulated in a developmental- and tissue-specific manner, with the transcript being detected in young leaves and floral organs of N. glutinosa plants. In mature leaves, the NGPI-1 gene is rapidly activated by distinct temporal induction patterns in response to pathogen-related (biotic) and wound-related (abiotic) stresses.

Identification of a novel human member of the DEAD box protein family.

The cDNA library of human pancreatic islets was screened with sera from patients with insulin-dependent diabetes mellitus (IDDM). From the library screening, we isolated a novel cDNA, RNA helicase-like protein (RHELP), which exhibited strong sequence homology to p68 RNA helicase, a prototypic member of the DEAD (Asp-Glu-Ala-Asp) box protein family. Sequence analysis of the cDNA revealed that RHELP contained DEAD sequence motif and other conserved motifs of the DEAD box protein family, indicating that RHELP is a new member of this family. DEAD box-containing proteins are involved in the RNA processing, ribosome assembly, spermatogenesis, embryogenesis, and cell growth and division. RHELP showed 42% and 44% amino acid sequence identity to human p68 RNA helicase and yeast DBP2 RNA helicase, respectively, among the DEAD box protein family. Northern blot analysis revealed that RHELP is expressed in most tissues including the liver, lung, tonsil, thymus, and muscle in addition to the pancreatic islets. In vivo or in vitro functions of RHELP as a putative RNA helicase and its potential role as a diabetic autoantigen need to be further investigated.

Genetic analyses of Schizosaccharomyces pombe dna2(+) reveal that dna2 plays an essential role in Okazaki fragment metabolism.

In this report, we investigated the phenotypes caused by temperature-sensitive (ts) mutant alleles of dna2(+) of Schizosaccharomyces pombe, a homologue of DNA2 of budding yeast, in an attempt to further define its function in vivo with respect to lagging-strand synthesis during the S-phase of the cell cycle. At the restrictive temperature, dna2 (ts) cells arrested at late S-phase but were unaffected in bulk DNA synthesis. Moreover, they exhibited aberrant mitosis when combined with checkpoint mutations, in keeping with a role for Dna2 in Okazaki fragment maturation. Similarly, spores in which dna2(+) was disrupted duplicated their DNA content during germination and also arrested at late S-phase. Inactivation of dna2(+) led to chromosome fragmentation strikingly similar to that seen when cdc17(+), the DNA ligase I gene, is inactivated. The temperature-dependent lethality of dna2 (ts) mutants was suppressed by overexpression of genes encoding subunits of polymerase delta (cdc1(+) and cdc27(+)), DNA ligase I (cdc17(+)), and Fen-1 (rad2(+)). Each of these gene products plays a role in the elongation or maturation of Okazaki fragments. Moreover, they all interacted with S. pombe Dna2 in a yeast two-hybrid assay, albeit to different extents. On the basis of these results, we conclude that dna2(+) plays a direct role in the Okazaki fragment elongation and maturation. We propose that dna2(+) acts as a central protein to form a complex with other proteins required to coordinate the multienzyme process for Okazaki fragment elongation and maturation.