[Open, set, and close: DNA clamp loading mechanism revealed by electron microscopy].
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Adenosine Triphosphate↗
Biomedical subjects
Hirofumi Suzuki
Publications and source records attributed to Hirofumi Suzuki.
Open clamp structure in the clamp-loading complex visualized by electron microscopic image analysis.
Ring-shaped sliding clamps and clamp loader ATPases are essential factors for rapid and accurate DNA replication. The clamp ring is opened and resealed at the primer-template junctions by the ATP-fueled clamp loader function. The processivity of the DNA polymerase is conferred by its attachment to the clamp loaded onto the DNA. In eukarya and archaea, the replication factor C (RFC) and the proliferating cell nuclear antigen (PCNA) play crucial roles as the clamp loader and the clamp, respectively. Here, we report the electron microscopic structure of an archaeal RFC-PCNA-DNA complex at 12-A resolution. This complex exhibits excellent fitting of each atomic structure of RFC, PCNA, and the primed DNA. The PCNA ring retains an open conformation by extensive interactions with RFC, with a distorted spring washer-like conformation. The complex appears to represent the intermediate, where the PCNA ring is kept open before ATP hydrolysis by RFC.
DNA Replication↗
Tumor necrosis factor alpha-mediated activation of c-Jun NH(2)-terminal kinase as a mechanism for fumonisin B(1) induced apoptosis in murine primary hepatocytes.
Fumonisin B(1) is a mycotoxin produced by Fusarium verticillioides, frequently associated with corn. It produces species-specific and organ-specific toxicity, including equine leukoencephalomalacia, porcine pulmonary edema, and hepatic or renal damage in most animal species. Fumonisin B(1) perturbs sphingolipid metabolism by inhibiting ceramide synthase. Our previous studies indicated that fumonisin B(1) caused localized activation of cytokines in liver produced by macrophages and other cell types that modulate fumonisin B(1) induced hepatic apoptosis in mice. The role of tumor necrosis factor alpha (TNFalpha) in fumonisin B(1) mediated hepatocyte apoptosis has been established; not much is known about the downstream events leading to apoptosis. In the current study, fumonisin B(1) induced apoptosis in primary culture of liver cells. In consistence with previous reports, fumonisin B(1) caused accumulation of sphingoid bases and led to increase in TNFalpha expression. Phosphorylated and total c-Jun NH(2)-terminal kinase (JNK) activities were increased after 24 h fumonisin B(1) treatment. JNK inhibitor (SP600125) and anti-TNFalpha reduced the apoptosis induced by fumonisin B(1). The role of JNK signaling in fumonisin B(1) induced apoptosis is downstream of TNFalpha production, as fumonisin B(1)-mediated activation of JNK was reduced by the presence of anti-TNFalpha in the medium, whereas the presence of JNK inhibitor did not change the fumonisin B(1) induced TNFalpha expression. Results of this study imply that generation of fumonisin B(1) induced TNFalpha results in modulation of mitogen activated protein kinases, particularly of JNK, and provides a possible mechanism for apoptosis in murine hepatocytes.
Animals↗
[A case of metastatic submandibular lymphnode treated successfully with palliative oral (5-FU + PSK) chemotherapy in the elderly].
The patient was a 87-year-old woman diagnosed as type 2 advanced colon cancer in the ascending colon. The patient underwent right hemicolectomy. The pathological diagnosis showed poorly-differentiated adenocarcinoma, si, ly2, v1, n0 (0/41) and Stage IIIa. The postoperative course was uneventful, and she was discharged on POD 23. But a left submandibular lymph node enlarged rapidly within two months after the operation. Aspiration cytology of the lymph node indicated poorly-differentiated adenocarcinoma, and she was diagnosed as recurrent colon cancer. Combined chemotherapy of 5-FU (200 mg/day/po) and PSK (3.0 g/day/po) was started as palliative chemotherapy. The metastatic lymph nodes were reduced in size within two months after the treatment. Oral administration of 5-FU+PSK succeeded without serious adverse effects or worsening of quality of life. Ten months later, no recurrence was detected on physical examination or computed tomography. We conclude that palliative oral (5-FU+PSK) chemotherapy is useful for recurrent colon cancer in the elderly because of its excellent safety and effectiveness.
Adenocarcinoma↗
beta-Helix is a likely core structure of yeast prion Sup35 amyloid fibers.
We have studied the core structure of amyloid fibers of yeast prion protein Sup35. We developed procedures to prepare straight fibers of relatively uniform diameters from three kinds of fragments; N (1-123), NMp (1-189), and NM (1-253). X-ray fiber diffraction patterns from dried oriented fibers gave common reflections in all three cases; a sharp meridional reflection at 4.7A, and a diffuse equatorial peak at around 9A, apparently supporting the typical "cross-beta" structure with stacked beta-sheets proposed for many different amyloid fibers. However, X-ray fiber diffraction from hydrated fibers showed the meridional reflection at 4.7A but no equatorial reflections at 9A in all three cases, indicating that the stack of beta-sheets in dried fibers is an artifact produced by drying process. Thus, the core structure of these amyloid fibers made of the N domain is likely to be beta-helix nanotube as proposed by Perutz et al.
Amyloid↗
Structure of the rotor of the bacterial flagellar motor revealed by electron cryomicroscopy and single-particle image analysis.
The FliF ring is the base for self-assembly of the bacterial flagellum and the FliF/FliG ring complex is the core of the rotor of the flagellar motor. We report the structures of these two ring complexes obtained by electron cryomicroscopy and single-particle image analysis at 22A and 25A resolution, respectively. Direct comparison of these structures with the flagellar basal body made by superimposing the density maps on the central section reveals many interesting features, such as how the mechanically stable connection between the ring and the rod is formed, how directly FliF domains are involved in the near axial density of the basal body forming the proximal end of the central channel for a potential gating mechanism, some indication of flexibility in the connection of FliF and FliG, and structural and functional similarities to the head-to-tail connectors of bacteriophages.
Bacteria↗
ATP-induced hexameric ring structure of the cyanobacterial circadian clock protein KaiC.
BACKGROUND: KaiA, KaiB and KaiC are cyanobacterial circadian clock proteins. KaiC contains two ATP/GTP-binding Walker's motif As, and mutations in these regions affect the clock oscillations. RESULTS: ATP induced the hexamerization of KaiC. The Km value for the ATP for the hexamerization was 1.9 micro m. Triphosphate nucleotides bound to the two Walker's motif As, and their binding functioned cooperatively for the hexamerization. An unhydrolysable substrate, 5'-adenylylimidodiphosphate (AMPPNP), also induced the hexamerization, indicating that nucleotide binding, but not its hydrolysis, is essential for the hexamerization. Mutations in each of the two Walker's motif As that affect the clock phenotype increased the Km value for ATP and inhibited the hexamerization. Thus, the KaiC hexamerization seems to be necessary for its clock function. The KaiC hexamer has the shape of a hexagonal pot with a diameter and height of approximately 100 A and with a relatively large cavity (73 A deep and 18-34 A wide) inside. This pot-shaped structure suggests that KaiC functions in a similar manner to F1-ATPase, helicase or ATP-dependent protease/chaperon, all of which have dynamic activities inside the central cavity of their hexameric rings. CONCLUSION: ATP-induced KaiC hexamerization is necessary for the clock function of KaiC.
Adenosine Triphosphate↗
Early phase metabolic bone disorders after gastrectomy: influence of active vitamin D treatment.
Metabolic bone disorders are recognized as one of the late complications after gastrectomy. However, the onset time and the extent of bone disorders are still unknown. We examined the influence of active vitamin D treatment on bone metabolism in the early period after gastrectomy. Sixty-three postgastrectomy patients were divided into two groups; active vitamin D treatment group [VD(+)] and no treatment group [VD(-)]. The level of serum calcium and phosphate was increased in the VD(+) group compared with the preoperative level, and parathyroid hormone (PTH-M) was decreased in the VD(+) group. Both 1,25-(OH)2D3 and bone-specific alkaline phosphatase (B-ALP) were increased in the VD(-) group. Cross-linked carboxyterminal telopeptide of type I collagen (I-CTP) was increased in the VD(+) group. There was no change in calcitonin in either group. In conclusion, metabolic bone disorders after gastrectomy have their onset in the early period, and active vitamin D treatment from the early period may be effective in preventing bone disorders.
Aged↗
Dihydropyrimidine dehydrogenase-related enzymes predict efficacy and adverse reactions of UFT1+cisplatin neoadjuvant chemotherapy for gastric cancer.
Dihydropyrimidine dehydrogenase (DPD) and dihydropyrimidinase (DHP) are metabolic enzymes of fluoropyrimidine. UFT containing uracil (U) and Tegafur is the first reported DPD-inhibitory fluoropyrimidine. To clarify the significance of the enzyme activities, we examined the relationships between the effects and adverse reactions, and DPD and DHP activities in gastric cancer treated with UFT1+cisplatin neoadjuvant chemotherapy. Twenty-five gastric cancer patients were administered UFT at 370 mg/m(2)/day for 21 days and cisplatin at 15 mg/m(2)/day for 2 days. Dihydrouracil (DU) and U levels in the urine and DPD activities in the resected tumors were measured. Chemotherapeutic effects were classified histologically into non-responder and responder groups. The responder group accounted for 48% of the patients. All six patients with high DPD activities (> or = 0.08 nmol/min/ww) belonged to the non-responder group and 11 of 19 patients with low DPD activities (<0.08 nmol/min/ww) belonged to the responder group; the difference was significant (p=0.0435). Adverse reactions to UFT occurred in four patients, all of whom were among the six patients with abnormal DU/U values. The incidence of UFT adverse reactions was estimated at 67%. In conclusion, the measured levels of DPD-related enzyme activities appear to be significant for predicting the effects and adverse reactions to chemotherapy.
Amidohydrolases↗
Apoptosis and apoptosis-associated gene products related to the response to neoadjuvant chemotherapy for gastric cancer.
To evaluate the effect of neoadjuvant chemotherapy on gastric cancer, we examined the correlation between induction of apoptosis and expression of p53, Bcl-2, and Bax. Eighty-five patients with advanced gastric cancer were retrospectively divided into the following two groups: 54 patients received 5-fluorouracil (5-FU) at 300 mg/body/day for 14 days and cisplatin (CDDP) at 15 mg/body/day for 2 days as group A; 31 patients without any preoperative chemotherapy as group B. According to histological changes in tumors due to neoadjuvant chemotherapy, the therapeutic effects on tumors were evaluated. The apoptotic index (AI) of group A was significantly higher than that of group B (1.12+/-0.40 vs. 0.67+/-0.24; p<0.01). In group A, the AI of p53-positive cases was significantly lower than that of negative cases (0.92+/-0.32 vs. 1.39+/-0.32; p<0.01). The AI of histological responders was significantly higher than that of non-responders (1.34+/-0.35 vs. 1.02+/-0.38; p<0.01). There was no significant correlation between AI and expression of Bcl-2 or Bax. In group A, histological responders, Bcl-2 positive, and high AI patients had better prognosis, respectively. In conclusion, neoadjuvant chemotherapy for gastric cancer enhanced induction of apoptosis, and AI might be useful to evaluate the effect of neoadjuvant chemotherapy.
Aged↗
[The current status and problems of the intestinal tuberculosis through a review of the Annual of the Pathological Autopsy Cases in Japan].
From 1997 to 1999 the incidence rate of tuberculosis increased and extrapulmonary tuberculosis has also increased during the same period. Among various types of extrapulmonary tuberculosis, intestinal tuberculosis is one of the diseases difficult to diagnose. Recently doctors could not make appropriate diagnosis of intestinal tuberculosis because they did not pay much attention to tuberculosis. With a background described above, we reviewed the Annual of the Pathological Autopsy Cases in Japan in previous five years 1994-1998, in order to investigate a current status of intestinal tuberculosis. Out of 140,358 autopsied cases, the number of cases with active and old tuberculosis was 5,103 (3.6%), in which the intestinal tuberculosis was accompanied in 80 (0.057%). The 80 cases were consisting of 45 males and 35 females and their mean age was 72.7 years old ranging from 28 to 96. Of the 80 intestinal tuberculosis cases, 71 were seen in pulmonary tuberculosis cases and six cases were considered to be intestinal tuberculosis alone. In terms of the location of intestinal tuberculosis the most commonly affected site was ileocecum which occupied 72.7% of all intestinal tuberculosis. Among the 80 cases, 14 cases were complicated with malignant tumor and only 30 cases (37.5%) were diagnosed clinically as intestinal tuberculosis before their death. Clinical diagnosis of other cases were ileus, simple constipation, severe diarrhea, malignant tumor or its tumor. The delay in diagnosing intestinal tuberculosis may result in fatal outcome in the aged persons, so that in general practice much attention should be paid not only to pulmonary but also extrapulmonary tuberculosis including intestinal tuberculosis.
Adult↗
Neoadjuvant chemotherapy with CDDP and 5-fluorouracil for gastric cancer with serosal invasion.
Many gastric cancer patients who recur peritoneally are initially diagnosed with serosal invasion. To clarify the usefulness of neoadjuvant chemotherapy with 5-fluorouracil (5-FU) +/- cisplatin (CDDP), neoadjuvant versus no preoperative chemotherapy for gastric cancer with preoperative serosal invasion was investigated. The patients were treated preoperatively with 5-FU 300 mg/m(2)/day for 2 weeks (F group; n=40), 5-FU 300 mg/m(2)/day for 2 weeks + CDDP 15 mg/m(2)/day for 2 days (FP group; n=80) or nothing (C group; n=100). A total of 78% of patients in C, 65.0% in F and 67.5% in FP group were classified as T3 or higher surgically. In patients without peritoneal metastasis, the positive peritoneal lavage cytology was 29.2% in C, 11.8% in F, and 12.2% in FP patients (p=0.0279). Serosal invasion was found histologically in 60.0% of C, 30.0% of F, and 33.8% of FP patients (p=0.001). There were no serious drug reactions and no increases in morbidity or mortality using either regimen. The 5-year survival rate was 47.0% in F and 50.9% in FP patients, but only 33.2% in C patients (p=0.0042). In conclusion, neoadjuvant chemotherapy with 5-FU +/- CDDP for gastric cancer patients with serosal invasion may reduce positive peritoneal cytology, eliminate cancer cells from the serosal surface, and improve prognosis.
Aged↗