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Biomedical subjects

Daisuke Maeda

Publications and source records attributed to Daisuke Maeda.

10 recordsLinked to original sources
PubMed2006

Ubc9- and mms21-mediated sumoylation counteracts recombinogenic events at damaged replication forks.

The Ubc9 SUMO-conjugating enzyme and the Siz1 SUMO ligase sumoylate several repair and recombination proteins, including PCNA. Sumoylated PCNA binds Srs2, a helicase counteracting certain recombination events. Here we show that ubc9 mutants depend on checkpoint, recombination, and replication genes for growth. ubc9 cells maintain stalled-fork stability but exhibit a Rad51-dependent accumulation of cruciform structures during replication of damaged templates. Mutations in the Mms21 SUMO ligase resemble the ubc9 mutations. However, siz1, srs2, or pcna mutants altered in sumoylation do not exhibit the ubc9/mms21 phenotype. Like ubc9/mms21 mutants, sgs1 and top3 mutants also accumulate X molecules at damaged forks, and Sgs1/BLM is sumoylated. We propose that Ubc9 and Mms21 act in concert with Sgs1 to resolve the X structures formed during replication. Our results indicate that Ubc9- and Mms21-mediated sumoylation functions as a regulatory mechanism, different from that of replication checkpoints, to prevent pathological accumulation of cruciform structures at damaged forks.

DNA Damage↗
PubMed2006

Knockdown of an arbuscular mycorrhiza-inducible phosphate transporter gene of Lotus japonicus suppresses mutualistic symbiosis.

cDNA for a major arbuscular mycorrhiza (AM)-inducible phosphate (Pi) transporter of Lotus japonicus, LjPT3, was isolated from Glomus mosseae-colonized roots. The LjPT3 transcript was expressed in arbuscule-containing cells of the inner cortex. The transport activity of the gene product was confirmed by the complementation of a yeast mutant that lacks high-affinity Pi transporters. In contrast to most AM-inducible Pi transporters thus far reported, LjPT3 has an amino acid sequence that has much in common with those of other members of the Pht1 family of plant Pi transporters, such as StPT3 of potato. To understand better the physiological role of this AM-inducible Pi transporter, knockdown transformants of the gene were prepared through hairy root transformation and RNA interference. Under Pi-limiting conditions, the transformants showed a reduction of Pi uptake via AM and growth retardation. The transformants also exhibited a decrease in G. mosseae arbuscules. Additionally, when Mesorhizobium loti was inoculated into the knockdown transformants in combination with G. mosseae, necrotic root nodules were observed. Based on these findings, we consider that the genetically engineered host plants had monitored insufficient Pi uptake via AM or low expression of LjPT3, excluding the existing fungi and rhizobia and/or preventing further development of the fungal and nodule structures.

Alphaproteobacteria↗
PubMed2005

[Development of three-dimensional kinematic analysis system for artificial knee implants using X-ray fluoroscopic imaging].

To achieve quantitative assessment of 3D dynamic motion of artificial knee implants under clinical conditions, we developed a 3D kinematic analysis system using X-ray fluoroscopic imaging. The 3D pose-estimation technique for knee implants was built on a 2D/3D registration algorithm, which determines the spatial pose for each femoral and tibial component from the knee implant contours and computer-assisted design (CAD) models of the implant. In order to validate the accuracy of the 3D pose estimation and the system, computer simulation and in vitro tests were performed using images of knee implants taken in 10 different poses with respect to X-ray focus. Computer simulation tests showed that the root mean square errors (RMSE) for all variables were less than 1.0 mm 1.0 degrees. In vitro tests showed that the RMSE for translation perpendicular to the X-ray image plane was about 1.5 mm, while the accuracy of the remaining two translational and three rotational variables was found to be sufficient for analyzing knee kinematics. Computation time in 3D pose estimation was then obtained in less than 30 seconds for each frame. In clinical application, dynamic movement in deep knee bending was quantitatively analyzed, and the feasibility and effectiveness of the system was demonstrated.

Algorithms↗
PubMed2005

Visualization of femorotibial contact in total knee arthroplasty using X-ray fluoroscopy.

The purpose of this study was to build a visualization technique of the femorotibial contact in fixed-bearing total knee arthroplasty (TKA) using X-ray fluoroscopy, and to apply this technique to a TKA patient during dynamic motion. In vivo kinametcis of the metallic knee implant was determined using a 2D/3D registration technique, which uses computer assisted design (CAD) model of the implant to estimate the 3D pose of radiopaque metallic femoral and tibial components from a single-plane fluoroscopic image. In fixed-bearing TKA, a 3D pose of radiolucent tibial polyethylene insert can be determined from the estimated pose of the tibial component. To visualize femorotibial contact, the proximity between surfaces of femoral component and tibial insert was calculated, and mapped onto the insert surface model. For the clinical application, dynamic states of contact on the tibial insert were observed including axial rotation and unilateral loading during knee flexion, and post-cam contact of posterior stabilized TKA. The present technique provided us new information and enabled us to better understand the relationship between in vivo knee kinematics and articular shape of the implant.

Arthroplasty, Replacement, Knee↗
PubMed2004

SMC6 is required for MMS-induced interchromosomal and sister chromatid recombinations in Saccharomyces cerevisiae.

SMC6 (RHC18) in Saccharomyces cerevisiae, which is a homologue of the Schizosaccharomyces pombe rad18+ gene and essential for cell viability, encodes a structural maintenance of chromosomes (SMC) family protein. In contrast to the rest of the SMC family of proteins, Smc1-Smc4, which are the components of cohesin or condensin, little is known about Smc6. In this study, we generated temperature sensitive (ts) smc6 mutants of budding yeast and characterized their properties. One ts-mutant, smc6-56, ceased growth soon after up-shift to a non-permissive temperature, arrested in the late S and G2/M phase, and gradually lost viability. smc6-56 cells at a permissive temperature showed a higher sensitivity than wild-type cells to various DNA damaging agents including methyl methanesulfonate (MMS). The rad52 smc6-56 double mutant showed a sensitivity to MMS similar to that of the rad52 single mutant, indicating that Smc6 is involved in a pathway that requires Rad52 to function. Moreover, no induction of interchromosomal recombination and sister chromatid recombination was observed in smc6-56 cells, which occurred in wild-type cells upon exposure to MMS.

Base Sequence↗
PubMed2004

Ubc9 is required for damage-tolerance and damage-induced interchromosomal homologous recombination in S. cerevisiae.

Ubc9 is an enzyme involved in the conjugation of small ubiquitin related modifier (SUMO) to target proteins. A Saccharomyces cerevisiae ubc9 temperature sensitive (ts) mutant showed higher sensitivity to various DNA damaging agents such as methylmethanesulfonate (MMS) and UV at a semi-permissive temperature than wild-type cells. The sensitivity of ubc9ts cells was not suppressed by the introduction of a mutated UBC9 gene, UBC9-C93S, whose product is unable to covalently bind to SUMO and consequently fails to conjugate SUMO to target proteins. Diploid ubc9ts cells were more sensitive to various DNA damaging agents than haploid ubc9ts cells suggesting the involvement of homologous recombination in the sensitivity of ubc9ts cells. The frequency of interchromosomal recombination between heteroalleles, his1-1/his1-7 loci, in wild-type cells was remarkably increased upon exposure to MMS or UV. Although the frequency of spontaneous interchromosomal recombination between the heteroalleles in ubc9ts cells was almost the same as that of wild-type cells, no induction of interchromosomal recombination was observed in ubc9ts cells upon exposure to MMS or UV.

DNA Damage↗
PubMed2004

In vivo kinematics of mobile-bearing knee arthroplasty in deep knee bending motion.

The current study aimed to analyze kinematics during deep knee bending motion by subjects with fully congruent mobile-bearing total knee arthroplasties allowing axial rotation and anteroposterior (AP) gliding. Twelve subjects were implanted with Dual Bearing Knee prostheses (DBK, slot type: Finsbury Orthopaedics, Surrey, UK). These implants include a mobile-bearing insert that is fully congruent with the femoral component throughout flexion and allows axial rotation and limited AP translation. Sequential fluoroscopic images were taken in the sagittal plane during loaded knee bending motion. In vivo kinematics were analyzed using a two- to three-dimensional registration technique, which uses computer-assisted design models to reproduce the spatial position of femoral and tibial components from single-view fluoroscopic images. The average femoral component demonstrated 13.4 degrees external axial rotation for 0-120 degrees flexion. On average, the medial condyle moved anteriorly 6.2 mm for 0-100 degrees flexion, then posteriorly 4.0 mm for 100-120 degrees flexion. On average, the lateral condyle moved anteriorly 1.0 mm for 0-40 degrees flexion, then posteriorly 8.7 mm for 40-120 degrees flexion. The typical subject exhibited a lateral pivot pattern from extension to 60 degrees flexion and a central pivot pattern from 60 degrees to 100 degrees flexion, patterns that are not usually observed in normal knees. Subsequently from 100 degrees to 120 degrees flexion, a rollback pattern was reproduced in which bilateral condyles moved backward.

Aged↗
PubMed2002

Ubc9 is essential for viability of higher eukaryotic cells.

Ubc9 is an enzyme involved in the conjugation of SUMO-1 (small ubiquitin related modifier 1) to target proteins. The SUMO-1 conjugation system is well conserved from yeasts to higher eukaryotes, but many SUMO-1 target proteins reported recently in higher eukaryotic cells, including IkappaBalpha, MDM2, p53, and PML, are not present in yeasts. To determine the physiological roles of SUMO-1 conjugation in higher eukaryotic cells, we constructed a conditional UBC9 mutant of chicken DT40 cells containing the UBC9 transgene under control of a tetracycline-repressible promoter and characterized their loss of function phenotypes. Ubc9 disappeared 3 days after the addition of tetracycline and the increase in viable cell number stopped 4 days after the addition of drug. In contrast to the cases of ubc9 mutants of budding and fission yeasts, which show defects in progression of G2 or early M phase and in chromosome segregation, respectively, we did not observe accumulation of cells in G2/M phase or a considerable increase in the frequency of chromosome missegregation upon depletion of Ubc9 but we did observe an increase in the number of cells containing multiple nuclei, indicating defects in cytokinesis. A considerable portion of the Ubc9-depleted cell population was committed to apoptosis without accumulating in a specific phase of the cell cycle, suggesting that chromosome damages are accumulated in Ubc9-depleted cells, and apoptosis is triggered without activating checkpoint mechanisms under conditions of SUMO-1 conjugation system impairment.

Animals↗