Effect of pedestrian physique differences on head injury prediction in car-to-pedestrian accidents using deep learning.

OBJECTIVE: The aim of this study is to identify the effects of pedestrian physique differences on head injury prediction in car-to-pedestrian accidents via deep learning. METHODS: A series of parametric studies was carried out using a family car finite element model and MADYMO pedestrian models (AM50, AF05, 6YO). The car model was developed and tuned by 11 impact tests. The initial gaits for the pedestrian models were obtained from volunteer experiments to reproduce 420 pre-crash reactions. Furthermore, by factoring the pedestrian models (3 types), pedestrian directions (2 each), impact positions (3 each), and car velocities (6 levels) with the pre-crash parameters, a total of 45,360 car-to-pedestrian impact simulations were performed. After the simulations, image datasets were created by labeling the pedestrian collision images with head injury criteria of 15 ms (HIC) and dividing the images into training and test data based on model type. Next, deep learning was conducted using the training dataset to obtain trained models. Finally, the effects of pedestrian physique differences on head injury predictions were investigated based on the accuracy of each trained model for test data. RESULTS: The results indicate that the head impact area and the amount of pedestrian information in the image differ depending on the pedestrian models. In head injury prediction with deep learning, AF05 showed the highest prediction accuracy (93.25%), followed by AM50 (90.61%) and 6YO (88.29%). These results using deep learning show that pedestrian physique differences affect the head injury prediction accuracies by 2.32-4.96 points. CONCLUSIONS: Based on the prediction results of the trained models that learned the relationships between the pedestrian collision images and HIC from simulations, we demonstrated the desirable performance of deep learning methods in head injury prediction for adult men, women with small physique, and children. Furthermore, our results confirmed the effect of pedestrian physique differences on the injury prediction accuracy.

A phase III, 52-week, open-label study to evaluate the safety and efficacy of 5% sofpironium bromide (BBI-4000) gel in Japanese patients with primary axillary hyperhidrosis.

A long-term study was conducted in Japanese patients with primary axillary hyperhidrosis who completed the preceding 6-week phase III, confirmatory study of 5% sofpironium bromide gel (hereinafter referred to as sofpironium) to evaluate the safety and efficacy of 52-week treatment with sofpironium. In the long-term study, 185 patients who completed the confirmatory study (94 and 91 patients in the vehicle and sofpironium groups, respectively) started to receive sofpironium (switching and extension groups, respectively), and all these patients were included in both the full analysis set (FAS) and the safety analysis set (SAF). In the FAS, there were more females than males (73.0% vs. 27.0%), and median age was 38.0 years. A total of 161 patients (86 and 75 patients in the switching and extension groups, respectively) completed the study at week 52. The proportions of patients with hyperhidrosis disease severity score of 1 or 2 and a 50% or more reduction in total gravimetric weight of sweat were 57.4% in the switching group and 58.2% in the extension group at week 52. The proportions of patients who achieved this efficacy end-point in the long-term study were similar to that (53.9%) in the sofpironium group in the confirmatory study. In the SAF, the incidences of adverse events (AEs) were 80.9% in the switching group and 83.5% in the extension group, and the incidences of adverse drug reactions were 39.4% and 45.1%, respectively. AEs that occurred in at least 20% of patients in both treatment groups were application site dermatitis (25.5% and 33.0%, respectively) and nasopharyngitis (31.9% and 23.1%, respectively). Reported AEs were generally mild, and there were no deaths. Serious AEs occurred in three patients, but none were considered related to the study drug. In this study, the efficacy of sofpironium was maintained during 52-week treatment, and no new safety risk was observed.

A phase 3, multicenter, randomized, double-blind, vehicle-controlled, parallel-group study of 5% sofpironium bromide (BBI-4000) gel in Japanese patients with primary axillary hyperhidrosis.

A phase 3 study was conducted to verify the efficacy and safety of 5% sofpironium bromide (BBI-4000) gel (hereinafter referred to as sofpironium) administrated for 6 weeks in Japanese patients with primary axillary hyperhidrosis. The primary efficacy end-point was the proportion of patients who satisfied both criteria of a Hyperhidrosis Disease Severity Score (HDSS) of 1 or 2 at the end of 6-week treatment and a 50% or more reduction in total gravimetric weight of sweat at the end of treatment relative to baseline. A total of 281 patients were randomized to receive 5% sofpironium (141 patients) or vehicle (140 patients), and all patients were included in the full analysis set (FAS). In the FAS, 70.1% of patients were female, and the median age was 35.0 years. The proportion of patients who achieved the primary efficacy end-point was 53.9% in the sofpironium group and 36.4% in the vehicle group, with a statistically significant difference of 17.5% (95% confidence interval, 6.02-28.93) between these two groups (P = 0.003). The incidence of adverse events was 44.0% in the sofpironium group and 30.7% in the vehicle group, and the incidence of adverse drug reactions was 16.3% in the sofpironium group and 5.0% in the vehicle group. Reported adverse events were generally mild or moderate in severity. In the sofpironium group, common events (incidence, ≥5%) were nasopharyngitis (14.2%) and dermatitis/erythema at the application site (8.5%/5.7%), with no serious adverse events reported. This study demonstrated the efficacy and safety of 5% sofpironium.

Efficacy of FGF-2 in Periodontal Regeneration in a Case of Severe Intrabony Defect and Furcation Involvement With 15-Month Follow-Up.

INTRODUCTION: Several studies have demonstrated that basic fibroblast growth factor (FGF-2) is one of the most effective growth factors for periodontal regeneration. The Ministry of Health, Labor and Welfare in Japan have approved 0.3% human recombinant FGF-2 for periodontal regeneration, and it has been commercially available since 2016. In this case report, a patient was treated with this periodontal regenerative medicine and demonstrated success at 15-month follow-up, as confirmed by dental X-ray and on cone-beam computed tomography (CBCT). CASE PRESENTATION: A 42-year-old woman with a one by two walled intrabony defect and Class III furcation involvement in tooth #19, and Class II furcation involvement in tooth #18 (lingual) underwent periodontal regenerative surgery with FGF-2 without any bone graft materials. Favorable clinical and radiographic outcomes were noted 15 months after the procedure. The vertical bone defect in tooth #19 showed a clinical attachment level gain of 8 mm. Moreover, CBCT analysis revealed considerable new bone formation in the Class II furcation involvement in tooth #18 and limited bone formation in the Class III furcation involvement in tooth #19. CONCLUSIONS: This case report indicates that FGF-2 showed a positive outcome in terms of periodontal regeneration in a case of one by two wall intrabony defects with Class III furcation involvement. A complete recovery of Class II furcation involvement was observed without artificial bone graft materials.

Application of Deep Learning Methods for Pedestrian Collision Detection Using Dashcam Videos.

The goal of this study is to clarify the usefulness of deep learning methods for pedestrian collision detection using dashcam videos for advanced automatic collision notification, focusing on pedestrians, as they make up the highest number of traffic fatalities in Japan. First, we created a dataset for deep learning from dashcam videos. A total of 78 dashcam videos of pedestrian-to-automobile accidents were collected from a video hosting website and from the Japan Automobile Research Institute (JARI). Individual frames were selected from the video data amounting to a total of 1,212 still images, which were added to our dataset with class and location information. This dataset was then divided to create training, validation, and test datasets. Next, deep learning was performed based on the training dataset to learn the features of pedestrian collision images, which are images that capture pedestrian behavior at the time of the collision. Pedestrian collision detection performance of the trained model was evaluated as the percentage of correct predictions of pedestrian collisions in image data according to varied test sets with different combinations of characteristics. Our results for the proposed method show high-precision collision detection for daytime, clear pedestrian wrap trajectory accident data, including accurate detection of pedestrian collision location information. However, nighttime, unclear accident data resulted in false detection or no detection. Reduction of exposure value and resolution was confirmed to reduce detection rate. The results of the present study suggest the possibility of pedestrian collision detection by deep learning using dashcam videos.

ZFAND5/ZNF216 is an activator of the 26S proteasome that stimulates overall protein degradation.

ZFAND5/ZNF216, a member of the zinc finger AN1-type domain family, is abundant in heart and brain, but is induced in skeletal muscle during atrophy (although not in proteotoxic stress). Because mice lacking ZFAND5 exhibit decreased atrophy, a role in stimulating protein breakdown seemed likely. Addition of recombinant ZFAND5 to purified 26S proteasomes stimulated hydrolysis of ubiquitinated proteins, short peptides, and ATP. Mutating its C-terminal AN1 domain abolished the stimulation of proteasomal peptidase activity. Mutating its N-terminal zinc finger A20 domain, which binds ubiquitin chains, prevented the enhanced degradation of ubiquitinated proteins without affecting peptidase activity. Mouse embryonic fibroblast (MEF) cells lacking ZFAND5 had lower rates of protein degradation and proteasomal activity than WT MEFs. ZFAND5 addition to cell lysates stimulated proteasomal activity and protein degradation. Unlike other proteasome regulators, ZFAND5 enhances multiple 26S activities and overall cellular protein breakdown.

A Ferric-Peroxo Intermediate in the Oxidation of Heme by IsdI.

The canonical heme oxygenases (HOs) catalyze heme oxidation via a heme-bound hydroperoxo intermediate that is stabilized by a water cluster at the active site of the enzyme. In contrast, the hydrophobic active site of IsdI, a heme-degrading enzyme from Staphylococcus aureus, lacks a water cluster and is expected to oxidize heme by an alternative mechanism. Reaction of the IsdI-heme complex with either H2O2 or m-chloroperoxybenzoic acid fails to produce a specific oxidized heme iron intermediate, suggesting that ferric-hydroperoxo or ferryl derivatives of IsdI are not involved in the catalytic mechanism of this enzyme. IsdI lacks a proton-donating group in the distal heme pocket, so the possible involvement of a ferric-peroxo intermediate has been evaluated. Density functional theory (DFT) calculations indicate that heme oxidation involving a ferric-peroxo intermediate is energetically accessible, whereas the energy barrier for a reaction involving a ferric-hydroperoxo intermediate is too great in the absence of a proton donor. We propose that IsdI catalyzes heme oxidation through nucleophilic attack by the heme-bound peroxo species. This proposal is consistent with our previous demonstration by nuclear magnetic resonance spectroscopy that heme ruffling increases the susceptibility of the meso-carbon of heme to nucleophilic attack.

Effect of rebamipide on gastric bleeding and ulcerogenic responses induced by aspirin plus clopidogrel under stimulation of acid secretion in rats.

BACKGROUND AND AIM: We examined the prophylactic effect of rebamipide on gastric bleeding induced by the perfusion of aspirin (acetylsalicylic acid [ASA]) plus clopidogrel under the stimulation of acid secretion in rats. METHODS: Under urethane anesthesia, acid secretion was stimulated by the i.v. infusion of histamine (8 mg/kg/h), and the stomach was perfused with 25 mmol/L ASA at a rate of 0.4 mL/min. Gastric bleeding was evaluated as the concentration of hemoglobin in the perfusate. Clopidogrel (30 mg/kg) was given p.o. 24 h before the perfusion. Rebamipide (3-30 mg/kg) or other antiulcer drugs were given i.d. before the ASA perfusion. RESULTS: Slight gastric bleeding or damage was observed with the perfusion of ASA under the stimulation of acid secretion, whereas these responses were significantly increased in the presence of clopidogrel. Both omeprazole and famotidine inhibited acid secretion and prevented these responses to ASA plus clopidogrel. Rebamipide had no effect on acid secretion, but dose-dependently prevented gastric bleeding in response to ASA plus clopidogrel, with the degree of inhibition being almost equivalent to that of the antisecretory drugs, and the same effects were obtained with the gastroprotective drugs, irsogladine and teprenone. These agents also reduced the severity of gastric lesions, although the effects were less than those of the antisecretory drugs. CONCLUSIONS: These results suggest that the antiplatelet drug, clopidogrel, increases gastric bleeding induced by ASA under the stimulation of acid secretion, and the gastroprotective drug, rebamipide, is effective in preventing the gastric bleeding induced under such conditions, similar to antisecretory drugs.

Hsp70-Bag3 interactions regulate cancer-related signaling networks.

Bag3, a nucleotide exchange factor of the heat shock protein Hsp70, has been implicated in cell signaling. Here, we report that Bag3 interacts with the SH3 domain of Src, thereby mediating the effects of Hsp70 on Src signaling. Using several complementary approaches, we established that the Hsp70-Bag3 module is a broad-acting regulator of cancer cell signaling by modulating the activity of the transcription factors NF-κB, FoxM1, Hif1α, the translation regulator HuR, and the cell-cycle regulators p21 and survivin. We also identified a small-molecule inhibitor, YM-1, that disrupts the Hsp70-Bag3 interaction. YM-1 mirrored the effects of Hsp70 depletion on these signaling pathways, and in vivo administration of this drug was sufficient to suppress tumor growth in mice. Overall, our results defined Bag3 as a critical factor in Hsp70-modulated signaling and offered a preclinical proof-of-concept that the Hsp70-Bag3 complex may offer an appealing anticancer target.

Comparative effects of the anti-platelet drugs, clopidogrel, ticlopidine, and cilostazol on aspirin-induced gastric bleeding and damage in rats.

AIMS: The present study compared the effects of frequently used anti-platelet drugs, such as clopidogrel, ticlopidine, and cilostazol, on the gastric bleeding and ulcerogenic responses induced by intraluminal perfusion with 25 mM aspirin acidified with 25 mM HCl (acidified ASA) in rats. MAIN METHODS: The stomach was perfused with acidified ASA at a rate of 0.4 ml/min for 60 min under urethane anesthesia, and gastric bleeding was measured as the concentration of hemoglobin in the luminal perfusate, which was collected every 15 min. Clopidogrel (10-100mg/kg), ticlopidine (10-300 mg/kg), or cilostazol (3-30 mg/kg) was given p.o. 24h or 90 min before the perfusion of acidified ASA, respectively. KEY FINDINGS: Perfusion of the stomach with acidified ASA alone led to slight bleeding and lesions in the stomach. The pretreatment with clopidogrel, even though it did not cause bleeding or damage by itself, dose-dependently increased the gastric bleeding and ulcerogenic responses induced by acidified ASA. Ticlopidine also aggravated the severity of damage by increasing gastric bleeding, and the effects of ticlopidine at 300 mg/kg were equivalent to those of clopidogrel at 100mg/kg. In contrast, cilostazol dose-dependently decreased gastric bleeding and damage in response to acidified ASA. SIGNIFICANCE: These results demonstrated that clopidogrel and ticlopidine, P2Y12 receptor inhibitors, increased gastric bleeding and ulcerogenic responses to acidified ASA, to the same extent, while cilostazol, a phosphodiesterase III inhibitor, suppressed these responses. Therefore, cilostazol may be safely used in dual anti-platelet therapy combined with ASA, without increasing the risk of gastric bleeding.

Synthesis of L-2,3-diaminopropionic acid, a siderophore and antibiotic precursor.

L-2,3-diaminopropionic acid (L-Dap) is an amino acid that is a precursor of antibiotics and staphyloferrin B a siderophore produced by Staphylococcus aureus. SbnA and SbnB are encoded by the staphyloferrin B biosynthetic gene cluster and are implicated in L-Dap biosynthesis. We demonstrate here that SbnA uses PLP and substrates O-phospho-L-serine and L-glutamate to produce a metabolite N-(1-amino-1-carboxyl-2-ethyl)-glutamic acid (ACEGA). SbnB is shown to use NAD(+) to oxidatively hydrolyze ACEGA to yield α-ketoglutarate and L-Dap. Also, we describe crystal structures of SbnB in complex with NADH and ACEGA as well as with NAD(+) and α-ketoglutarate to reveal the residues required for substrate binding, oxidation, and hydrolysis. SbnA and SbnB contribute to the iron sparing response of S. aureus that enables staphyloferrin B biosynthesis in the absence of an active tricarboxylic acid cycle.

Animal models of gastric bleeding induced by dual antiplatelet therapy using aspirin and clopidogrel--prophylactic effect of antiulcer drugs.

We set up two models of gastric bleeding in rats using low-dose aspirin (ASA) and the antiplatelet drug clopidogrel, a P2Y12 receptor antagonist, and examined the effect of antiulcer drugs on gastric bleeding and ulcerogenic responses under such conditions. Under urethane anesthesia, two catheters were inserted into the rat stomach, one from the esophagus and another through the pylorus via an incision in the duodenum. In the first model, the stomach was perfused with 25 mM ASA dissolved in 50 mM HCl using an infusion pump, and gastric bleeding was measured as the hemoglobin concentration in perfusate collected every 15 min. In the second model, the stomach was perfused with ASA under stimulation of acid secretion by a continuous i.v. infusion of histamine (8 mg/kg/hr). Clopidogrel (30 mg/kg) was given p.o. 24 h before the ASA perfusion, while antiulcer drugs were given i.d. or i.v. 30 min before. Perfusion of the stomach with acidified ASA or ASA under histamine-stimulated acid secretion caused minimal bleeding in the stomach with few lesions. The ulcerogenic and bleeding responses to ASA under these conditions were markedly aggravated by pretreatment with clopidogrel, which by itself provoked neither bleeding nor damage. Antiulcer drugs, such as prostaglandin E2, irsogladine, rebamipide and teprenone, reduced the severity of gastric bleeding and damage in response to ASA plus clopidogrel in the presence of both exogenous and endogenous acid. In contrast, antisecretory drugs such as a proton pump inhibitor and histamine H2 receptor antagonists markedly suppressed the gastric bleeding and lesion responses to ASA plus clopidogrel under histamine-stimulated acid secretion, but had no effect on the responses to acidified ASA plus clopidogrel. These results suggest that clopidogrel increases gastric bleeding induced by ASA and that antiulcer drugs are useful for preventing gastric bleeding caused by the dual antiplatelet therapy.

Single molecule force spectroscopy reveals that iron is released from the active site of rubredoxin by a stochastic mechanism.

Metal centers in metalloproteins involve multiple metal-ligand bonds. The release of metal ions from metalloproteins can have significant biological consequences, so understanding of the mechanisms by which metal ion dissociates has broad implications. By definition, the release of metal ions from metalloproteins involves the disruption of multiple metal-ligand bonds, and this process is often accompanied by unfolding of the protein. Detailed pathways for metal ion release from metalloproteins have been difficult to elucidate by classical ensemble techniques. Here, we combine single molecule force spectroscopy and protein engineering techniques to investigate the mechanical dissociation mechanism of iron from the active site of the simplest iron-sulfur protein, rubredoxin, at the single molecule level. Our results reveal that the mechanical rupture of this simplest iron center is stochastic and follows multiple, complex pathways that include concurrent rupture of multiple ferric-thiolate bonds as well as sequential rupture of ferric-thiolate bonds that lead to the formation of intermediate species. Our results uncover the surprising complexity of the rupture process of the seemingly simple iron center in rubredoxin and provide the first unambiguous experimental evidence concerning the detailed mechanism of mechanical disruption of a metal center in its native protein environment in aqueous solution. This study opens up a new avenue to investigating the rupture mechanism of metal centers in metalloproteins with unprecedented resolution by using single molecule force spectroscopy techniques.

Cytochrome b5 from Giardia lamblia.

The protozoan intestinal parasite Giardia lamblia lacks mitochondria and the ability to make haem yet encodes several putative haem-binding proteins, including three of the cytochrome b(5) family. We cloned one of these (gCYTb5-I) and expressed it within Escherichia coli as a soluble holoprotein. UV-visible and resonance Raman spectra of gCYTb5-I resemble those of microsomal cytochrome b(5), and homology modelling supports a structure in which a pair of invariant histidine residues act as axial ligands to the haem iron. The reduction potential of gCYTb5-I is -165 mV vs. SHE and is relatively low compared to most values (-110 to +80 mV) for this class of protein. The amino- and carboxy-terminal sequences that flank the central haem-binding core of the Giardia cytochromes are highly charged and differ from those of other family members. A core gCYTb5-I variant lacking these flanking sequences was also able to bind haem. The presence of one actual and two probable functional cytochromes b(5) in Giardia is evidence of uncharacterized cytochrome-mediated metabolic processes within this medically important protist.

Inactivation of the heme degrading enzyme IsdI by an active site substitution that diminishes heme ruffling.

IsdG and IsdI are paralogous heme degrading enzymes from the bacterium Staphylococcus aureus. Heme bound by these enzymes is extensively ruffled such that the meso-carbons at the sites of oxidation are distorted toward bound oxygen. In contrast, the canonical heme oxygenase family degrades heme that is bound with minimal distortion. Trp-66 is a conserved heme pocket residue in IsdI implicated in heme ruffling. IsdI variants with Trp-66 replaced with residues having less bulky aromatic and alkyl side chains were characterized with respect to catalytic activity, heme ruffling, and electrochemical properties. The heme degradation activity of the W66Y and W66F variants was approximately half that of the wild-type enzyme, whereas the W66L and W66A variants were inactive. A crystal structure and NMR spectroscopic analysis of the W66Y variant reveals that heme binds to this enzyme with less heme ruffling than observed for wild-type IsdI. The reduction potential of this variant (-96 ± 7 mV versus standard hydrogen electrode) is similar to that of wild-type IsdI (-89 ± 7 mV), so we attribute the diminished activity of this variant to the diminished heme ruffling observed for heme bound to this enzyme and conclude that Trp-66 is required for optimal catalytic activity.

Kinematics of the unrestrained vehicle occupants in side-impact crashes.

A test series involving direct right-side impact of a moving wall on unsupported, unrestrained cadavers with no arms was undertaken to better understand human kinematics and injury mechanisms during side impact at realistic speeds. The tests conducted provided a unique opportunity for a detailed analysis of the kinematics resulting from side impact. Specifically, this study evaluated the 3-dimensional (3D) kinematics of 3 unrestrained male cadavers subjected to lateral impact by a multi-element load wall carried by a pneumatically propelled rail-mounted sled reproducing a conceptual side crash impact. Three translations and 3 rotations characterize the movement of a solid body in the space, the 6 degrees of freedom (6DoF) kinematics of 15 bone segments were obtained from the 3D marker motions and computed tomography (CT)-defined relationships between the maker array mounts and the bones. The moving wall initially made contact with the lateral aspect of the pelvis, which initiated lateral motion of the spinal segments beginning with the pelvis and moving sequentially up through the lumbar spine to the thorax. Analyzing the 6DoF motions kinematics of the ribs and sternum followed right shoulder contact with the wall. Overall thoracic motion was assessed by combining the thoracic bone segments as a single rigid body. The kinematic data presented in this research provides quantified subject responses and boundary condition interactions that are currently unavailable for lateral impact.

Hydrogen bond strength modulates the mechanical strength of ferric-thiolate bonds in rubredoxin.

It has long been recognized that hydrogen bonds formed by protein backbone amides with cysteinyl S(γ) atoms play important roles in modulating the functional and structural properties of the iron-sulfur centers in proteins. Here we use single molecule atomic force microscopy, cyclic voltammetry, and protein engineering techniques to investigate directly how the strength of N-H···S(γ) hydrogen bonds in the secondary coordination sphere affects the mechanical stability of Fe(III)-thiolate bonds of rubredoxin. Our results show that the mechanical stability of Fe(III)-thiolate bonds in rubredoxin correlates with the strength of N-H···S(γ) hydrogen bonds as reflected by the midpoint reduction potential, providing direct evidence that N-H···S(γ) hydrogen bonds play important roles in modulating the mechanical and kinetic properties of the Fe(III)-thiolate bonds of iron-sulfur proteins and corroborating the important roles of the protein environment in tuning the properties of metal-thiolate bonds.

Electronic properties of the highly ruffled heme bound to the heme degrading enzyme IsdI.

IsdI, a heme-degrading protein from Staphylococcus aureus, binds heme in a manner that distorts the normally planar heme prosthetic group to an extent greater than that observed so far for any other heme-binding protein. To understand better the relationship between this distinct structural characteristic and the functional properties of IsdI, spectroscopic, electrochemical, and crystallographic results are reported that provide evidence that this heme ruffling is essential to the catalytic activity of the protein and eliminates the need for the water cluster in the distal heme pocket that is essential for the activity of classical heme oxygenases. The lack of heme orientational disorder in (1)H-NMR spectra of the protein argues that the catalytic formation of ß- and δ-biliverdin in nearly equal yield results from the ability of the protein to attack opposite sides of the heme ring rather than from binding of the heme substrate in two alternative orientations.

N-terminal short fragment of TUP1 confers resistance to 5-bromodeoxyuridine in the yeast Saccharomyces cerevisiae.

Small molecules that exhibit biological activity have contributed to the understanding of the molecular mechanisms of various biological phenomena. 5-Bromodeoxyuridine (BrdU) is a thymidine analogue that modulates various biological phenomena such as cellular differentiation and cellular senescence in cultured mammalian cells. Although BrdU is thought to function through changing chromatin structure and gene expression, its precise molecular mechanisms are not understood. To study the molecular mechanism for the action of BrdU, we have employed the yeast Saccharomyces cerevisiae as a model system, and screened multi-copy suppressor genes that confer resistance to BrdU. Our genetic screen has revealed that expression of the N-terminal short fragment of TUP1, and also disruption of HDA1 or HOS1, histone deacetylases that interact with TUP1, conferred resistance to BrdU. These results suggest the implication of the chromatin proteins in the function of BrdU, and would provide novel clues to answer the old question of how BrdU modulates various biological phenomena.

BAG3 directly interacts with mutated alphaB-crystallin to suppress its aggregation and toxicity.

A homozygous disruption or genetic mutation of the bag3 gene causes progressive myofibrillar myopathy in mouse and human skeletal and cardiac muscle disorder while mutations in the small heat shock protein αB-crystallin gene (CRYAB) are reported to be responsible for myofibrillar myopathy. Here, we demonstrate that BAG3 directly binds to wild-type αB-crystallin and the αB-crystallin mutant R120G, via the intermediate domain of BAG3. Peptides that inhibit this interaction in an in vitro binding assay indicate that two conserved Ile-Pro-Val regions of BAG3 are involved in the interaction with αB-crystallin, which is similar to results showing BAG3 binding to HspB8 and HspB6. BAG3 overexpression increased αB-crystallin R120G solubility and inhibited its intracellular aggregation in HEK293 cells. BAG3 suppressed cell death induced by αB-crystallin R120G overexpression in differentiating C2C12 mouse myoblast cells. Our findings indicate a novel function for BAG3 in inhibiting protein aggregation caused by the genetic mutation of CRYAB responsible for human myofibrillar myopathy.