The Ser176 of T4 endonuclease IV is crucial for the restricted and polarized dC-specific cleavage of single-stranded DNA implicated in restriction of dC-containing DNA in host Escherichia coli

Endonuclease (Endo) IV encoded by denB of bacteriophage T4 is an enzyme that cleaves single-stranded (ss) DNA in a dC-specific manner. Also the growth of dC-substituted T4 phage and host Escherichia coli cells is inhibited by denB expression presumably because of the inhibitory effect on replication of dC-containing DNA. Recently, we have demonstrated that an efficient cleavage by Endo IV occurs exclusively at the 5′-proximal dC (dC1) within a hexameric or an extended sequence consisting of dC residues at the 5′-proximal and the 3′-proximal positions (dCs tract), in which a third dC residue within the tract affects the polarized cleavage and cleavage rate. Here we isolate and characterize two denB mutants, denB(W88R) and denB(S176N). Both mutant alleles have lost the detrimental effect on the host cell. Endo IV(W88R) shows no enzymatic activity (<0.4% of that of wild-type Endo IV). On the other hand, Endo IV(S176N) retains cleavage activity (17.5% of that of wild-type Endo IV), but has lost the polarized and restricted cleavage of a dCs tract, indicating that the Ser176 residue of Endo IV is implicated in the polarized cleavage of a dCs tract which brings about a detrimental effect on the replication of dC-containing DNA

Iowa Mutant Apolipoprotein A-I (ApoA-IIowa) Fibrils Target Lysosomes

The single amino acid mutation G26R in human apolipoprotein A-I (apoA-IIowa) is the first mutation that was associated with familial AApoA1 amyloidosis. The N-terminal fragments (amino acid residues 1–83) of apoA-I containing this mutation deposit as amyloid fibrils in patients’ tissues and organs, but the mechanisms of cellular degradation and cytotoxicity have not yet been clarified. In this study, we demonstrated degradation of apoA-IIowa fibrils via the autophagy-lysosomal pathway in human embryonic kidney 293 cells. ApoA-IIowa fibrils induced an increase in lysosomal pH and the cytosolic release of the toxic lysosomal protease cathepsin B. The mitochondrial dysfunction caused by apoA-IIowa fibrils depended on cathepsin B and was ameliorated by increasing the degradation of apoA-IIowa fibrils. Thus, although apoA-IIowa fibril transport to lysosomes and fibril degradation in lysosomes may have occurred, the presence of an excess number of apoA-IIowa fibrils, more than the lysosomes could degrade, may be detrimental to cells. Our results thus provide evidence that the target of apoA-IIowa fibrils is lysosomes, and we thereby gained a novel insight into the mechanism of AApoA1 amyloidosis

The Selective Arterial Calcium Injection Test is a Valid Diagnostic Method for Invisible Gastrinoma with Duodenal Ulcer Stenosis : A Case Report

The localization and diagnosis of microgastrinomas in a patient with multiple endocrine neoplasia type 1 is difficult preoperatively. The selective arterial calcium injection (SACI) test is a valid diagnostic method for the preoperative diagnosis of these invisible microgastrinomas. We report a rare case of multiple invisible duodenal microgastrinomas with severe duodenal stenosis diagnosed preoperatively by using the SACI test. A 50-year-old man was admitted to our hospital with recurrent duodenal ulcers. His serum gastrin level was elevated to 730 pg/ml. It was impossible for gastrointestinal endoscopy to pass through to visualize the inferior part of the duodenum, because recurrent duodenal ulcers had resulted in severe duodenal stenosis. The duodenal stenosis also prevented additional endoscopic examinations such as endoscopic ultrasonography. Computed tomography did not show any tumors in the duodenum and pancreas. The SACI test provided the evidence for a gastrinoma in the vascular territory of the inferior pancreatic-duodenal artery. We diagnosed a gastrinoma in the peri- ampullary lesion, so we performed Subtotal Stomach-Preserving Pancreatico- duodenectomy with regional lymphadenectomy. Histopathological findings showed multiple duodenal gastrinomas with lymph node metastasis and nonfunctioning pancreatic neuroendocrine tumors. Twenty months after surgery, the patient is alive with no evidence of recurrence and a normal gastrin level. In conclusion, the SACI test can enhance the accuracy of preoperative localization and diagnosis of invisible microgastrinomas, especially in the setting of severe duodenal stenosis

Study on the reusability of fluorescent nuclear track detectors using optical bleaching

Fluorescent nuclear track detectors (FNTDs) based on Al${_2}$O${_3}$:C,Mg crystals are luminescent detectors that can be used for dosimetry and detection of charged particles and neutrons. These detectors can be utilised for imaging applications where a reasonably high track density, approximately of the order of 1 $\times$ $10^4$ tracks in an area of 100 $\times$ 100 $\mu$m$^2$, is required. To investigate the reusability of FNTDs for imaging applications, we present an approach to perform optical bleaching under the required track density conditions. The reusability was assessed through seven irradiation-bleaching cycles. For the irradiation, the studied FNTD was exposed to alpha-particles from an $^{241}$Am radioactive source. The optical bleaching was performed by means of ultraviolet laser light with a wavelength of 355 nm. Three dedicated regions on a single FNTD with different accumulated track densities and bleaching conditions were investigated. After every irradiation-bleaching cycle, signal-to-noise ratio was calculated to evaluate FNTD performance. It is concluded that FNTDs can be reused at least seven times for applications where accumulation of a high track density is required

Trajectories of 3T3 fibroblasts cultured on A1-A3 and B1-B3 gels.

<p>Time-lapse microscopy was performed for 30 hr and images were acquired at 15 min intervals. The starting position of each cell trajectory was mapped to the origin of the graph. A band of single unit including the origin is highlighted. n=40 cells.</p

Long-range cell migration on gels with saw-like asymmetric elasticity patterns and perpendicular soft lanes (PSLs).

<p>The conditions used to prepare the gels are described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078067#pone-0078067-t001" target="_blank">Table 1</a>. (a) Phase contrast microscopy image of the gel. Sale bar: 100 µm. (b) Distribution of Young’s modulus in a single unit of the gel. Unit size: 90 µm. Elasticity peak: ca. 500 kPa. (c) Trajectories of 3T3 fibroblasts cultured on gels with PSLs. Cells were observed by time-lapse microscopy for 24 hr and images were acquired at 15 min intervals. (d) Time-course of ensemble averaged trajectories with standard deviation in the presence (red) and absence (blue) of PSLs. n=30 cells.</p

Distribution of Young’s moduli in gels with different unit sizes and peak elasticities.

<p>The width of each unit in series A and B was 100 µm and 120 µm, respectively. Series 1, 2, and 3 had a peak elasticity of ca. 100 kPa, 300 kPa, and 500 kPa, respectively. The ratio of ascending:descending elasticity gradients in each unit was 1:2, i.e., the X-position of the peak of elasticity was located at ca. 30 µm and 40 µm in series A and B, respectively. The conditions used to prepare each of the gels are described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078067#pone-0078067-t001" target="_blank">Table 1</a>.</p