Lats2 is an essential mitotic regulator required for the coordination of cell division

Tumor suppressor Lats2 is a member of the conserved Dbf2 kinase family. It localizes to the centrosome and has been implicated in regulation of the cell cycle and apoptosis. However, the in vivo function of this kinase remains unclear. Here, we show that complete disruption of the gene encoding Lats2 in mice causes developmental defects in the nervous system and embryonic lethality. Furthermore, mutant cells derived from total LATS2-knock-out embryos exhibit mitotic defects including centrosome fragmentation and cytokinesis defects, followed by nuclear enlargement and multinucleation. We show that the Mob1 family, a regulator of mitotic exit, associates with Lats2 to induce its activation. We also show that the complete LATS2-knock-out cells exhibit an acceleration of exit from mitosis and marked down-regulation of critical mitotic regulators. These results suggest that Lats2 plays an essential mitotic role in coordinating accurate cytokinesis completion, governing the stabilization of other mitotic regulators.     

Acetylation of NDPK-D Regulates Its Subcellular Localization and Cell Survival

Nucleoside diphosphate kinases (NDPK) are ubiquitous enzymes that catalyze the reversible phosphotransfer of γ-phosphates between di- and triphosphonucleosides. NDPK-D (Nm23-H4) is the only member of the NDPK family with a mitochondrial targeting sequence. Despite the high expression of NDPK-D in the developing central nervous system, its function remains to be determined. In this study, we show that NDPK-D knockdown induces apoptosis in neuroblastoma cells as well as in mouse cortex, suggesting that NDPK-D is required for neuronal survival. We identified NDPK-D as a binding partner of NAD⁺-dependent histone deacetylase, SIRT1, by yeast two-hybrid screening. NDPK-D co-localized with SIRT1, and the association of these molecules was confirmed by co-immunoprecipitation. Inhibition of SIRT1 increases the acetylation of NDPK-D. Overexpression of NDPK-D along with SIRT1, or mutation in the acetylated lysine residues in NDPK-D, increases its nuclear accumulation. Furthermore, the NDPK-D acetylation-mimic mutant increased apoptosis in N1E-115 cells. Our data demonstrate that acetylation regulates the shuttling of NDPK-D between nucleus and cytoplasm, and increased acetylation of NDPK-D causes apoptosis.     

Purification, characterization and sequence analyses of the extracellular giant hemoglobin from Oligobrachia mashikoi

We purified an extracellular hemoglobin with the molecular mass of ca. 440 kDa from the whole homogenates of Oligobrachia mashikoi (phylum Pogonophora) by a one-step gel-filtration. The preparation was pure to be crystallized. The P50 values of the hemoglobin and the fresh blood prepared from O. mashikoi were about 0.82 Torr and 0.9 Torr, respectively, which were much lower than the P50 value of human hemoglobin. However, the n values of the hemoglobin and the blood were about 1.2 and 1.1, respectively.Using the improved tricine SDS-PAGE, we could separate O. mashikoi hemoglobin into four kinds of the globin chains, A1, A2, B1 and B2, and succeeded for the first time in cloning and sequencing of the complete cDNA encoding B1 globin gene, in addition to A1, A2 and B2 globin genes in full length. We found that all globin genes have the extracellular signal sequences in each molecule and the distal His of the B1 globin chain is replaced to Gln. Finally, we constructed phylogenetic trees of the hemoglobins from Pogonophora, Vestimentifera and Annelida.     

Differences in gene expression profile among SH-SY5Y neuroblastoma subclones with different neurite outgrowth responses to nerve growth factor

Nerve growth factor (NGF) plays a key role in the differentiation of neurons. In this study, we established three NGF-induced neurite-positive (NIN+) subclones that showed high responsiveness to NGF-induced neurite outgrowth and three NGF-induced neurite-negative (NIN-) subclones that abolished NGF-induced neurite outgrowth from parental SH-SY5Y cells, and analyzed differences in the NGF signaling cascade. The NIN+ subclones showed enhanced responsiveness to FK506-mediated neurite outgrowth as well. To clarify the mechanism behind the high frequency of NGF-induced neurite outgrowth, we investigated differences in NGF signaling cascade among subclones. Expression levels of the NGF receptor TrkA, and NGF-induced increases in mRNAs for the immediate-early genes (IEGs) c-fos and NGF inducible (NGFI) genes NGFI-A, NGFI-B and NGFI-C, were identical among subclones. Microarray analysis revealed that the NIN+ cell line showed a very different gene expression profile to the NIN- cell line, particularly in terms of axonal vesicle-related genes and growth cone guidance-related genes. Thus, the difference in NGF signaling cascade between the NIN+ and NIN- cell lines was demonstrated by the difference in gene expression profile. These differentially expressed genes might play a key role in neurite outgrowth of SH-SY5Y cells in a region downstream from the site of induction of IEGs, or in a novel NGF signaling cascade.     

Unpaired Ultimobranchial Glands of the African Lungfish, Protopterus dolloi

The ultimobranchial glands of juvenile African lungfish (Protopterus dolloi) (14 individuals; total body length 25-205 mm) were immunohistochemically examined. In individuals larger than 36 mm, one ultimobranchial gland was close to the left afferent branchial arteries. The topography of the ultimobranchial gland was similar to that of salamanders and sharks, but not to teleosts. With body growth, the ultimobranchial gland was vascularized and the parenchymal cells were gradually immunostained with anti-calcitonin antibody. In all individuals examined, the ultimobranchial gland existed only on the left side of the pharynx. These observations are discussed from a phylogenetic viewpoint.     

Internodal elongation under submergence in the Amazonian wild rice species <Emphasis Type="Italic">Oryza glumaepatula</Emphasis>: the growth response is induced by hypoxia but not by ethylene

Phosphate (Pi) plays important roles in plant development and architecture. With the goal of identifying genomic regions that influence tolerance to Pi deficiency (TPDE) in hybrid rice (Oryza sativa L.), quantitative trait loci (QTL) were mapped using recombinant inbred lines (RILs) that were derived from a cross between tolerant ‘XieqingzaoB’ (XB) and susceptible ‘Zhonghui9308’. Six TPDE-related traits, including the root length, root dry weight, tillers number, shoot dry weight, total plant dry weight and root-to-shoot ratio, were evaluated for QTL analysis during both the tillering and heading stages. A correlation analysis showed that most of the traits were correlated with each other. Twenty-one additive QTL were detected and jointly explained between 10–49% of the trait variance, tending to cluster on chromosomes 4, 6, 10 and 11. Three QTL, qTPDE4 ^XB , qTPDE10 ^XB and qTPDE11.3 ^XB , were validated by the phenotypic evaluation using near isogenic lines (NILs, BC₄F₃) during the seedling stage. qTPDE4 ^XB showed the most stable tolerance against Pi deficiency. These QTL will enrich the genetic resources and accelerate hybrid rice breeding against Pi deficiency.     

Correlation among clinicopathological parameters of myocardial damage in rats treated with isoproterenol.

The correlation among clinicopathological parameters of myocardial damage was investigated in rats administered a single subcutaneous dose of isoproterenol at 0 (saline), 0.04, 0.4 and 4 mg/kg. Total lactate dehydrogenase (LDH), creatine kinase (CK), and their isoenzymes (LDH-1, LDH-2 and CK-MB), as well as troponin I and tropinin T, were measured 4 h after the administration of the drug. Troponin I was determined by a chemiluminescence method using Bayer Centaur and DPC Immulyze, as well as by ELISA. Troponin T was assayed semi-quantitatively using Trop T Sensitive. A high correlation was found among LDH isoenzymes, troponin I (Centaur) and troponin T. The present result provides a baseline for interpreting changes in the different parameters of myocardial damage assayed by different methods in toxicity studies.     

What Time Periods of the Day Are Concerning for Parents of Children with Attention Deficit Hyperactivity Disorder?

Background/Aim

The questionnaire-children with difficulties (QCD) is a parent-assessed questionnaire designed to evaluate a child’s difficulties in functioning during specific time periods of the day. In this study, the QCD was applied to determine the time periods of the day that are concerning for the parents of children with attention deficit hyperactivity disorder (ADHD). The results were compared with those for a community sample.

Methods

Elementary and junior high school students with ADHD (243 boys, 55 girls) and a community sample of children (518 boys, 618 girls) were enrolled in this study. Their behaviors were assessed by the QCD, the ADHD-rating scale (ADHD-RS), and the Oppositional Defiant Behavior Inventory (ODBI). The effects of gender (boy/girl) and diagnosis (ADHD/community sample) on the total QCD score were analyzed across each school grade (elementary/junior high school). Correlation coefficients between QCD and ADHD-RS/ODBI scores were analyzed.

Results

The QCD score for the ADHD group was significantly lower than that for the community sample (P < 0.001). There were significantly strong correlations between “evening” and ADHD-RS and ODBI scores for all children with ADHD (r > 0.41, P < 0.001) and between “night” and inattention and oppositional symptoms for the girls with ADHD (r > 0.40, P < 0.001).

Conclusions

Parents reported that children with ADHD faced greater difficulties in completing basic daily activities compared with the community controls, particularly in the evening. Furthermore, these difficulties were related to the severity of ADHD symptoms. The parents’ perceptions depended on the gender, ADHD and oppositional symptoms, and the time period of the day. This study determined that children with ADHD face greater difficulties in daily functioning compared with community sample children, that these difficulties are time-dependent, and that these difficulties were particularly experienced in the evening.     

Cloning of two members of the calcitonin-family receptors from stingray, Dasyatis akajei: possible physiological roles of the calcitonin family in osmoregulation

In cartilaginous fish, two cDNAs encoding calcitonin-family receptors were isolated for the first time from the stingray brain. The open reading frame of one receptor cDNA coded a 525-amino acid protein. The amino acid identity of this receptor to human calcitonin-receptor-like receptor (CRLR) is 64.5%, frog CRLR is 64.7%, and flounder CRLR is 61.2% and this was higher than to human calcitonin receptor (CTR) (46.1%), frog CTR (54.7%), and flounder CTR (48.9%). We strongly suggested that this receptor is a ray CRLR based on phylogenetic analysis. In case of the second receptor, amino acid identity among CRLRs (human 50.5%, frog 50.7%, flounder 48.0%) and CTRs (human 43.2%, frog 49.1%, flounder 41.8%) was similar. From phylogenetic analysis of both CRLRs and CTRs, we believe that this receptor is ray CTR. The expression of ray CRLR mRNA was predominantly detected in the nervous system (brain) and vascular system (atrium, ventricle, and gill), which reflects the similar localization of CGRP in the nervous and vascular systems as mammals. It was observed that the second receptor was expressed in several tissues, namely cartilage, brain, pituitary gland, gill, atrium, ventricle, pancreas, spleen, liver, gall bladder, intestine, rectal gland, kidney, testis and ovary. This localization pattern was very similar to flounder CTR. Both receptor mRNAs were strongly expressed in the gill. This suggests that the calcitonin-family members are involved in the osmoregulation of stingray as this fish is known to be euryhaline. When a stingray was transferred to diluted seawater (20% seawater), the expression of both receptors significantly decreased in the gill. Similar results were obtained in the kidney of the stingray. Thus, our cloning and isolation of both receptors in the stingray will be helpful for elucidation of their physiological role(s) such as osmoregulation including calcium metabolism of cartilaginous fish.