The physiological significance of the xanthurenic acid-insulin comples.

The xanthurenic acid-insulin complex was found to have similar immunological properties to native Zn-insulin. This complex showed less hormonal activity on glucose metabolism in adipose tissue than native An-insulin, but its activity was increased by addition of Zn2+ ions.     

Ribonucleic acid polymerase of germinating Bacillus cereus T.

It appears that a de novo synthesis of the deoxyribonucleic acid-dependent ribonucleic acid-polymerase in Bacillus cereus T takes place fairly late in outgrowth, at the onset of the vegetative cycle. Therefore, the ribonucleic acid-polymerase used by germinating spores is the one carried on from sporulating cells. However, the sporal enzyme is less soluble that the vegetative one, and its "core" is bound to two extra peptides. This complexing to other molecules could play a role in the regulation of gene expression during germination.     

Molecular Sizes of the Catalytic Units of Oxygen Evolution and the Reaction Center in Photosystem II of Spinach Thylakoids

Target analysis of the PS II reaction in spinach thylakoidsshowed that the respective molecular masses of the catalyticunits for oxygen evolution and the reaction center are about120 kDa and 250 kDa based on a kinetic separation of the tworeaction rates. The size of the oxygen-evolving enzyme agreedwith that determined for the PS II preparation from a thermophiliccyanobacterium by the same means [Nugent and Atkinson (1984)FEBS Lett. 170: 89]. Single hit-inactivation of oxygen evolutionand the PS II reaction center units indicates that each functionis driven by a structurally assembled unit. (Received August 6, 1984; Accepted December 17, 1984)     

T-maze Forced Alternation and Left-right Discrimination Tasks for Assessing Working and Reference Memory in Mice

Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in rodents. In our laboratory, we evaluated the two types of memory in more than 30 strains of genetically engineered mice using the automated version of this apparatus. Here, we present the modified T-maze apparatus operated by a computer with a video-tracking system and our protocols in a movie format. The T-maze apparatus consists of runways partitioned off by sliding doors that can automatically open downward, each with a start box, a T-shaped alley, two boxes with automatic pellet dispensers at one side of the box, and two L-shaped alleys. Each L-shaped alley is connected to the start box so that mice can return to the start box, which excludes the effects of experimenter handling on mouse behavior. This apparatus also has an advantage that in vivo microdialysis, in vivo electrophysiology, and optogenetics techniques can be performed during T-maze performance because the doors are designed to go down into the floor. In this movie article, we describe T-maze tasks using the automated apparatus and the T-maze performance of α-CaMKII+/- mice, which are reported to show working memory deficits in the eight-arm radial maze task. Our data indicated that α-CaMKII+/- mice showed a working memory deficit, but no impairment of reference memory, and are consistent with previous findings using the eight-arm radial maze task, which supports the validity of our protocol. In addition, our data indicate that mutants tended to exhibit reversal learning deficits, suggesting that α-CaMKII deficiency causes reduced behavioral flexibility. Thus, the T-maze test using the modified automatic apparatus is useful for assessing working and reference memory and behavioral flexibility in mice.     

Two ANGUSTIFOLIA genes regulate gametophore and sporophyte development in Physcomitrella patens

In Arabidopsis thaliana the ANGUSTIFOLIA (AN) gene regulates the width of leaves by controlling the diffuse growth of leaf cells in the medio‐lateral direction. In the genome of the moss Physcomitrella patens, we found two normal ANs (PpAN1‐1 and 1‐2). Both PpAN1 genes complemented the A. thaliana an‐1 mutant phenotypes. An analysis of spatiotemporal promoter activity of each PpAN1 gene, using transgenic lines that contained each PpAN1‐promoter– uidA (GUS) gene, showed that both promoters are mainly active in the stems of haploid gametophores and in the middle to basal region of the young sporophyte that develops into the seta and foot. Analyses of the knockout lines for PpAN1‐1 and PpAN1‐2 genes suggested that these genes have partially redundant functions and regulate gametophore height by controlling diffuse cell growth in gametophore stems. In addition, the seta and foot were shorter and thicker in diploid sporophytes, suggesting that cell elongation was reduced in the longitudinal direction, whereas no defects were detected in tip‐growing protonemata. These results indicate that both PpAN1 genes in P. patens function in diffuse growth of the haploid and diploid generations but not in tip growth. To visualize microtubule distribution in gametophore cells of P. patens, transformed lines expressing P. patens α‐tubulin fused to sGFP were generated. Contrary to expectations, the orientation of microtubules in the tips of gametophores in the PpAN1‐1/1‐2 double‐knockout lines was unchanged. The relationships among diffuse cell growth, cortical microtubules and AN proteins are discussed.     

Loss of aPKCλ in Differentiated Neurons Disrupts the Polarity Complex but Does Not Induce Obvious Neuronal Loss or Disorientation in Mouse Brains

Cell polarity plays a critical role in neuronal differentiation during development of the central nervous system (CNS). Recent studies have established the significance of atypical protein kinase C (aPKC) and its interacting partners, which include PAR-3, PAR-6 and Lgl, in regulating cell polarization during neuronal differentiation. However, their roles in neuronal maintenance after CNS development remain unclear. Here we performed conditional deletion of aPKCλ, a major aPKC isoform in the brain, in differentiated neurons of mice by camk2a-cre or synapsinI-cre mediated gene targeting. We found significant reduction of aPKCλ and total aPKCs in the adult mouse brains. The aPKCλ deletion also reduced PAR-6β, possibly by its destabilization, whereas expression of other related proteins such as PAR-3 and Lgl-1 was unaffected. Biochemical analyses suggested that a significant fraction of aPKCλ formed a protein complex with PAR-6β and Lgl-1 in the brain lysates, which was disrupted by the aPKCλ deletion. Notably, the aPKCλ deletion mice did not show apparent cell loss/degeneration in the brain. In addition, neuronal orientation/distribution seemed to be unaffected. Thus, despite the polarity complex disruption, neuronal deletion of aPKCλ does not induce obvious cell loss or disorientation in mouse brains after cell differentiation.     

Evidence of a landlocked reproducing population of the marine pejerrey Odontesthes argentinensis (Actinopterygii; Atherinopsidae)

In South America, the order Atheriniformes includes the monophyletic genus Odontesthes with 20 species that inhabit freshwater, estuarine and coastal environments. Pejerrey Odontesthes argentinensis is widely distributed in coastal and estuarine areas of the Atlantic Ocean and is known to foray into estuaries of river systems, particularly in conditions of elevated salinity. However, to our knowledge, a landlocked self-sustaining population has never been recorded. In this study, we examined the pejerrey population of Salada de Pedro Luro Lake (south-east of Buenos Aires Province, Argentina) to clarify its taxonomic identity. An integrative taxonomic analysis based on traditional meristic, landmark-based morphometrics and genetic techniques suggests that the Salada de Pedro Luro pejerrey population represents a novel case of physiological and morphological adaptation of a marine pejerrey species to a landlocked environment and emphasises the environmental plasticity of this group of fishes.     

Molecular characterization of rotaviruses obtained from patients with rotavirus-associated encephalitis/encephalopathy

Group A rotavirus (RVA) rarely causes severe complications such as encephalitis/encephalopathy. However, the pathophysiology of this specific complication remains unclear. Next-generation sequence analysis was used to compare the entire genome sequences of RVAs detected in patients with encephalitis/encephalopathy and gastroenteritis. This study enrolled eight patients with RVA encephalitis/encephalopathy and 10 with RVA gastroenteritis who were treated between February 2013 and July 2014. Viral RNAs were extracted from patients' stool, and whole-genome sequencing analysis was carried out to identify the specific gene mutations in RVA obtained from patients with severe neurological complications. Among the eight encephalitis/encephalopathy cases, six strains were DS-1-like G1P[8] and the remaining two were Wa-like G1P[8] (G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1). Meanwhile, eight of the 10 viruses detected in rotavirus gastroenteritis patients were DS-1-like G1P[8], and the remaining two were Wa-like G1P[8]. These strains were further characterized by conducting phylogenetic analysis. No specific clustering was demonstrated in RVAs detected from encephalitis/encephalopathy patients. Although the DS-1-like G1P[8] strain was predominant in both groups, no specific molecular characteristics were detected in RVAs from patients with severe central nervous system complications.     

Visualization of multivalent histone modification in a single cell reveals highly concerted epigenetic changes on differentiation of embryonic stem cells

Combinations of histone modifications have significant biological roles, such as maintenance of pluripotency and cancer development, but cannot be analyzed at the single cell level. Here, we visualized a combination of histone modifications by applying the in situ proximity ligation assay, which detects two proteins in close vicinity (∼30 nm). The specificity of the method [designated as imaging of a combination of histone modifications (iChmo)] was confirmed by positive signals from H3K4me3/acetylated H3K9, H3K4me3/RNA polymerase II and H3K9me3/H4K20me3, and negative signals from H3K4me3/H3K9me3. Bivalent modification was clearly visualized by iChmo in wild-type embryonic stem cells (ESCs) known to have it, whereas rarely in Suz12 knockout ESCs and mouse embryonic fibroblasts known to have little of it. iChmo was applied to analysis of epigenetic and phenotypic changes of heterogeneous cell population, namely, ESCs at an early stage of differentiation, and this revealed that the bivalent modification disappeared in a highly concerted manner, whereas phenotypic differentiation proceeded with large variations among cells. Also, using this method, we were able to visualize a combination of repressive histone marks in tissue samples. The application of iChmo to samples with heterogeneous cell population and tissue samples is expected to clarify unknown biological and pathological significance of various combinations of epigenetic modifications.