Translation of nonSTOP mRNA is repressed post-initiation in mammalian cells

We investigated the fate of aberrant mRNAs lacking in-frame termination codons (called nonSTOP mRNA) in mammalian cells. We found that translation of nonSTOP mRNA was considerably repressed although a corresponding reduction of mRNA was not observed. The repression appears to be post-initiation since (i) repressed nonSTOP mRNAs were associated with polysomes, (ii) translation of IRES-initiated and uncapped nonSTOP mRNA were repressed, and (iii) protein production from nonSTOP mRNA associating with polysomes was significantly reduced when used to program an in vitro run-off translation assay. NonSTOP mRNAs distributed into lighter polysome fractions compared to control mRNAs encoding a stop codon, and a significant amount of heterogeneous polypeptides were produced during in vitro translation of nonSTOP RNAs, suggesting premature termination of ribosomes translating nonSTOP mRNA. Moreover, a run-off translation assay using hippuristanol and RNAse protection assays suggested the presence of a ribosome stalled at the 3' end of nonSTOP mRNAs. Taken together, these data indicate that ribosome stalling at the 3' end of nonSTOP mRNAs can block translation by preventing upstream translation events.     

Enrichment of longevity phenotype in mtDNA haplogroups D4b2b, D4a, and D5 in the Japanese population

We report new results from the re-analysis of 672 complete mitochondrial (mtDNA) genomes of unrelated Japanese individuals stratified into seven equal sized groups by the phenotypes: diabetic patients, diabetic patients with severe angiopathy, healthy non-obese young males, obese young males, patients with Alzheimer’s disease, patients with Parkinson’s disease and centenarians. Each phenotype had 96 samples over 27 known haplogroups: A, B4a, B4b, B4c, B*, B5, D*, F1, F2, M*, M7a, M7b, M8, M9, D4a, D4b1, D4b2, D4d, D4e, D4g, D4h, D5, G, Z, M*, N9a, and N9b. A t-test comparing the fraction of samples in a haplogroup to healthy young males showed a significant enrichment of haplogroups D4a, D5, and D4b2 in centenarians. The D4b2 enrichment was limited to a subgroup of 40 of 61 samples which had the synonymous mutation 9296C > T. We identified this cluster as a distinct haplogroup and labeled it as D4b2b. Using an exhaustive procedure, we constructed the complete list of “mutation patterns” for centenarians and showed that the most significant patterns were in D4a, D5, and D4b2b. We argue that if a selection for longevity appeared only once, it was probably an autosomal event which could be dated to after the appearance of the D mega-group but before the coalescent time of D4a, D5, and D4b2b. Using a simple procedure, we estimated that this event occurred 24.4 ± 0.9 kYBP. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Gabriela Alexe and Noriyuki Fuku are joint first authors.     

Genetic and molecular characterization of pathogenic isolates of Pyricularia grisea from wheat (Triticum aestivum Lam.) and triticale (x Triticosecale Wittmack) in the state of Paraná, Brazil

Isolates of Pyricularia grisea from wheat (Triticum aestivum Lam.) and triticale (x Triticosecale Wittmack) spikes with blast symptoms were analyzed by classical (VCG) and molecular (RAPD) techniques. P. grisea mutants, unable to use sodium nitrate (nit) as nitrogen source, were obtained with potassium chlorate. For vegetative compatibility (VCG) tests, genetically complementary nit mutant pairs were inoculated in a medium with sodium nitrate as a single nitrogen source. P. grisea isolates were divided into two vegetative compatibility groups and two RAPD groups. Since vegetative compatible strains may mutually exchange genetic and cytoplasmatic material, the contribution of the parasexual cycle in the genetic variability of Brazilian P. grisea isolates is discussed.     

The genome size evolution of medaka (Oryzias latipes) and fugu (Takifugu rubripes)

Evolution of the genome size in eukaryotes is often affected by changes in the noncoding sequences, for which insertions and deletions (indels) of small nucleotide sequences and amplification of repetitive elements are considered responsible. In this study, we compared the genomic DNA sequences of two kinds of fish, medaka (Oryzias latipes) and fugu (Takifugu rubripes), which show two-fold difference in the genome size (800 Mb vs. 400 Mb). We selected a contiguous DNA sequence of 790 kb from the medaka chromosome LG22 (linkage group 22), and made a precise comparison with the sequence (387 kb) of the corresponding region of Takifugu. The sequence of 178 kb in total was aligned common between two fishes, and the remaining sequences (612 kb for medaka and 209 kb for fugu) were found abundant in various repetitive elements including many types of unclassified low copy repeats, all of which accounted for more than a half (54%) of the genome size difference. Furthermore, we identified a significant difference in the length ratio of the unaligned sequences that locate between the aligned sequences (USBAS), particularly after eliminating known repetitive elements. These USBAS with no repetitive elements (USBAS-nr) located within the intron and intergenic region. These results strongly indicated that amplification of repetitive elements and compilation of indels are major driving forces to facilitate changes in the genome size.     

Mitochondrial genomes of two luminous beetles, Rhagophthalmus lufengensis and R. ohbai (Arthropoda, Insecta, Coleoptera)

We determined the mitochondrial DNA (mtDNA) sequences of two luminous beetles (Arthropoda, Insecta, Coleoptera), Rhagophthalmus lufengensis from Yunnan, China and Rhagophthalmus ohbai from Yaeyama Island, Japan. We identified all the 37 mtDNA genes of R. lufengensis (15,982 bp) and the 34 genes of R. ohbai (15,704 bp). R. lufengensis and R. ohbai genomes have higher A + T contents than other coleopteran genomes although the gene arrangements are similar. Interestingly, in a study of the evolutionary relationship among R. lufengensis, R. ohbai and the firefly Pyrocoelia rufa, the phylogenetic tree inferred from lrRNA genes from mitochondrial genomes indicates a biogeographic relationship among the bioluminescent insects in East Asia and the phylogenetic tree inferred from luciferase-related genes from nuclear genomes shows an appropriate relationship among coleopterans, reflecting the evolutionary origin of bioluminescence. Thus, the mtDNAs of luminescent beetles can provide an insight into their evolutionary origin and biogeography.     

Identification of three novel proteins (SGSM1, 2, 3) which modulate small G protein (RAP and RAB)-mediated signaling pathway

We report a novel protein family consisting of three members, each of which contains RUN and TBC motifs and appears to be associated with small G protein-mediated signal transduction pathway. We named these proteins as small G protein signaling modulators (SGSM1/2/3). Northern blot analysis revealed that human SGSM2/3 are expressed ubiquitously in various tissues, whereas SGSM1 is expressed mainly in brain, heart, and testis. Mouse possessed the same protein family genes, and the in situ hybridization and immunohistochemical staining of tissue sections revealed that mouse Sgsm1/2/3 are expressed in the neurons of central nervous system, indicating the strong association of Sgsm family with neuronal function. Furthermore, endogenous Sgsm1 protein was localized in the trans-Golgi network of mouse Neuro2a cells by immunofluorescence microscopy. Expression of various cDNA constructs followed by immunoprecipitation assay revealed that human SGSM1/2/3 proteins are coprecipitated with RAP and RAB subfamily members of the small G protein superfamily. Based on these results, we postulated that the SGSM family members function as modulators of the small G protein RAP and RAB-mediated neuronal signal transduction and vesicular transportation pathways.     

Unprecedented intraspecific diversity of the MHC class I region of a teleost medaka, Oryzias latipes

The major histocompatibility complex (MHC) is present at a single chromosomal locus of all jawed vertebrate analyzed so far, from sharks to mammals, except for teleosts whose orthologs of the mammalian MHC-encoded genes are dispersed at several chromosomal loci. Even in teleosts, several class IA genes and those genes directly involved in class I antigen presentation preserve their linkage, defining the teleost MHC class I region. We determined the complete nucleotide sequence of the MHC class I region of the inbred HNI strain of medaka, Oryzias latipes (northern Japan population-derived), from four overlapping bacterial artificial chromosome (BAC) clones spanning 540,982 bp, and compared it with the published sequence of the corresponding region of the inbred Hd-rR strain of medaka (425,935 bp, southern Japan population-derived) as the first extensive study of intraspecies polymorphisms of the ectotherm MHC regions. A segment of about 100 kb in the middle of the compared sequences encompassing two class Ia genes and two immunoproteasome subunit genes, PSMB8 and PSMB10, was so divergent between these two inbred strains that a reliable sequence alignment could not be made. The rest of the compared region (about 320 kb) showed a fair correspondence, and an approximately 96% nucleotide identity was observed upon gap-free segmental alignment. These results indicate that the medaka MHC class I region contains an ∼100-kb polymorphic core, which is most probably evolving adaptively by accumulation of point mutations and extensive genetic rearrangements such as insertions, deletions and duplications. The nucleotide sequence data of HNI MHC class I region reported in this paper have been submitted to the DDBJ/EMBL/GenBank and were assigned the accession number AB183488.     

An isozyme of earthworm serine proteases acts on hydrolysis of triacylglycerol

An enzyme catalyzing the hydrolysis of triacylglycerol was purified from an earthworm. The N-terminal amino acid sequence and the catalytic function of the purified enzyme were identical to those of Isozyme C, an isozyme of the earthworm-serine proteases. No other lipase proteins were found in the earthworm cells. The isozyme might act on the hydrolysis of triacylglycerol as well as the protein decomposition.