Lineage-Specific Homogenization of the Polyubiquitin Gene Among Human and Great Apes

Ubiquitin is a highly conserved protein, and is encoded by a multigene family among eukaryote species. The polyubiquitin genes, UbB and UbC, comprise tandem multiple ubiquitin coding units without a spacer region or intron. We determined nucleotide sequences for the UbB and UbC of human, chimpanzee, gorilla, and orangutan. The ubiquitin repeat number of UbB was constant (3) in human and great apes, while that of UbC varied: 6 to 11 for human, 10 to 12 for chimpanzee, 8 for gorilla, and 10 for orangutan. The heterogeneity of the repeat number within closely related hominoid species suggests that a lineage-specific unequal crossover and/or gene duplication occurred. A marked homogenization of UbC occurred in gorilla with a low level of synonymous difference (p_s). The homogenization of UbC also occurred in chimpanzee and less strikingly in human. The first and last ubiquitin coding units of UbC were clustered independently between species, and less affected by homogenization during the hominoid evolution. Therefore, the homogenization of ubiquitin coding units is likely due to an unequal crossing-over inside the ubiquitin units. The lineage-specific homogenization of UbC among closely related species suggests that concerted evolution has a key role in the short-term evolution of UbC.     

Interspecific divergence, intrachromosomal recombination, and phylogenetic utility of Y-chromosomal genes in Drosophila

Reconstruction of phylogenetic relationships among recently diverged species is complicated by three general problems: segregation of polymorphisms that pre-date species divergence, gene flow during and after speciation, and intra-locus recombination. In light of these difficulties, the Y chromosome offers several important advantages over other genomic regions as a source of phylogenetic information. These advantages include the absence of recombination, rapid coalescence, and reduced opportunity for interspecific introgression due to hybrid male sterility. In this report, we test the phylogenetic utility of Y-chromosomal sequences in two groups of closely related and partially inter-fertile Drosophila species. In the D. bipectinata species complex, Y-chromosomal loci unambiguously recover the phylogeny most consistent with previous multi-locus analysis and with reproductive relationships, and show no evidence of either post-speciation gene flow or persisting ancestral polymorphisms. In the D. simulans species complex, the situation is complicated by the duplication of at least one Y-linked gene region, followed by intrachromosomal recombination between the duplicate genes that scrambles their genealogy. We suggest that Y-chromosomal sequences are a useful tool for resolving phylogenetic relationships among recently diverged species, especially in male-heterogametic organisms that conform to Haldane's rule. However, duplication of Y-linked genes may not be uncommon, and special care should be taken to distinguish between orthologous and paralogous sequences.     

新疆西部雏蝗属一新种记述（直翅目：网翅蝗科）

记述采自新疆西部网翅蝗科雏蝗属 Chorthippus 1新种,即新源雏蝗 Ch. xinyuanensis sp. nov.。新种近似于狭窝雏蝗Ch. foveatus Xia & Jin, 1982和长角雏蝗Ch. longicornis (Latreille),1804。与二者的主要区别为:前胸背板沟前区与沟后区等长;前胸背板侧隆线间最宽处为最狭处的1.7倍;前翅到达第6节背板中部(♂);中脉域约与肘脉域等宽(♂)。模式标本保存于陕西师范大学动物研究所标本室。     

新疆雏蝗属短翅亚属一新种（直翅目：网翅蝗科）

记述采自新疆地区网翅蝗科雏蝗属 Chorthippus 短翅亚属1新种,即果子沟雏蝗 Chorthippus (Altichorthippus) guozigouensis sp. nov.。新种近似于姜氏雏蝗Ch. charpini Chang, 1939 与积石山雏蝗 Ch. jishishanensis Zheng et Xie, 2000。与后两者的主要区别为:♂头侧窝长为宽的3.00倍;♂触角短,刚不到达后足股节基部;♂前翅到达第7节腹节背板后缘,其前缘脉域宽为中脉域宽0.67倍,前后肘脉合并,肘脉域消失;♂后翅到达第7节腹节背板中部;尾须顶端到达肛上板长的一半,膝部与后足胫节端部棕色,非黑色。模式标本保存于陕西师范大学动物研究所标本室。     

四川省雏蝗属一新种（直翅目：网翅蝗科）

记述采自中国四川省网翅蝗科雏蝗属Chorthippus(Altichorthippus)1新种:周氏雏蝗Chorthippus(Altichorthippus)choui sp.nov.。本新种同昌都雏蝗Chorthippus(A.)changtunensis Yin,1984近似,其区别特征为:后足股节和胫节桔红色;雄性腹部末端桔红色;前翅中脉域宽为肘脉域宽的1.2倍;本新种同西藏雏蝗Ch.(A.)tibetanus Uvarov,1935也近似,其区别特征为前胸背板侧隆线在沟前区明显,呈角状弯曲,在前横沟和后横沟之间不消失。     

Defective transfer of seminal-fluid materials during matings of semi-fertile fruitless mutants in Drosophila

In context of the semi-sterility exhibited by Drosophila males expressing certain mating-enabling fruitless (fru) mutant genotypes, we examined the transfer of seminal fluid using a transgene that encodes the Sex Peptide (SP) oligopeptide fused to Green Fluorescent Protein (GFP). We found that this fusion construct expresses SP-GFP in a valid manner within accessory glands of the male reproductive system in normal and fru-mutant males. Transfer of SP-GFP to live females was readily detectable during and after copulation. With respect to the pertinent combinations of fru mutations, we demonstrated that these abnormal genotypes cause males to transmit mating-related materials in two aberrant ways: one involving whether any seminal-fluid entities are transferred at all during a given mating; the other revealing an intriguing aspect of these fruitless effects, such that the mutations in question cause males to transfer female-affecting materials in a manner that varies among copulations. In this regard, certain mutant males that do not transfer SP nevertheless are able to transfer sperm: a fru-mated female possessing no GFP who was not fecund initially could produce progeny when seminal-fluid proteins were subsequently supplied by mating with a male that was spermless owing to the effects of a tudor mutation.     

Molecular Mechanisms for the Variation of Mitochondrial Gene Content and Gene Arrangement Among Chigger Mites of the Genus Leptotrombidium (Acari: Acariformes)

The gene content of a mitochondrial (mt) genome, i.e., 37 genes and a large noncoding region (LNR), is usually conserved in Metazoa. The arrangement of these genes and the LNR is generally conserved at low taxonomic levels but varies substantially at high levels. We report here a variation in mt gene content and gene arrangement among chigger mites of the genus Leptotrombidium. We found previously that the mt genome of Leptotrombidium pallidum has an extra gene for large-subunit rRNA (rrnL), a pseudo-gene for small-subunit rRNA (PrrnS), and three extra LNRs, additional to the 37 genes and an LNR typical of Metazoa. Further, the arrangement of mt genes of L. pallidum differs drastically from that of the hypothetical ancestor of the arthropods. To find to what extent the novel gene content and gene arrangement occurred in Leptotrombidium, we sequenced the entire or partial mt genomes of three other species, L. akamushi, L. deliense, and L. fletcheri. These three species share the arrangement of all genes with L. pallidum, except trnQ (for tRNA-glutamine). Unlike L. pallidum, however, these three species do not have extra rrnL or PrrnS and have only one extra LNR. By comparison between Leptotrombidium species and the ancestor of the arthropods, we propose that (1) the type of mt genome present in L. pallidum evolved from the type present in the other three Leptotrombidium species, and (2) three molecular mechanisms were involved in the evolution of mt gene content and gene arrangement in Leptotrombidium species. [Reviewing Editor: Dr. Martin Kreitman]     

Rapid Adaptive Evolution of the Tumor Suppressor Gene Pten in an Insect Lineage

The Pten gene was initially identified in humans as a tumor suppressor. It has since been shown to play important roles in the control of cell size, cell motility, apoptosis, and organ size, and it has also been implicated in aging. Pten is highly conserved among organisms as diverse as nematodes, insects, and vertebrates. In contrast, a phylogenetic analysis by maximum likelihood of a 133-amino acid region showed an average nonsynonymous-to-synonymous rate ratio of 10.4 for Pten in the lineage leading to parasitoid wasps of the Nasonia genus, indicating very strong positive selection. A previous study identified Pten as a potential QTL candidate gene for differences in male wing size in Nasonia. Most of the amino acid replacements that occurred in the Nasonia lineage cluster in a small region of the protein surface, suggesting that they might be involved in an interaction between Pten and another protein. The phenotypic changes due to Pten are not yet known, although it is not associated with known differences in male wing size. Introgression of Pten from one species to another does affect longevity, but a causal relationship is not established. [Reviewing Editor: Dr. Willie J. Swanson]     

‘潘那利’番茄碱性螺旋环螺旋转录因子基因的克隆与表达分析

碱性螺旋环螺旋(basic/Helix Loop Helix, bHLH)转录因子是植物最大的转录因子家族之一,其广泛参与植物逆境胁迫响应。该研究从野生‘潘那利’番茄(Solanum pennellii Correll)中成功克隆出bHLH转录因子基因SpbHLH89(Sol Genomics登录号Sopen04g001150),采用qRT PCR分析其在干旱胁迫下的表达模式,并利用异源表达初步分析其对非生物胁迫的响应。结果表明：(1)SpbHLH89编码区包含684 bp,编码227个氨基酸,具有典型的碱性螺旋环螺旋区,主要定位于细胞核中;进化树结果显示,SpbHLH89转录因子高度保守,与拟绒毛烟草NtbHLH94(Nicotiana tomentosiformis)存在高度相似性。(2)qRT PCR结果显示,SpbHLH89在‘潘那利’番茄的茎、叶和花中均有表达,其表达量受干旱胁迫诱导。(3)SDS PAGE与Western bloting结果显示,pET 30a SpbHLH89重组蛋白大小约为31 kD。(4)在盐胁迫(400 mmol/L NaCl)和干旱胁迫(600 mmol/L甘露醇)条件下,异源表达重组蛋白的E. coli BL21(DE3)重组菌生长速度提高,说明异源表达SpbHLH89转录因子基因可提高细菌对非生物胁迫的耐受性。     

Gene duplication,allelic diversity,selection processes and adaptive value of MHC class II <Emphasis Type="Italic">DRB</Emphasis> genes of the bank vole, <Emphasis Type="Italic">Clethrionomys glareolus</Emphasis>

The generation and maintenance of allelic polymorphism in genes of the major histocompatibility complex (MHC) is a central issue in evolutionary genetics. Recently, the focus has changed from ex situ to in situ populations to understand the mechanisms that determine adaptive MHC polymorphism under natural selection. Birth-and-death evolution and gene conversion events are considered to generate sequence diversity in MHC genes, which subsequently is maintained by balancing selection through parasites. The ongoing arms race between the host and parasites leads to an adaptive selection pressure upon the MHC, evident in high rates of non-synonymous vs synonymous substitution rates. We characterised the MHC class II DRB exon 2 of free living bank voles, Clethrionomys glareolus by single-strand conformation polymorphism and direct sequencing. Unlike other arvicolid species, the DRB locus of the bank vole is at least quadruplicated. No evidence for gene conversion events in the Clgl-DRB sequences was observed. We found not only high allelic polymorphism with 26 alleles in 36 individuals but also high rates of silent polymorphism. Exceptional for MHC class II genes is a purifying selection pressure upon the majority of MHC-DRB sequences. Further, we analysed the association between certain DRB alleles and the parasite burden with gastrointestinal trichostrongyle nematodes Heligmosomum mixtum and Heligmosomoides glareoli and found significant quality differences between specific alleles with respect to infection intensity. Our findings suggest a snapshot in an evolutionary process of ongoing birth-and-death evolution. One allele cluster has lost its function and is already silenced, another is loosing its adaptive value in terms of gastrointestinal nematode resistance, while a third group of alleles indicates all signs of classical functional MHC alleles.     

Heterogeneity of Chlorophyll Fluorescence over Thalli of Several Foliose Macrolichens Exposed to Adverse Environmental Factors: Interspecific Differences as Related to Thallus Hydration and High Irradiance

Spatial heterogeneity of chlorophyll (Chl) fluorescence over thalli of three foliose lichen species was studied using Chl fluorescence imaging (CFI) and slow Chl fluorescence kinetics supplemented with quenching analysis. CFI values indicated species-specific differences in location of the most physiologically active zones within fully hydrated thalli: marginal thallus parts (Hypogymnia physodes), central part and close-to-umbilicus spots (Lasallia pustulata), and irregulary-distributed zones within thallus (Umbilicaria hirsuta). During gradual desiccation of lichen thalli, decrease in Chl fluorescence parameters (F_O - minimum Chl fluorescence at point O, F_P - maximum Chl fluorescence at P point, ₂ - effective quantum yield of photochemical energy conversion in photosystem 2) was observed. Under severe desiccation (>85 % of water saturation deficit), substantial thalli parts lost their apparent physiological activity and the resting parts exhibited only a small Chl fluorescence. Distribution of these active patches was identical with the most active areas found under full hydration. Thus spatial heterogeneity of Chl fluorescence in foliose lichens may reflect location of growth zones (pseudomeristems) within thalli and adjacent newly produced biomass. When exposed to high irradiance, fully-hydrated thalli of L. pustulata and U. hirsuta showed either an increase or no change in F_O, and a decrease in F_P. Distribution of Chl fluorescence after the high irradiance treatment, however, remained the same as before the treatment. After 60 min of recovery in the dark, F_O and F_P did not recover to initial values, which may indicate that the lichen used underwent a photoinhibition. The CFI method is an effective tool in assessing spatial heterogeneity of physiological activity over lichen thalli exposed to a variety of environmental factors. It may be also used to select a representative area at a lichen thallus before application of single-spot fluorometric techniques in lichens.     

Long Repeats in a Huge Genome: Microsatellite Loci in the Grasshopper Chorthippus biguttulus

It is commonly believed that both the average length and the frequency of microsatellites correlate with genome size. We have estimated the frequency and the average length for 69 perfect dinucleotide microsatellites in an insect with an exceptionally large genome: Chorthippus biguttulus (Orthoptera, Acrididae). Dinucleotide microsatellites are not more frequent in C. biguttulus, but repeat arrays are 1.4 to 2 times longer than in other insect species. The average repeat number in C. biguttulus lies in the range of higher vertebrates. Natural populations are highly variable. At least 30 alleles per locus were found and the expected heterozygosity is above 0.95 at all three loci studied. In contrast, the observed heterozygosity is much lower (≤0.51), which could be caused by long null alleles. [Reviewing Editor: Dr. Dmitri Petrov] Sequence data from this article have been deposited with the EMBL/GenBank databases under accession numbers AY532396–AY532400.     

Concerted evolution of a tandemly arrayed family of mating-specific genes in Phytophthora analyzed through inter- and intraspecific comparisons

Multigene families are features of most eukaryotic genomes, which evolve through a variety of mechanisms. This study describes the structure, expression, and evolution of a novel family in the oomycete Phytophthora. In the heterothallic species P. infestans, M96 is expressed specifically during sexual sporogenesis, and encodes a low-complexity extracellular protein that may be a component of oospore walls. Intriguingly, M96 exists in P. infestans as 22 relatively homogeneous loci tandemly repeated at a single site, which is partitioned by inversions and retroelements into subclusters exhibiting semi-independent evolution. M96 relatives were detected in other heterothallic and homothallic oomycetes including species closely (P. mirabilis, P. phaseoli) or distantly (P. ramorum, P. sojae) related to P. infestans. Those M96 relatives also exhibit oosporogenesis-specific expression and are arrayed multigene families. Nucleotide changes and repeat expansion diversify M96 in each species, however, paralogues are more related than orthologues. Concerted evolution through gene conversion and not strong purifying selection appears to be the major contributor to intraspecific homogenization. Divergence and concerted evolution was also detected between isolates of P. infestans. The divergence of M96 proteins between P. infestans, P. ramorum, and P. sojae exceeds that of typical proteins, reflecting trends in reproductive proteins from other kingdoms. Sequence data from this article have been deposited with the GenBank Data Libraries under accessions DQ196155 to DQ196175.     

Interspecific differences in differentiation and dedifferentiation patterns in the genusNicotiana

Differentiation on hormoneless media, habituation ability and crown gall induction inNicotiana tissue cultures have been used as physiological parameters of evolutionary differentiation between species. Some of them on hormone free media differentiated whole plantlets, others produced only shoots or roots or showed undifferentiated growth (habituation), some eventually died. Moreover, the same genotypes showed a differential behaviour as far as tumor formation byAgrobacterium tumefaciens was concerned. Particularly, the competence for crown gall transformation inNicotiana species seems negatively correlated with differentiation capacity and may be ascribed to differences in the plants capacity to synthesize growth regulators. The correlation between the results obtained and the phylogenetic position of the genotypes tested is finally discussed.     

Baculovirus-mediated Expression of a Gene for Trehalase of the Mealworm Beetle, Tenebrio molitor, in Insect Cells, SF-9, and Larvae of the Cabbage Armyworm, Mamestra brassicae

It is of interest to understand what kinds of physiological and biochemical changes occur in insects if the homeostasis of trehalose in the hemolymph is disrupted by the infection with a recombinant baculovirus containing a secretory-trehalase gene. For this purpose, two recombinant non-occluded Autographa california multicapsid nucleopolyhedroviruses (AcMNPVs), vTREVL and vERTVL, containing a trehalase cDNA of the mealworm beetle, Tenebrio molitor, were constructed. The trehalase cDNA was inserted in the sense orientation downstream of the polyhedrin promoter for vTREVL, and in the anitsense orientation for vERTVL. The active trehelase of T. molitor was found outside of cells when SF-9 cells or larvae of the cabbage armyworm, Mamestra brassicae, were infected with vTREVL. In the hemolymph of vTREVL-infected larvae, expression of the active trehelase was followed by disappearance of trehalose and appearance of glucose. However, the mortality time of virus-infected 5th instar larvae increased in the following order: AcMNPV C6 (wild-type virus) ≤ vERTVL < vTREVL. The symptoms (the browning and liquefying of the host body) of NPV infection were moderated considerably in vTREVL-infected larvae.     

Interspecific differences in Zn,Cd and Pb accumulation by freshwater algae and bryophytes

The relationships between the concentrations of zinc, cadmium and lead in aquatic plants and the concentrations of these metals in the ambient water have been compared for three algae (Lemanea fluviatilis, Cladophora glomerata, Stigeoclonium tenue), one liverwort (Scapania undulata) and three mosses (Amblystegium riparium, Fontinalis antipyretica, Rhynchostegium riparioides). The data to establish these relationships are all based on our own studies, some published already, some here for the first time. They come from a wide range of streams and rivers in Belgium, France, Germany, Ireland, Italy and the U.K. There were significant bivariate positive relationships between concentrations of Zn, Cd and Pb in water and plant for all species except Cd and Pb in Stigeoclonium tenue. When relationships were compared using datasets with total or filtrable metals in water, most differences were slight. However there were marked differences both between species and between metals. Comparison for the seven species of Zn in the plant when aqueous Zn is 0.01 mg l^–1, a concentration at which all seven were found, shows that the four bryophytes had the highest concentrations; however the two green algae had steeper slopes (representing change in concentration in plant in response to change in aqueous concentration). Lemanea fluviatilis had a slope closer to that of the bryophytes, but the concentration was about one order of magnitude lower. All seven species were found at a concentration of 0.01 mg l^–1 Pb, and at this concentration there were almost two orders of magnitude difference between the species which accumulated the most (Scapania undulata) and the one which accumulated the least (Cladophora glomerata). The steepest slope was however shown by C. glomerata.When multiple stepwise regression was applied, the aqueous metal under consideration was the first variable extracted in only nine of the 21 regressions. However one of the other heavy metals (aqueous or accumulated) was extracted first in all but one of the other regressions, presumably because the occurrences of Zn, Cd and Pb were strongly cross-correlated. The principal non-heavy metal factor extracted for Zn and Cd, but not Pb, was aqueous Ca. The relevance of these results to the use of aquatic plants for monitoring heavy metals is discussed.     

Evolution of restriction sites of ribosomal DNA in natural populations of the field mouse,Apodemus speciosus

An analysis by restriction endonuclease digestion of ribosomal DNA (rDNA) was carried out in natural populations of Apodemus speciosus, a field mouse that is endemic to Japan. Two restriction sites, the EcoRI (E3) and DraI (D4) sites, in the nontranscribed spacer region downstream from the gene for 28S RNA showed polymorphism within and between individuals in the populations from the Japanese main islands. By contrast, populations from the small adjoining islands which are thought to have separated from the main islands 1–2 × 10⁴ years ago showed relatively low levels of polymorphism within and between individuals, i.e., one of the polymorphic bands in the case of each enzyme was predominant in these populations, irrespective of the variants. These results indicate that the rate of fixation of site variations depends on population size and that the direction of fixation is random. Furthermore, each polymorphic restriction site seems to be fixed independently.Correspondence to: H. Suzuki     

Interspecific Differences of Leaf Gas Exchange and Water Relations of Three Evergreen Mediterranean Shrub Species

Leaf gas exchange and plant water relations of three co-occurring evergreen Mediterranean shrubs species, Quercus ilex L. and Phillyrea latifolia L. (typical evergreen sclerophyllous shrubs) and Cistus incanus L. (a drought semi-deciduous shrub), were investigated in order to evaluate possible differences in their adaptive strategies, in particular with respect to drought stress. C. incanus showed the highest annual rate of net photosynthetic rate (P _N) and stomatal conductance (g _s) decreasing by 67 and 69 %, respectively, in summer. P. latifolia and Q. ilex showed lower annual maximum P _N and g _s, although P _N was less lowered in summer (40 and 37 %, respectively). P. latifolia reached the lowest midday leaf water potential (₁) during the drought period (–3.54±0.36 MPa), 11 % lower than in C. incanus and 19 % lower than in Q. ilex. Leaf relative water content (RWC) showed the same trend as ₁. C. incanus showed the lowest RWC values during the drought period (60 %) while they were never below 76 % in P. latifolia and Q. ilex; moreover C. incanus showed the lowest recovery of ₁ at sunset. Hence the studied species are well adapted to the prevailing environment in Mediterranean climate areas, but they show different adaptive strategies that may be useful for their co-occurrence in the same habitat. However, Q. ilex and P. latifolia by their water use strategy seem to be less sensitive to drought stress than C. incanus.     

Cloning, Expression and Characterization of a Thiolase Gene from Clostridium pasteurianum

A thl gene encoding the thiolase (EC 2.3.1.9) of Clostridium pasteurianum was cloned by thermal asymmetric interlaced (TAIL) PCR. It consists of 1179 bp with 36.8% GC content and encodes 392 amino acids with a deduced molecular mass of 40,954 Da and shows 77% identity and 88% similarity to that of Clostridium tetani E88 and should be classified as a biosynthetic thiolase with three conserved residues Cys89, Cys382 and His352. The gene was over-expressed in Escherichia coli and the thiolase was purified with Ni-NTA agarose column to homogeneity. The K _m of this thiolase for acetoacetyl-CoA is 0.13 mM with 0.06 mM CoASH at pH 8.2, 25°C and a V _max value of 46 μmol min⁻¹ mg⁻¹.