Positioning the expanded akirin gene family of Atlantic salmon within the transcriptional networks of myogenesis

Vertebrate akirin genes usually form a family with one-to-three members that regulate gene expression during the innate immune response, carcinogenesis and myogenesis. We recently established that an expanded family of eight akirin genes is conserved across salmonid fish. Here, we measured mRNA levels of the akirin family of Atlantic salmon (Salmo salar L.) during the differentiation of primary myoblasts cultured from fast-skeletal muscle. Using hierarchical clustering and correlation, the data was positioned into a network of expression profiles including twenty further genes that regulate myogenesis. akirin1(2b) was not significantly regulated during the maturation of the cell culture. akirin2(1a) and 2(1b), along with IGF-II and several igfbps, were most highly expressed in mononuclear cells, then significantly and constitutively downregulated as differentiation proceeded and myotubes formed/matured. Conversely, akirin1(1a), 1(1b), 1(2a), 2(2a) and 2(2b) were expressed at lowest levels when mononuclear cells dominated the culture and highest levels when confluent layers of myotubes were evident. However, akirin1(2a) and 2(2a) were first upregulated earlier than akirin1(1a), 1(1b) and 2(2b), when rates of myoblast proliferation were highest. Interestingly, akirin1(1b), 1(2a), 2(2a) and 2(2b) formed part of a module of co-expressed genes involved in muscle differentiation, including myod1a, myog, mef2a, 14-3-3β and 14-3-3γ. All akirin paralogues were expressed ubiquitously across ten tissues, although mRNA levels were regulated between cell-types and family members. Gene expression patterns were often highly correlated between akirin paralogues, suggesting that natural selection has maintained an intricate network of co-regulation among family members. We concluded that the Atlantic salmon akirin family performs a multifaceted role during myogenesis and has physiological functions spanning many cell-types.     

Functional differentiation of bmp2a and bmp2b genes in zebrafish

Bone morphogenetic protein 2 plays an important role in the regulation of osteoblast proliferation and differentiation. Phylogenetic analysis showed that the bmp2 ortholog evolved from the same ancestral gene family in vertebrates and was duplicated in teleost, which were named bmp2a and bmp2b. The results of whole-mount in situ hybridization showed that the expression locations of bmp2a and bmp2b in zebrafish were different in different periods (24 hpf, 48 hpf, 72 hpf), which revealed potential functional differentiation between bmp2a and bmp2b. Phenotypic analysis showed that bmp2a mutations caused partial rib and vertebral deformities in zebrafish, while bmp2b^−/− embryos died massively after 12 hpf due to abnormal somite formation. We further explored the expression pattern changes of genes (bmp2a, bmp2b, smad1, fgf4, runx2b, alp) related to skeletal development at different developmental stages (20 dpf, 60 dpf, 90 dpf) in wild-type and bmp2a^−/− zebrafish. The results showed that the expression of runx2b in bmp2a^−/− was significantly downregulated at three stages and the expression of other genes were significantly downregulated at 90 dpf compared with wild-type zebrafish. The study revealed functional differentiation of bmp2a and bmp2b in zebrafish embryonic and skeletal development.     

Discovery of a new fragrance allele and development of functional markers for identifying diverse fragrant genotypes in rice

Discovery of new fragrance alleles provides important genetic resources for breeding fragrant rice. In this study, a hybrid complementation test demonstrated the association of a new fragrance allele without mutation in the coding region with flavor formation in a fragrant rice variety Nankai 138. The new allele (badh2-p-5′UTR) has a 3-bp deletion in the 5′ untranslated region and an 8-bp insertion in the promoter (?1,314 site upstream from the initiation codon). Surprisingly, we found that there is also an 8-bp insertion in the promoter of the badh2-E7 allele. We developed a new sequence tagged site functional marker to identify the badh2-p-5′UTR and badh2-E7 alleles according to the 8-bp insertion in their promoters. A cleaved amplified polymorphic sequence (AluI) functional marker targeting a common base substitution in the intron 2 of three badh2 alleles, viz. badh2-p-5′UTR, badh2-E7 and badh2-E2, was developed to identify diverse genotypes for fragrance in rice. Based on the results of sequence alignments among the three badh2 alleles, we suggest that the badh2-E7 and badh2-p-5′UTR alleles may have the same genetic origin. In addition, the genetic distance between the badh2-E7 and badh2-p-5′UTR alleles may be closer than that between the badh2-E2 and the badh2-p-5′UTR alleles, or between the badh2-E2 and the badh2-E7 alleles.     

Genetic specifics of the breeding of cotton varieties with cleistogamous flowers

As a prerequisite to studying the genetics and breeding of chasmogamous and cleistogamous flowers, a preliminary experiment was performed to estimate the extent of cross-pollination in cotton varieties and hybrids. Vicinism estimates varied from 0.53 to 15.36%, i.e., the proportion of cross-pollination was relatively high, leading to a biological contamination. As a result of such contamination, genetic collection lines and varieties lose genetic homogeneity and become heterozygous and genetically heterogeneous. The genetic control of the flower type was studied in the Gossipium hirsutum L. × G. barbadense L. interspecific hybrids, and phenotypic segregation of the 3: 1 and 15: 1 types with monogenic (3: 1) and digenic (15: 1) differences of noncumulative polymerization was observed. The corresponding types of genotypic segregation were 1: 2: 1 (1Cg ₁ Cg ₁ cg ₂ cg ₂ : 2Cg ₁ cg ₁ cg ₂ cg ₂ : 1cg ₁ cg ₁ cg ₂ cg ₂ ) and 1: 2: 2: 4: 1: 2: 1: 2: 1 (1) Cg ₁ Cg ₁ Cg ₂ Cg ₂ -1; (2) Cg ₁ Cg ₁ cg ₂ cg ₂ -2; (3) Cg ₁ cg ₁ Cg ₂ Cg ₂ -2; (4) Cg ₁ cg ₁ Cg ₂ cg ₂-4; (5) Cg ₁ Cg ₁ cg ₂ cg ₂ -1; (6) Cg ₁ cg ₁ cg ₂ cg ₂ -2; (7) cg ₁ cg ₁ Cg ₂ Cg ₂-1; (8) cg ₁ cg ₁ Cg ₂ cg ₂ -2; (9) cg ₁ cg ₁ cg ₂ cg ₂ -1. Genotypes (1)–(8) had chasmogamous flowers, while double-recessive genotype (9) had cleistogamous flowers. Based on this, genotypes with individual phenotypic expression were identified in F2, and their correlation with the most important morphological, biological, and agricultural features was studied. Special attention was paid to the productivity of hybrid plants intended for use in breeding to obtain intensive varieties. The study made it possible to isolate forms, families, genetic collection lines, and varieties with isogenic or nonisogenic determination of these characters and chasmogamous and cleistogamous flowers of G. hirsutum L. and G. barbadense L. prototypes by using original methods to examine the two types of flowers; the methods do not have analogs in cotton breeding worldwide.     

Identification of an amino acid residue required for differential recognition of a viral movement protein by the Tomato mosaic virus resistance gene Tm-2(2)

The Tm-2 gene of tomato and its allelic gene, Tm-2², confer resistance to Tomato mosaic virus (ToMV) and encode a member of the coiled-coil/nucleotide binding-ARC/leucine-rich repeat (LRR) protein class of plant resistance (R) genes. Despite exhibiting only four amino acid differences between the products of Tm-2 and Tm-2², Tm-2² confers resistance to ToMV mutant B7, whereas Tm-2 is broken by ToMV-B7. An Agrobacterium-mediated transient expression system was used to study the mechanism of differential recognition of the movement proteins (MPs), an avirulence factor for ToMV resistance, of ToMV-B7 by Tm-2 and Tm-2². Although resistance induced by Tm-2 and Tm-2² is not usually accompanied by hypersensitive response (HR), Tm-2 and Tm-2² induced HR-like cell death by co-expression with MP of a wild-type ToMV, a strain that causes resistance for these R genes, and Tm-2² but not Tm-2 induced cell death with B7-MP in this system. Site-directed amino acid mutagenesis revealed that Tyr-767 in the LRR of Tm-2² is required for the specific recognition of the B7-MP. These results suggest that the Tyr residue in LRR contributes to the recognition of B7-MP, and that Tm-2 and Tm-2² are involved in HR cell death.     

A mating-type factors of Coprinus cinereus have variable numbers of specificity genes encoding two classes of homeodomain proteins

We have identified the seven genes that constitute the A43 mating-type factor of Coprinus cinereus and compare the organisation of A43 with the previously characterised A42 factor. In both, the genes that trigger clamp cell development, the so-called specificity genes, are separated into α and β loci by 7 kb of noncoding sequence and are flanked by homologous genes α-fg and β-fg. The specificity genes are known to encode two classes of dissimilar homeodomain (HD1 and HD2) proteins and have different allelic forms which show little or no cross-hybridisation. By partial sequencing we identified a divergently transcribed HD1 (a1-2) and HD2 (a2-2) gene in the A43 α locus. a2-2 failed to elicit clamp cell development in three different hosts, suggesting that it is non-functional. a1-2 elicited clamp cells in an A42 host that has only an HD2 gene (a2-1) in its α locus, thus demonstrating that the compatible Aα mating interaction is between an HD1 and an HD2 protein. The A43 β locus contains three specificity genes, the divergently transcribed HD1 and HD2 genes b1-2 and b2-2 and a third HD1 gene (d1-1) that was shown by hybridisation and transformation analyses to be functionally equivalent to d1-1 in A42. An untranscribed footprint of a third A42 HD1 gene, c1-1, was detected between the A43 b2-2 and d1-1 genes by Southern hybridisation.     

Selective Impairment of a Subset of Ran-GTP-binding Domains of Ran-binding Protein 2 (Ranbp2) Suffices to Recapitulate the Degeneration of the Retinal Pigment Epithelium (RPE) Triggered by Ranbp2 Ablation

Retinal pigment epithelium (RPE) degeneration underpins diseases triggered by disparate genetic lesions, noxious insults, or both. The pleiotropic Ranbp2 controls the expression of intrinsic and extrinsic pathological stressors impinging on cellular viability. However, the physiological targets and mechanisms controlled by Ranbp2 in tissue homeostasis, such as RPE, are ill defined. We show that mice, RPE-cre::Ranbp2^−/−, with selective Ranbp2 ablation in RPE develop pigmentary changes, syncytia, hypoplasia, age-dependent centrifugal and non-apoptotic degeneration of the RPE, and secondary leakage of choriocapillaris. These manifestations are accompanied by the development of F-actin clouds, metalloproteinase-11 activation, deregulation of expression or subcellular localization of critical RPE proteins, atrophic cell extrusions into the subretinal space, and compensatory proliferation of peripheral RPE. To gain mechanistic insights into what Ranbp2 activities are vital to the RPE, we performed genetic complementation analyses of transgenic lines of bacterial artificial chromosomes of Ranbp2 harboring loss of function of selective Ranbp2 domains expressed in a Ranbp2^−/− background. Among the transgenic lines produced, only Tg^RBD2/3*-HA::RPE-cre::Ranbp2^−/−-expressing mutations, which selectively impair binding of RBD2/3 (Ran-binding domains 2 and 3) of Ranbp2 to Ran-GTP, recapitulate RPE degeneration, as observed with RPE-cre::Ranbp2^−/−. By contrast, Tg^RBD2/3*-HA expression rescues the degeneration of cone photoreceptors lacking Ranbp2. The RPE of RPE-cre::Ranbp2^−/− and Tg^RBD2/3*-HA::RPE-cre::Ranbp2^−/− share proteostatic deregulation of Ran GTPase, serotransferrin, and γ-tubulin and suppression of light-evoked electrophysiological responses. These studies unravel selective roles of Ranbp2 and its RBD2 and RBD3 in RPE survival and functions. We posit that the control of Ran GTPase by Ranbp2 emerges as a novel therapeutic target in diseases promoting RPE degeneration.     

Cytogenetic characterization of ten araneomorph spiders (Araneae): Karyotypes and meiotic features

Karyotypes, sex chromosome systems and meiotic characteristics are reported for ten spider species belonging to the families Gnaphosidae, Philodromidae, Salticidae, Oxyopidae and Sicariidae by using standard Giemsa staining. The male diploid numbers (2n) and sex chromosome systems are as follows: Berinda hakani 2n = 22 (X₁X₂), Berinda ensigera 2n = 22 (X₁X₂), Trachyzelotes lyonneti 2n = 22 (X₁X₂), Trachyzelotes malkini 2n = 22 (X₁X₂), Zelotes caucasius 2n = 22 (X₁X₂) (Gnaphosidae); Thanatus pictus 2n = 28 (X₁ X₂), Tibellus macellus 2n = 24 (X₁ X₂) (Philodromidae); Neon reticulatus 2n = 21 (X₀) (Salticidae); Peucetia virescens 2n = 28 (X₁X₂) (Oxyopidae) and Loxosceles rufescens 2n = 21 (X₁ X₂Y) (Sicariidae). All species have monoarmed chromosomes with the exception of L. rufescens that has biarmed (metacentric and submetacentric) chromosomes. The obtained data are the first results for the genera Berinda, Trachyzelotes and Neon. Additionally, with the exception of L. rufescens, all species are being chromosomally analyzed for the first time.     

Change in State following Transposition of a Regulatory Element of the Enhancer System in Maize

From an original A2 allele (colored aleurone), a mutable allele, a2-m-4 1629, that changes from a2 to A2 is described. Mutability is expressed as a very distinct pattern limited to the last cell division.—The mutability of a2-m-4 1629 is autonomously controlled by an En at the a2 locus. This En, inactive on standard a testers for En, is partially active on a2-m-1, an a2 tester for En, and expresses varied levels of activity from limited to nearly full suppression of the a2-m-1 color phenotype.—When the En of the a2-m-4 1629 allele transposes from the a2 locus, it behaves, at the new position, like a standard En in triggering a2-m-1, a-m-1 and a-m(r), which express colored spots on a colorless background. The activity of En is therefore different following the change in chromosome location. This finding supports the "position" hypothesis that has been proposed to explain diverse patterns observed among controlling elements. In this case mutation is related to the terminal cell state and not to tissue differences as shown with some phase-variation regulatory elements.     

Study of the ref(2)P locus of Drosophila melanogaster

The ref(2)P gene of Drosophila melanogaster is implicated in sigma rhabdovirus multiplication. Two common alleles of ref(2)P are known, ref(2)P ⁰ which permits sigma virus multiplication and ref(2)P ^pwhich is restrictive for most sigma virus strains. This gene maps to the cytogenetic region 37E3-F3. Using Df(2L)E55 (=Df(2L)37D2-El;37F5-38A1), we have screened for lethal, semi-lethal and visible mutations following diepoxybutane (DEB) or ethyl methanesulfonate (EMS) mutagenesis. Our data confirm than DEB is mor efficient than EMS at inducing deletions. The mutations obtained in this region define 14 complementation groups. One of them, l(2)37Dh, appears to be a general enhancer of Minute and Minute-like mutations. None of the mutations were allelic to the ref(2)P locus. Loss-of-function alleles of ref(2)P (called null) were selected following DEB mutagenesis. Homozygous or hemizygous ref(2)P ^nullflies are male sterile. These flies, like homozygous or hemizygous ref(2)P ⁰flies, are fully permissive for sigma virus replication. We suggest that the ref(2)P products interact with viral products, but that this interaction is not necessary for an efficient viral cycle.     

Karyology of eight species of bats (Mammalia: Chiroptera) from Hainan Island,China

Karyotypes and chromosomal data are presented for eight bat species representing two families (Rhinolophidae and Vespertilionidae) from Hainan Island, China. The species investigated were Rhinolophus lepidus (2n = 62, FN = 60), R. pusillus (2n = 62, FN = 60), R. affinis (2n = 62, FN = 60), R. sinicus (2n = 36, FN = 60), Myotis horsfieldi (2n = 44, FN = 52), Pipistrellus abramus (2n = 26, FN = 44), Miniopterus australis (2n = 46, FN = 50) and M. schreibersii (2n = 46, FN = 50). The karyotype of Rhinolophus lepidus is reported for the first time.     

Variance of actual inbreeding

The variances of actual inbreeding and coancestry in terms of their corresponding identities by descent were studied for finite populations. For inbreeding at a single locus, the total variance σ² = F(1 ? F) (F is the inbreeding coefficient) is comprised of a component σ_w² within populations and a component σ_b² between replicate populations. These variances increase in time to a maximum at about 1.1N_e generations for σ_w², about 2.3N_e generations for σ_b², and about 1.4N_e generations for σ², and decrease thereafter (N_e is effective population size). The ratio $\text{σ}{}_{\mathrm{<mtext>b</mtext>}}{}^2\text{σ}{}^2$ is ever increasing to an asymptote in the range 0.4-0.5 depending on N_e and the mating system. For finite populations with variation in pedigree F's, there are contributions σ_wF² within and σ_bF² between populations. The component σ_bF² is insignificant except for very small populations, and σ_wF² is largest in the early generations and then decreases roughly as $\text{(1 ? F)}{}^2\text{KN}{}_{\mathrm{<mtext>e</mtext>}}\text{where}\text{K}$ is formulated in terms of the mating strategy and the degree of avoidance of mating relatives. An additional degree of avoidance increases K by a factor of 4. In a large population at equilibrium with respect to mixed self and random mating, σ_wF² accounts for onehalf to two-thirds of σ_w². Bringing in more loci leads to the decomposition of the total variance into four components whose values are affected by linkages among the loci. The relationships between these components and σ_w², σ_b², σ_wF², and σ_bF², are elaborated in terms of tight and loose linkage. The exact computations of σ_wF² and σ_bF² require the use of two locus descent measures without linkage. The variances of various averages of actual identities by descent, such as the proportions for individuals or populations, are formulated for a sample of individuals.     

Multinuclear NMR study and crystal structures of complexes of the types cis- and trans-Pt(amine)2I2

Complexes of the types cis- and trans-Pt(amine)₂I₂ were studied by spectroscopic methods, especially by multinuclear NMR spectroscopy. In ¹⁹⁵Pt NMR, the cis diiodo compounds with primary amines were observed between −3342 and −3357 ppm in acetone, while the trans compounds were found between −3336 and −3372 ppm. For the secondary amines, the chemical shifts were observed at lower fields. In ¹H NMR, the trans complexes were observed at higher fields than the cis compounds, while in ¹³C NMR, the reverse was observed. The ²J(¹⁹⁵Pt-¹H) and ³J(¹⁹⁵Pt-¹H) coupling constants are larger for the cis compounds (ave. 67 and 45 Hz, respectively) than for the trans isomers (ave. 59 and 38 Hz). In ¹³C NMR, the values of ²J(¹⁹⁵Pt-¹³C) and ³J(¹⁹⁵Pt-¹³C) were also found to be larger for the cis complexes (ave. 17 and 39 Hz versus 11 and 28 Hz). There seems to be a slight dependence of the pK_a values of the protonated amines or the proton affinity in the gas phase with the δ(Pt) chemical shifts. The crystal structures of eight diiodo complexes were determined. These compounds are cis-Pt(CH₃NH₂)₂I₂, cis-Pt(n-C₄H₉NH₂)₂I₂, cis-Pt(Et₂NH)₂I₂, trans-Pt(n-C₃H₇NH₂)₂I₂, trans-Pt(iso-C₃H₇NH₂)₂I₂, trans-Pt(n-C₄H₉NH₂)₂I₂, trans-Pt(t-C₄H₉NH₂)₂I₂ and trans-Pt(Me₂NH)₂I₂. The Pt-N bond distances located in trans position to the iodo ligands were compared to those located in trans position to the amines. The Pt-N bond in cis-Pt(Et₂NH)₂I₂ are much longer than the others, probably caused by the steric hindrance of the two very bulky ligands located in cis positions.     

Combination of the Loss of cmnmU34 with the Lack of sU34 Modifications of tRNA, tRNA, and tRNA Altered Mitochondrial Biogenesis and Respiration

Yeast Saccharomyces cerevisiae MTO2, MTO1, and MSS1 genes encoded highly conserved tRNA modifying enzymes for the biosynthesis of carboxymethylaminomethyl (cmnm)⁵s²U₃₄ in mitochondrial tRNA^Lys, tRNA^Glu, and tRNA^Gln. In fact, Mto1p and Mss1p are involved in the biosynthesis of the cmnm⁵ group (cmnm⁵U₃₄), while Mto2p is responsible for the 2-thiouridylation (s²U₃₄) of these tRNAs. Previous studies showed that partial modifications at U₃₄ in mitochondrial tRNA enabled mto1, mto2, and mss1 strains to respire. In this report, we investigated the functional interaction between MTO2, MTO1, and MSS1 genes by using the mto2, mto1, and mss1 single, double, and triple mutants. Strikingly, the deletion of MTO2 was synthetically lethal with a mutation of MSS1 or deletion of MTO1 on medium containing glycerol but not on medium containing glucose. Interestingly, there were no detectable levels of nine tRNAs including tRNA^Lys, tRNA^Glu, and tRNA^Gln in mto2/mss1, mto2/mto1, and mto2/mto1/mss1 strains. Furthermore, mto2/mss1, mto2/mto1, and mto2/mto1/mss1 mutants exhibited extremely low levels of COX1 and CYTB mRNA and 15S and 21S rRNA as well as the complete loss of mitochondrial protein synthesis. The synthetic enhancement combinations likely resulted from the completely abolished modification at U₃₄ of tRNA^Lys, tRNA^Glu, and tRNA^Gln, caused by the combination of eliminating the 2-thiouridylation by the mto2 mutation with the absence of the cmnm⁵U₃₄ by the mto1 or mss1 mutation. The complete loss of modifications at U₃₄ of tRNAs altered mitochondrial RNA metabolisms, causing a degradation of mitochondrial tRNA, mRNA, and rRNAs. As a result, failures in mitochondrial RNA metabolisms were responsible for the complete loss of mitochondrial translation. Consequently, defects in mitochondrial protein synthesis caused the instability of their mitochondrial genomes, thus producing the respiratory-deficient phenotypes. Therefore, our findings demonstrated a critical role of modifications at U₃₄ of tRNA^Lys, tRNA^Glu, and tRNA^Gln in maintenance of mitochondrial genome, mitochondrial RNA stability, translation, and respiratory function.     

An approximation of the minimum-variance estimator of heritability based on variance component analysis

An approximate minimum-variance estimate of heritability (h²) is proposed, using the sire and dam components of variance from a hierarchical analysis of variance. The minimum sampling variance is derived for unbalanced data. Optimum structures for the estimation of h² are given for the balanced case. The degree to which ĥ² is more precise than the equally weighted estimate ĥ²_S+D is a function of the size and structure of the sample used. However, computer simulation reveals that ĥ² has less desirable behavior than ĥ²_S+D. An iterative procedure improved the estimation of h², especially in small populations, when those values of ĥ²_S or ĥ²_D outside the range of the parameter were constrained to zero or unity.