Evolutionary modes of emergence of short interspersed nuclear element (SINE) families in grasses

Short interspersed nuclear elements (SINEs) are non‐autonomous transposable elements which are propagated by retrotransposition and constitute an inherent part of the genome of most eukaryotic species. Knowledge of heterogeneous and highly abundant SINEs is crucial for de novo (or improvement of) annotation of whole genome sequences. We scanned Poaceae genome sequences of six important cereals (Oryza sativa, Triticum aestivum, Hordeum vulgare, Panicum virgatum, Sorghum bicolor, Zea mays) and Brachypodium distachyon to examine the diversity and evolution of SINE populations. We comparatively analyzed the structural features, distribution, evolutionary relation and abundance of 32 SINE families and subfamilies within grasses, comprising 11 052 individual copies. The investigation of activity profiles within the Poaceae provides insights into their species‐specific diversification and amplification. We found that Poaceae SINEs (PoaS) fall into two length categories: simple SINEs of up to 180 bp and dimeric SINEs larger than 240 bp. Detailed analysis at the nucleotide level revealed that multimerization of related and unrelated SINE copies is an important evolutionary mechanism of SINE formation. We conclude that PoaS families diversify by massive reshuffling between SINE families, likely caused by insertion of truncated copies, and provide a model for this evolutionary scenario. Twenty‐eight of 32 PoaS families and subfamilies show significant conservation, in particular either in the 5′ or 3′ regions, across Poaceae species and share large sequence stretches with one or more other PoaS families.     

Short interspersed nuclear element (SINE) sequences of the Bovidae

DNA sequences from Bovidae (cattle, goats and sheep) in the EMBL nucleotide database contain several short interspersed repeated sequences (SINEs). Three different SINEs have been found: Bov-A2, containing two 115-bp A elements; Bov-tA, a tRNA pseudogene coupled to an A element; and Bov-B of 560 bp or less and partially homologous to the A element. Bov-A2, Bov-tA and Bov-B occupy about 1.8%, 1.6% and 0.5%, respectively, of the bovine genome as represented in the nucleotide database. Apart from a tRNA-like sequence in both Bov-tA and the porcine SINEs, there was no similarity with the porcine SINEs. Apparently, the artiodactyle SINEs were established after the divergence leading to the Suidae and Bovidae but before the radiation within these families. Oligonucleotides were designed for a specific amplification of DNA from Bovidae.     

Gene conversion as a secondary mechanism of short interspersed element (SINE) evolution.

The Alu repetitive family of short interspersed elements (SINEs) in primates can be subdivided into distinct subfamilies by specific diagnostic nucleotide changes. The older subfamilies are generally very abundant, while the younger subfamilies have fewer copies. Some of the youngest Alu elements are absent in the orthologous loci of nonhuman primates, indicative of recent retroposition events, the primary mode of SINE evolution. PCR analysis of one young Alu subfamily (Sb2) member found in the low-density lipoprotein receptor gene apparently revealed the presence of this element in the green monkey, orangutan, gorilla, and chimpanzee genomes, as well as the human genome. However, sequence analysis of these genomes revealed a highly mutated, older, primate-specific Alu element was present at this position in the nonhuman primates. Comparison of the flanking DNA sequences upstream of this Alu insertion corresponded to evolution expected for standard primate phylogeny, but comparison of the Alu repeat sequences revealed that the human element departed from this phylogeny. The change in the human sequence apparently occurred by a gene conversion event only within the Alu element itself, converting it from one of the oldest to one of the youngest Alu subfamilies. Although gene conversions of Alu elements are clearly very rare, this finding shows that such events can occur and contribute to specific cases of SINE subfamily evolution.     

The Au family, a novel short interspersed element (SINE) from Aegilops umbellulata

A novel plant short interspersed nuclear element (SINE) was identified in the second intron of the acetyl CoA carboxylase gene of Aegilops umbellulata which has been designated ”Au”, for the host species in which it was discovered. Au elements have a tRNA-related region, direct flanking repeats, and a short stretch of T at the 3′ end, which are features common to Au and previously characterized SINEs. Au elements are detected in the genomes of several monocots and dicots by DNA dot hybridization and are also found in the tobacco genome by database searching. Au elements are present at an especially high copy number (approximately 10⁴ copies per haploid genome) in wheat and Ae. umbellulata. This suggests a recent amplification of Au in the Triticum and Aegilops species. In situ hybridization revealed a dispersed distribution of Au elements on wheat chromosomes. Au elements were amplified by PCR from monocot and dicot species and the phylogenetic relationships among Au elements were inferred. This phylogenetic analysis suggests amplification of Au elements in a manner consistent with the retrotransposon model for SINE dispersion. The high copy number of Au elements and their dispersed distribution in wheat are desirable characteristics for a molecular marker system in this important species. Received: 15 April 2000 / Accepted: 24 August 2000     

Divergence of 3′ ends as a driver of short interspersed nuclear element (SINE) evolution in the Salicaceae

Short interspersed nuclear elements (SINEs) are small, non‐autonomous and heterogeneous retrotransposons that are widespread in plants. To explore the amplification dynamics and evolutionary history of SINE populations in representative deciduous tree species, we analyzed the genomes of the six following Salicaceae species: Populus deltoides, Populus euphratica, Populus tremula, Populus tremuloides, Populus trichocarpa, and Salix purpurea. We identified 11 Salicaceae SINE families (SaliS‐I to SaliS‐XI), comprising 27 077 full‐length copies. Most of these families harbor segmental similarities, providing evidence for SINE emergence by reshuffling or heterodimerization. We observed two SINE groups, differing in phylogenetic distribution pattern, similarity and 3′ end structure. These groups probably emerged during the ‘salicoid duplication’ (~65 million years ago) in the Salix–Populus progenitor and during the separation of the genus Salix (45–65 million years ago), respectively. In contrast to conserved 5′ start motifs across species and SINE families, the 3′ ends are highly variable in sequence and length. This extraordinary 3′‐end variability results from mutations in the poly(A) tail, which were fixed by subsequent amplificational bursts. We show that the dissemination of newly evolved 3′ ends is accomplished by a displacement of older motifs, leading to various 3′‐end subpopulations within the SaliS families.     

Short interspersed element (SINE) depletion and long interspersed element (LINE) abundance are not features universally required for imprinting

Genomic imprinting is a form of gene dosage regulation in which a gene is expressed from only one of the alleles, in a manner dependent on the parent of origin. The mechanisms governing imprinted gene expression have been investigated in detail and have greatly contributed to our understanding of genome regulation in general. Both DNA sequence features, such as CpG islands, and epigenetic features, such as DNA methylation and non-coding RNAs, play important roles in achieving imprinted expression. However, the relative importance of these factors varies depending on the locus in question. Defining the minimal features that are absolutely required for imprinting would help us to understand how imprinting has evolved mechanistically. Imprinted retrogenes are a subset of imprinted loci that are relatively simple in their genomic organisation, being distinct from large imprinting clusters, and have the potential to be used as tools to address this question. Here, we compare the repeat element content of imprinted retrogene loci with non-imprinted controls that have a similar locus organisation. We observe no significant differences that are conserved between mouse and human, suggesting that the paucity of SINEs and relative abundance of LINEs at imprinted loci reported by others is not a sequence feature universally required for imprinting.     

First application of the SINE (short interspersed repetitive element) method to infer phylogenetic relationships in reptiles: an example from the turtle superfamily Testudinoidea

Although turtles (order Testudines) constitute one of the major reptile groups, their phylogenetic relationships remain largely unresolved. Hence, we attempted to elucidate their phylogeny using the SINE (short interspersed repetitive element) method, in which the sharing of a SINE at orthologous loci is indicative of synapomorphy. First, a detailed characterization of the tortoise polIII/SINE was conducted using 10 species from eight families of hidden-necked turtles (suborder Cryptodira). Our analysis of 382 SINE sequences newly isolated in the present study revealed two subgroups, namely Cry I and Cry II, which were distinguishable according to diagnostic nucleotides in the 3' region. Furthermore, four (IA-ID) and five (IIA-IIE) different SINE types were identified within Cry I and Cry II subgroups, respectively, based on features of insertions/deletions located in the middle of the SINE sequences. The relative frequency of occurrence of the subgroups and the types of SINEs in this family were highly variable among different lineages of turtles, suggesting active differential retroposition in each lineage. Further application of the SINE method using the most retrotranspositionally active types, namely IB and IC, challenged the established phylogenetic relationships of Bataguridae and its related families. The data for 11 orthologous loci demonstrated a close relationship between Bataguridae and Testudinidae, as well as the presence of the three clades within Bataguridae. Although the SINE method has been used to infer the phylogenies of a number of vertebrate groups, it has never been applied to reptiles. The present study represents the first application of this method to a phylogenetic analysis of this class of vertebrates, and it provides detailed information on the SINE subgroups and types. This information may be applied to the phylogenetic resolution of relevant turtle lineages.     

Robertsonian translocation (8;14) in an infertile bitch (Canis familiaris)

A new case of a Robertsonian translocation diagnosed in an infertile bitch of West Highland White Terrier breed is reported. Cytogenetic analyses were performed with the use of Giemsa staining, Q-, C- and Ag-I banding techniques and FISH with canine whole X chromosome paint. The karyotype of this bitch was described as 77,XX der (8;14)(q10;q10).     

Isotope recycling in lactating dogs (Canis familiaris)

Isotope-based techniques for the measurement of water turnover, energy expenditure, and milk intake often assume that there is no recycling of isotopes once they have left the labeled animal. In experiments involving lactating females or their suckling offspring, there are several possible routes of isotope recycling. These include the consumption of labeled milk by offspring, the ingestion of labeled excreta, and the rebreathing of exhaled labeled CO(2) or water vapor by both mother and offspring. Isotope recycling might be especially important during lactation because the offspring are in close contact with each other and their mother for prolonged periods. We show here in 24- to 30-day-old domestic dog Canis familiaris puppies that there was no detectable transfer of (18)O or (2)H from labeled to unlabeled pups in two litters (16 pups, 8 labeled, 8 unlabeled) that were weaned early and independent of their mother. However, there was a significant transfer of both isotopes from labeled to unlabeled pups and from labeled pups to their mothers in nine equivalent nursing litters of the same age (27 labeled, 26 unlabeled pups). The increases in enrichment of isotopes in unlabeled offspring were greater than the increases in enrichment of the mothers. This indicates that maternal ingestion of offspring excreta and subsequent transfer of isotope in milk is not the sole pathway of recycling. Additional routes must also be important, such as exchange of isotope between pups on saliva-coated nipples and perhaps direct ingestion of excreta by unweaned young. Recycling is unlikely to be an important factor when determining maternal metabolic rate during peak lactation in domestic dogs. However, experiments that are designed to assess the energy demands of pups and isotope-based estimates of water turnover in offspring may need to take into account any effects of isotope recycling. In a theoretical example, removing the effects of recycling increased the measured energy expenditure in pups by up to 7% and increased the calculated elimination rates of both isotopes by up to 11.1% in (18)oxygen and 10.9% in (2)hydrogen.     

Peptidergic regulation of feeding in the dog (Canis familiaris)

Although the incidence of obesity in the domesticated dog is high, few studies have investigated the regulation of food intake in this species. In the present study we investigated the response of the dog to a number of putative satiety agents including cholecystokinin (CCK), bombesin, calcitonin and naloxone. CCK significantly suppressed food intake during a scheduled fifteen minute meal in intact dogs and in dogs receiving total subdiaphragmatic vagotomies. Emesis occurred following injection of higher doses of CCK in most dogs. Bombesin and calcitonin reduced intake in both normal and vagotomized dogs, although higher doses of calcitonin were needed to significantly suppress feeding in vagotomized dogs compared with intact animals. Naloxone reduced feeding by as much as 60% in intact and vagotomized animals. Glucagon suppressed feeding in intact dogs, but not in vagotomized animals. Somatostatin and pancreatic polypeptide did not alter food intake. Thus the domesticated dog responds somewhat differently to some neuropeptides compared with the laboratory rat stressing the importance of examining the regulation of food intake across species.     

Pstl repeat: a family of short interspersed nucleotide element (SINE)-like sequences in the genomes of cattle, goat, and buffalo.

The PstI family of elements are short, highly repetitive DNA sequences interspersed throughout the genome of the Bovidae. We have cloned and sequenced some members of the PstI family from cattle, goat, and buffalo. These elements are approximately 500 bp, have a copy number of 2 x 10(5) - 4 x 10(5), and comprise about 4% of the haploid genome. Studies of nucleotide sequence homology indicate that the buffalo and goat PstI repeats (type II) are similar types of short interspersed nucleotide element (SINE) sequences, but the cattle PstI repeat (type I) is considerably more divergent. Additionally, the goat PstI sequence showed significant sequence homology with bovine serine tRNA, and is therefore likely derived from serine tRNA. Interestingly, Southern hybridization suggests that both types of SINEs (I and II) are present in all the species of Bovidae. Dendrogram analysis indicates that cattle PstI SINE is similar to bovine Alu-like SINEs. Goat and buffalo SINEs formed a separate cluster, suggesting that these two types of SINEs evolved separately in the genome of the Bovidae.     

Two-item sentence comprehension by a dog (Canis familiaris)

Syntax use by non-human animals remains a controversial issue. We present here evidence that a dog may respond to verbal requests composed of two independent terms, one referring to an object and the other to an action to be performed relative to the object. A female mongrel dog, Sofia, was initially trained to respond to action (point and fetch) and object (ball, key, stick, bottle and bear) terms which were then presented as simultaneous, combinatorial requests (e.g. ball fetch, stick point). Sofia successfully responded to object-action requests presented as single sentences, and was able to flexibly generalize her performance across different contexts. These results provide empirical evidence that dogs are able to extract the information contained in complex messages and to integrate it in directed performance, an ability which is shared with other linguistically trained animals and may represent a forerunner of syntactic functioning.     

On the allometry of long bones in dogs (Canis familiaris)

The allometric relations of diameter and length of humerus, ulna, femur, and tibia of 108 specimens, from 63 different breeds of dogs and 12 specimens of wolves, were calculated by means of model II of regression or major axis method. Only for the tibia were the values of wolves included in the cluster formed for dog breeds. Consequently, separate lines of regression were calculated for the other bones. Results agree in general with the exponents predicted by the theory of geometric similarity; however, the slope obtained for femur (0.865) differed significantly from this. Morphology of the long bones of the legs does not differentiate dogs and wolves; this probably reflects secondary convergence among wolves with relatively modern breeds of dogs.     

Characterization of swine short interspersed repetitive sequences

Swine genomic DNA segments containing repetitive sequences were isolated from a porcine genomic library using genomic DNA as a probe. Three fragments containing the repetitive sequences from two of the primary phage clones were subcloned for sequence analysis, which revealed six new PRE-1 repetitive families other than those reported earlier by Singer et al. (Nucleic Acids Research 15, 2780, 1987). The frequency of the repetitive sequences in the swine genome was estimated at 2 x 10(6) per diploid genome. Sequence analysis revealed similarities between these repetitive sequences and that of arginine-tRNA gene.     

The mechanics of jumping by a dog (Canis familiaris)

A force platform has been used to obtain records of the forces exerted on the ground by an Alsatian dog, during take-off for running long jumps and standing scale jumps. The records have been analysed in conjunction with cinematograph film, taken simultaneously, and anatomical data. Stresses in the principal muscles of the hind limb, and in the triceps, have been calculated and the values obtained are compared with the stresses found by other investigators in isometric experiments with excised mammal muscles. Stresses in certain tendons and bones have been calculated, and the values obtained are compared with published values for the strength of tendon and bone. Evidence is presented that the gastrocnemius and plantaris muscles behave, in take-off for a jump, essentially as passive elastic bodies. Most of the elastic energy is probably stored in their tendons. A tendency for distal limb muscles to be pinnate, with much shorter fibres than proximal limb muscles, is noted and discussed.