The spread of sequence variants in Rattus satellite DNAs

The genus Rattus has two related families of satellite DNA: Satellite I consists of tandem arrays of a 370 base pair repeat unit which is a dimer of two 185 base pair portions (a, b) which are about 60% homologous. Satellite I' consists of tandem arrays of a 185 base pair repeat unit (a') which is about 85% homologous to a and 60% homologous to b. R. norvegicus contains only satellite I but R. rattus contains both satellites I and I'. We examined certain aspects of satellite DNA evolution by comparing the spacing at which variant repeat units of each satellite have spread among non-variant repeat units in these two species. With but one exception, in R. rattus, 15 different variant repeat units have spread among non-variant repeat units of satellite I, with a spacing equal to the length of the (a,b) dimer. Similarly, fourteen different variant repeat units of the monomeric satellite I' have mixed among non-variant repeat units with a spacing equal to the length of the (a') monomer. These results suggest that a mechanism involving homologous interaction among satellite sequences could account for the spread of variant family members. We also found that a sequence variant present in certain portions of the dimeric repeat unit of satellite I is more efficiently amplified (or less efficiently corrected) than variants occurring in other regions. This was not true for the monomeric repeat unit of satellite I'.     

Different repeat lengths in rat satellite I DNA containing chromatin and bulk chromatin.

The nucleosome repeat structure of a rat liver chromatin component containing the satellite I DNA (repeat length 370 bp) was investigated. Digestion experiments with micrococcal nuclease, DNAase II, and the Ca2+/Mg2+-dependent endogenous nuclease of rat liver nuclei revealed a repeat unit of 185 nucleotide pairs which is shorter by approximately 10 bp than the repeat unit of the bulk chromatin of this cell type. The difference seems not to be related to the histone composition which was found to be similar in the two types of chromatin.     

Structure of 1.71 lb gm/cm(3) bovine satellite DNA: evolutionary relationship to satellite I

The Eco RI fragments from the 2600 bp repeating unit of 1.711b gm/Cm(3) bovine satellite DNA were cloned in pBR322. The structure of the repeat unit was determined and compared to bovine satellite I DNA (rho CsCl = 1.715 gm/cm(3)). All of the DNA in the 1402 bp repeat of satellite I is represented in the sequence of the 2600 bp 1.711b gm/cm(3) repeat. The difference between the two repeats is due to a 1200 bp piece of DNA (INS) residing in the middle of the 1.711b gm/cm(3) repeat. The INS is AT-rich and has some repetitive components; it bears only limited similarity to the structure of eukaryotic transposable elements. We propose that the 1.711b gm/cm(3) satellite DNA arose via the amplification of a 1.715 gm/cm(3) satellite repeat altered by a 1200 bp insertion of DNA.     

Ag-NOR chromosomal association patterns in Rattus rattus (Rafinesque, 1814) and Rattus norvegicus (Berkenhout, 1796) from the southern Iberian peninsula

A high frequency of satellite association phenomena in a number of specimens of Rattus rattus and R. norvegicus, captured on the coastal fringe of Málaga province (Southern Spain), was found. Only Ag-NOR bearing chromosomes participated in associations in both species. R. rattus is characterized by 14/18 heterologous and R. norvegicus by 5/5 and 13/13 homologous association types.     

Nucleotide sequence of a highly repetitive component of rat DNA.

A highly repetitive component of rat DNA which could not yet be enriched by density gradient centrifugation was isolated with the help of the restriction nuclease Sau3AI. This nuclease converted the bulk of the DNA to small fragments and left a repetitive DNA component as large fragments which were subsequently purified by gel filtration and electrophoresis. This DNA component which was termed rat satellite DNA I is composed of tandemly repeated 370 bp blocks. According to sequence analysis the 370 bp repeats consist of alternating 92 and 93 bp units with homologous but not identical sequences. Methylation of CpG residues was correlated to the rate of cleavage by restriction nucleases. Significant homologies exist between the sequences of rat satellite DNA I and satellite DNAs of several other organisms. The divergence of the sequence of rat satellite DNA I was discussed with respect to evolutionary considerations.     

Rat mitochondrial DNA polymorphism: sequence analysis of a hypervariable site for insertions/deletions

Three closely related variants of rat (Rattus norvegicus) mtDNA have been shown to differ in the number of T residues found in a run of Ts (light strand) which spans the junction between the tRNACys and tRNATyr genes. The number of Ts in the repeat varies from 6 to 8 in these DNAs. Another, less closely related, R. norvegicus variant has a run of 5 Ts at this site and in the related species, Rattus rattus, a run of 4 Ts is found. In R. norvegicus mtDNA runs of 5 As and 5 Gs are found just to the 3' side of the variable T repeat, and it is suggested that the three runs of repeated nucleotides may stabilize heteroduplexes which result from strand slippage and which give rise to the insertions and/or deletions. Among 17 mtDNA clones derived from an individual with the 8T repeat, one clone was found which possessed a 9T repeat. This variant may represent an additional DNA type originally present within the individual.     

Sequence analysis of bovine satellite I DNA (1.715 gm/cm3).

The 1402 bp Eco RI repeating unit of bovine satellite I DNA (rho CsCl = 1.715 gm/cm3) has been cloned in pBR322. The sequence of this cloned repeat has been determined and is greater than 97% homologous to the sequence reported for another clone of satellite I (48) and for uncloned satellite I DNA (49). The internal sequence structure of the Eco RI repeat contains imperfect direct and inverted repeats of a variety of lengths and frequencies. The most outstanding repeat structures center on the hexanucleotide CTCGAG which, at a stringency of greater than 80% sequence homology, occurs at 26 locations within the RI repeat. Two of these 6 bp units are found within the 31 bp consensus sequence of a repeating structure which spans the entire length of the 1402 bp repeat (49). The 31 bp consensus sequence contains an internal dodecanucleotide repeat, as do the consensus sequences of the repeat units determined for 3 other bovine satellite DNAs (rho CsCl = 1.706, 1.711a, 1.720 gm/cm3). Based on this evidence, we present a model for the evolutionary relationship between satellite I and the other bovine satellites.     

Nucleotide sequence and functional properties of DNA encoding incompatibility in the broad host-range plasmid R1162

Summary A 370 base pair (bp) fragment of R1162 DNA encoding the incompatibility determinant has been cloned and sequenced. The DNA is located between 6.1 and 6.5 on the R1162 map, near the origin of replication. The sequence contains three perfectly conserved 20 bp direct repeats, with 11 bp of this sequence repeated a fourth time. The direct repeat unit shows some homology with that of another, unrelated broad host-range plasmid, RK2. The cloned DNA has two other properties: it lowers the copy number of R1162 when cloned into this plasmid, and it is required in cis for replication of R1162 satellite plasmids.     

Non-random arrangement of nucleosomes in satellite I containing chromatin of rat liver.

The location of nucleosomes on the nucleotide sequence of rat satellite I DNA was investigated using micrococcal nuclease, exonuclease III, and restriction nucleases as tools. Hae III cleaved the satellite DNA containing chromatin very preferentially in the linker region. Nucleosomes were found predominantly in three defined positions on the 370 bp satellite I monomer unit. This type of arrangement occurs on not more than half of the satellite DNA containing chromatin while the rest of this chromatin is arranged differently. The arrangement of nucleosomes with high probability in preferred frames and with low probability in less preferred frames may be a general phenomenon which can be discussed as a possible mechanism to modulate sequence recognition.     

Insertion and amplification of a DNA sequence in satellite DNA of Cucumis sativus L. (cucumber)

Summary Another satellite DNA repeat (type IV) in the genome of Cucumis sativus (cucumber) was found and investigated with respect to DNA sequence, methylation, and evolution. This satellite shows a repeat length of 360 bp and a GC-content of 47%. The repeats of type IV are highly conserved among each other. Evidence for CG and CNG methylation is presented. By comparison to the previously described satellites (type I/II and type III) from cucumber, it is evident that this repeat is created by an insertion of a 180 bp DNA sequence similar to type I–III into another DNA sequence (or vice versa), and subsequent amplification forming a new satellite repeat. The different satellites of the type I/II, type III, and the 180 bp insert of type IV show a sequence homology of 60%–70%, indicating that the complex satellite DNA of cucumber is originated from a common progenitor by mutation, additional insertion, and amplification events. Copies of a sequence similar to a part of type IV are present in the genome of the related species Cucumis melo (melon).     

Characterization of X-chromosome specific satellite DNA of Muntiacus muntjak vaginalis

A highly repeated DNA (designated satellite IA) was isolated from cultured cells of Muntiacus muntjak vaginalis and its organization analyzed by the use of restriction nucleases and hybridization experiments with cloned DNA-fragments. Several restriction nucleases cleave the satellite IA DNA into a series of fragments, which are multiples of a basic repeat unit of 800 bp. Sequences homologous to the satellite IA DNA were also found in a second highly repetitive DNA component of Muntiacus muntjak vaginalis (satellite IB). Its organization is more complex than the one of satellite IA and does not conform to a simple periodicity of a basic repeat unit. — Hybridization in situ revealed, that both satellites are confined in their entirety to the X-chromosome, where they are located at both arms close to the centromere. No satellite DNA was found at the Y1-chromosome, which is considered to be homologous to the long arm of the X-chromosome. These results have interesting implications for the evolution of the X-chromosome.     

Concerted evolution of primate alpha satellite DNA. Evidence for an ancestral sequence shared by gorilla and human X chromosome alpha satellite

To understand evolutionary events in the formation of higher-order repeat units in alpha satellite DNA, we have examined gorilla sequences homologous to human X chromosome alpha satellite. In humans, alpha satellite on the X chromosome is organized as a tandemly repeated, 2.0 x 10(3) base-pairs (bp) higher-order repeat unit, operationally defined by the restriction enzyme BamHI. Each higher-order repeat unit is composed of 12 tandem approximately 171 base-pair monomer units that have been classified into five distinct sequence homology groups. BamHI-digested gorilla genomic DNA hybridized with the cloned human 2 x 10(3) bp X alpha satellite repeat reveals three bands of sizes approximately 3.2 x 10(3), 2.7 x 10(3) and 2 x 10(3) bp. Multiple copies of all three repeat lengths have been isolated and mapped to the centromeric region of the gorilla X chromosome by fluorescence in situ hybridization. Long-range restriction mapping using pulsed-field gel electrophoresis shows that the 2.7 x 10(3) and 3.2 x 10(3) bp repeat arrays exist as separate but likely neighboring arrays on the gorilla X, each ranging in size from approximately 200 x 10(3) to 500 x 10(3) bp, considerably smaller than the approximately 2000 x 10(3) to 4000 x 10(3) bp array found on human X chromosomes. Nucleotide sequence analysis has revealed that monomers within all three gorilla repeat units can be classified into the same five sequence homology groups as monomers located within the higher-order repeat unit on the human X chromosome, suggesting that the formation of the five distinct monomer types predates the divergence of the lineages of contemporary humans and gorillas. The order of 12 monomers within the 2 x 10(3) and 2.7 x 10(3) bp repeat units from the gorilla X chromosome is identical with that of the 2 x 10(3) bp repeat unit from the human X chromosome, suggesting an ancestral linear arrangement and supporting hypotheses about events largely restricted to single chromosome types in the formation of alpha satellite higher-order repeat units.     

Recent amplification of an alpha satellite DNA in humans.

A repeat sequence 682 base pairs (bp) long produced by cleavage of human DNA with Xba I restriction enzyme is composed of four tandemly arranged subunits with lengths of 171, 170, 171, and 170 bp each. The sequence organization of the 682 bp Xba I repeat bears a striking resemblance to other complex satellite DNAs of primates, including the Eco RI human alpha satellite family which also occurs as a 170 bp repeat. The Eco RI tetramer and the 682 bp Xba I repeat show a sequence divergence of 21%. The 682 bp Xba I repeat sequence is restricted to humans and is only distantly related to the previously reported 340 bp Xba human repeated DNA sequence. These finding are consistent with the concept of occasional amplifications of members or groups of members of alpha satellite DNA during human evolution. Amplifications apparently occurred after humans, apes and gibbons diverged from Old World monkeys (Eco RI satellite), after humans and apes diverged from gibbons (340 bp Xba I satellite) and after humans diverged from the great apes (682 bp Xba I satellite).     

Conservation and antigenicity of N-terminal sequences of GP185 from different Plasmodium falciparum isolates

Complementary DNA (cDNA) clones for GP185, a major antigenically diverse glycoprotein of Plasmodium falciparum, were isolated from a cDNA library of the Honduras I/CDC (Honduras I) isolate, and 1052 bp were sequenced. The expression of cDNA fragments in Escherichia coli using the vector pCQV2 allowed verification of the reading frame. This GP185 cDNA sequence, like the cDNA sequence for a homologous gene of the K1 isolate [Hall et al., Nature 311 (1984) 379-382], codes for a polypeptide which is truncated due to multiple, in-frame stop codons. This polypeptide corresponds to the N-terminal 15% of the proposed coding region of the GP185 gene [Holder et al., Nature 317 (1985) 270-273]. Comparison of the nucleotide sequences for the GP185 gene of Honduras I and five other isolates indicated that there are two areas of conserved DNA sequence, one of 310 bp (beginning 181 bp upstream from the proposed initiation codon) and the other of greater than or equal to 360 bp (located entirely within the coding region), separated by a region encoding isolate-specific tandem amino acid repeats. Rat antiserum was raised to a fusion protein derived from the conserved regions and the intervening repeat region of this Honduras I protein. This antiserum bound GP185 on immunoblots of the homologous Honduras I isolate and the heterologous K1 isolate, which has different tandem repeats. Serum from owl monkeys and humans previously infected with P. falciparum reacted with the fusion protein on immunoblots demonstrating that determinants in the N-terminal 15% of GP185 were immunogenic in infected individuals and suggesting that some of these sites are conserved among isolates.(ABSTRACT TRUNCATED AT 250 WORDS)     

The 1360 bp long basic repeat unit of calf satellite I DNA contains GC rich nucleus of about 140 bp.

Fine melting profiles of calf satellite I DNA and its fragments obtained after digestion with endoR.EcoRI and endoR.AluI nucleases were investigated. It is shown that the 1360 bp basic repeat unit of calf satellite I DNA contains an about 140 bp long GC rich nucleus. It is localized on the 600 bp restriction fragment obtained after digestion of 1360 bp fragment with endoR.AluI nuclease. The main part of satellite I DNA melts as loops between such GC rich nuclei which strongly influence the melting properties of this satellite. There exist significant differences between the thermal stabilities of fragments containing many nuclei, one nucleus and those in which such nucleus is absent.     

Seasonal studies on commensal rats and their ectoparasites in a rural area of Egypt: the relationship of ectoparasites to the species, locality, and relative abundance of the host.

The present study was carried out in 3 villages, namely Kafr Ayoub Soliman, Kafr Ibrahim El-Aidi, and El-Sa'adat, Sharqiya Governorate, Egypt. A total of 519 rats was collected from the 3 study sites: 46.6% Rattus rattus, and 53.4% Rattus norvegicus. A total of 20,643 ectoparasites was recovered from R. rattus: 33.3% mites, 33.8% fleas, and 32.9% lice. From R. norvegicus a total of 40,997 ectoparasites was recovered: 28.9% mites, 31% fleas, and 40.1% lice. Three common mite species were recovered from both rat hosts, i.e., Ornithonyssus bacoti, Radfordia ensifera, and Laelaps nuttalli. Three common flea species were also recovered from both rat hosts, i.e., Echidnophaga gallinacea, Leptopsylla segnis, and Xenopsylla cheopis. Polyplax spinulosa was the only dominant louse species that infested both rat hosts. Rats did not show a definite breeding season, and the seasonal rat indices were generally low in different study sites. There were no significant differences between the prevalence of each of mites, fleas, and lice in both rat species. The total general indices of mites and fleas, on the other hand, was significantly higher in R. norvegicus. The general index of X. cheopis was high and ranged between 5.9 in R. rattus and 14.5 in R. norvegicus. Season-related changes were observed in the general index of each of L. segnis infesting both rat species and R. ensifera and O. bacoti infesting R. norvegicus. The prevalence and general indices of some ectoparasites showed differences related to the locality of their rat hosts. Seasonal changes in the general indices of some ectoparasites paralleled seasonal changes in the relative abundance of their rat hosts.     

The organisation of the long range periodicity calf satellite DNA I variants as revealed by restriction enzyme analysis.

The analysis of a large number of restriction sites within the long range periodicity calf satellite DNA I does not reveal a superimposable shorter repeat. Although some restriction sites are present in almost all the 100,000 tandemly arranged copies of the 1460 bp repetition unit, other sites such as Atu CI occur at much lower frequencies. When present they are distributed randomly along the satellite DNA molecules. The missing sites appear to result from random and presumably single base alterations. Digestion with the enzymes Hha I and Kpn I showed another type of variant to exist within the calf satellite DNA I. Unlike Atu CI the distributions of the variants detected by these enzymes are not random and organised on long stretches of satellite DNA. The possible functional significance and evolutionary implication of these results are discussed.     

DNA-protein interactions at the telomeric repeats of Schizosaccharomyces pombe.

Gel retardation assays using a probe containing the repeat region of a Schizosaccharomyces pombe chromosomal telomere identified four specific DNA- protein complexes in S. pombe total protein extracts (I, I', IIa and IIb). The proteins responsible for these complexes bound to the telomeric repeat region irrespective of whether or not the repeats were in close proximity to the end of a DNA molecule, and none of them bound strongly to single-stranded DNA. The protein responsible for complex I (TeRF I) was separated from the activity responsible for complexes IIa and IIb (TeRF II) using heparin-Sepharose chromatography. Both factors were efficiently cross-competed by an oligonucleotide containing the 18 bp sequence 5'-GGTTACAGGTTACAGGTT-3', which corresponds to two complete telomeric repeat units. Mutation of the T residues at positions 4 and 11 in the oligonucleotide dramatically reduced binding by TeRF II, but had no affect on binding by TeRF I. The protein responsible for complex I' did not bind strongly to either the wild-type or mutant oligonucleotide.     

Characterization and chromosomal distribution of satellite DNA sequences of the water buffalo (Bubalus bubalis).

Satellite DNA sequences were isolated from the water buffalo (Bubalus bubalis) after digestion with two restriction endonucleases, BamHI and StuI. These satellite DNAs of the water buffalo were classified into two types by sequence analysis: one had an approximately 1,400 bp tandem repeat unit with 79% similarity to the bovine satellite I DNA; the other had an approximately 700 bp tandem repeat unit with 81% similarity to the bovine satellite II DNA. The chromosomal distribution of the satellite DNAs were examined in the river-type and the swamp-type buffaloes with direct R-banding fluorescence in situ hybridization. Both the buffalo satellite DNAs were localized to the centromeric regions of all chromosomes in the two types of buffaloes. The hybridization signals with the buffalo satellite I DNA on the acrocentric autosomes and X chromosome were much stronger than that on the biarmed autosomes and Y chromosome, which corresponded to the distribution of C-band-positive centromeric heterochromatin. This centromere-specific satellite DNA also existed in the interstitial region of the long arm of chromosome 1 of the swamp-type buffalo, which was the junction of the telomere-centromere tandem fusion that divided the karyotype in the two types of buffaloes. The intensity of the hybridization signals with buffalo satellite II DNA was almost the same over all the chromosomes, including the Y chromosome, and no additional hybridization signal was found in noncentromeric sites.     

Dating of divergences within the Rattus genus phylogeny using whole mitochondrial genomes

The timing and order of divergences within the genus Rattus have, to date, been quite speculative. In order to address these important issues we sequenced six new whole mitochondrial genomes from wild-caught specimens from four species, Rattus exulans, Rattus praetor, Rattus rattus and Rattus tanezumi. The only rat whole mitochondrial genomes available previously were all from Rattus norvegicus specimens. Our phylogenetic and dating analyses place the deepest divergence within Rattus at approximately 3.5 million years ago (Mya). This divergence separates the New Guinean endemic R. praetor lineage from the Asian lineages. Within the Asian/Island Southeast Asian clade R. norvegicus diverged earliest at approximately 2.9Mya. R. exulans and the ancestor of the sister species R. rattus and R. tanezumi subsequently diverged at approximately 2.2Mya, with R. rattus and R. tanezumi separating as recently as approximately 0.4Mya. Our results give both a better resolved species divergence order and diversification dates within Rattus than previous studies.